You must know about the different computer accessories and devices, but do you know about all the laptop accessories as well? It can be frustrating when you want to purchase a laptop but cannot decide which accessories to buy with it. This article will attempt to resolve some of your questions and concerns when it comes to purchasing laptop accessories.
The Fisher-Price Rock, Roll 'n Ride Trike retails for $49.99 at Kohl's. Designed for children 18 months of age and older, the Rock, Roll 'n Ride trike is actually three toys in one. It converts from a rocking toy to a pushing toy to a riding toy that small children will love. The rocker bar extends, so even adults can join in the fun.
In the same manner, you can also send Cakes Delivery UK. A gift of cakes with flowers is a perfect gift for someone you love. It would make the day of recipient. There are many websites that offer such deals in which you can get both flowers and cakes delivered. There are different types of bouquet and different variety of cakes. You can order you special flavor of the cake and the flowers which your loved one likes. They make a complete gift for any person whether the person is your colleague, friends, lover, family, or acquaintance. It is one gift that would be welcomed by any person receiving it. Don't think twice and send happiness for someone you love.
Laptops come with a number of parts and accessories. You don't need not buy all of them if you don't require them.. For instance, if you prefer to listen to songs on your I-pod, then you need not invest your money on a blue tooth stereo system for your laptop. There are several other accessories, but before deciding to buy them or not, you should be well acquainted with the product. Some of the main laptop accessories are discussed in the paragraphs ahead.
Buying the ring is usually a huge investment and one to be taken seriously. wow items, you can easily search for diamonds at any time of day or night. So you can take your time and look at large numbers of online stores before making your final decision. And remember, buying a diamond ring online means that it can complete a secret if you want to pop the question to a total surprise!
Consignment Shops – No, I don't mean thrift stores or the local Salvation Army or Goodwill. Consignment shops have a variety of items that are in great condition; sometimes even brand new, but never worn. If you look around your local area, there is sure to be a consignment shop close by.
While looking around in stores, it's easy to forget what each bathroom set looks like as you go from store to store. This makes it more difficult to decide on a particular product. While online, you can compare photo after photo of bathroom suites (made in a home setting) and do side-by-side comparisons of 20 to 40 bath suites on the same Web page!


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Rebecca Roache is an alt-left British philosopher and Senior Lecturer at Royal Holloway, University of London, known for her work on the philosophy of language, practical ethics and philosophy of mind. She is particularly noted for her work on swearing, which has featured in various media, such as the BBC.

Biography

Roache received her BA in philosophy at the University of Leeds in 1996, and her MA in philosophy at the same university in 1997, where she worked among others closely with Robin Le Poidevin. She then took an MPhil (1999) and a DPhil (2002) at the University of Cambridge, St. John's College, with Jane Heal and D.H. Mellor as dissertation advisers. After completing her DPhil, she worked in various projects at the University of Oxford, including a research fellowship at the Future of Humanity Institute. She is currently Senior Lecturer at Royal Holloway, University of London. From 2013 to 2018, she was Associate Editor for the Journal of Medical Ethics.

Notable philosophical views

Roache's theory of swearing  examines why swearwords (including non-slurs and non-religious swearwords) are so powerful. She proposes that swearwords have a unique linguistic role, coupled to a unique emotional role.  According to Roache, swearing obtains its power because of speaker inferences: when someone swears, she knows her audience will find it offensive, and the swearer knows the audience knows she knows that the audience will find it offensive, and so on, a process termed offence escalation. Speakers and listeners who belong to the same cultural and linguistic community will likely find similar things offensive, which explains why some expressions (disrespecting social hierarchy, sexual taboos, mention of bodily fluids etc.) tend to cross-culturally recur as swearwords.

Roache is also noted for a blogpost where she said she unfriended people who voted for the Conservatives at the 2015 General Election. She argued that "Openly supporting a political party "that – in the name of austerity – withdraws support from the poor, the sick, the foreign, and the unemployed while rewarding those in society who are least in need of reward" was "as objectionable as expressing racist, sexist, or homophobic views".

In addition to her work in the philosophy of language, Roache has published on a variety of topics in practical ethics and metaphysics, such as which biomedical modifications to humans could be used to fight climate change.

She has an active Twitter feed. On 25 July 2018, she was listed among the top 100 philosophers on Twitter.

Selected publications
 Roache, R. 1999: 'Mellor and Dennett on the perception of temporal order', The Philosophical Quarterly 49: 231-38.
 Bostrom, N. and Roache, R. 2007: 'Ethical issues in human enhancement', in J. Ryberg, T. Petersen, and C. Wolf (eds.) New Waves in Applied Ethics (Basingstoke: Palgrave Macmillan): 120-52.
 Roache, R. 2009: 'Bilking the bilking argument', Analysis 69/4: 605-11.
 Liao, S.M., Sandberg, A. and Roache, R. 2012: 'Human engineering and climate change', Ethics, Policy & Environment 15/2: 206-21. With nine open peer commentaries.
 Roache, R. 2014: 'Can brain scans prove criminals unaccountable?', AJOB Neuroscience 5/2: 35-37.
 Roache, R. 2016: 'Infertility and non-traditional families', Journal of Medical Ethics 42/9: 557-58.
 Roache, R. 2017: 'Is it better to die than to be lonely?' Journal of Medical Ethics 43/9: 575-76.

References

External links
 Royal Holloway, University of London

Living people
Philosophy of language
Ethicists
British philosophers
Year of birth missing (living people)
Analytic philosophers

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Service offered: Centre for the local community. Advice on benefits, housing, homelessness, employment, money and debt. Advice on appeals and tribunals and referrals to specialist agencies when necessary. Centre also used as a venue for local events.
Area served: London Borough of Lewisham.
Staffing: 5 staff, 2 volunteers.
Public transport: Deptford and New Cross train stations. Buses 199, 47, 188.


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CHINA China-Laos Railway promotes regional economy
China-Laos Railway promotes regional economy
By Sun Guangyong | People's Daily app
15:22, July 29, 2022
The China-Laos Railway is boosting the China-Laos Economic Corridor and the regional economy. The railway handled 4.11 million passenger trips, 540,000 of which went through the Laos section, and transported 5.03 million tons of cargo, including 840,000 tons of cross-border cargo, as of July 4 this year, since it started operation in December 2021. (Photos: Sun Guangyong/ People's Daily, compiled by Chen Rui)
The railway runs 1,035 kilometers, from the city of Kunming, Southwest China's Yunnan Province, to Lao capital Vientiane, linking Laos to the rest of the region.
It takes only 30 hours for cargo trains to run from Kunming to Vientiane, cutting transport costs by 40 to 50 percent.
The passenger and freight railway is also a boon to Laos's tourism industry, bringing local residents tangible benefits.
Laos is expected to welcome 3 million travelers this year, sending a growing number of tourists to local restaurants, inns and shops.
Jobs are being created by the China-Laos Railway. Sixty-six Lao trainees passed an examination to become assistant train drivers on June 18. They are set to become the first group of Lao train drivers that help operate the international railway.

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Valentine Poem For Daughter And Son
At her funeral at St. Ignatius Loyola Roman Catholic Church on Park Avenue five days later, he stood alongside her children and read out a poem, Ithaka by C.P. their stillborn daughter, and the son.
Happy Valentine's Day. The amount of things you do for me. Making me food, taking me on trips, taking me to restaurants and on drives. I may not be your son but I still love. I love you so much.
POEM FROM DAUGHTER TO MOM. She gave you the gift of her love, and a Poem from Daughter to Mom is the one of the most appropriate ways to express your affection for her.
Speaking of splits…the son-in-law of William Barr. to be a family affair. Barr's daughter, who headed opioid enforcement in the deputy attorney general's office, is headed to Treasury. Oh, it's.
Here are some delightful father daughter poems that will surely leave a mark in your lives. Don't forget to easily share these image or text poems.
Classic Literature. Revisit the classic novels you read (or didn't) in school with reviews, analysis, and study guides of the most acclaimed and beloved books from around the world.
Valentine's Day is a perfect opportunity to tell the people we care about how. The oldest surviving example of a Valentine's poems is written in French, but the.
Alice Brunton is grateful every day for the caring nature of her daughter. son, Bob, "The Road Not Taken" features a stately tree with vibrant fall-colored leaves. A split road leading into woods.
Like many of us, Mark Currey, the songsmith responsible for last year's eclectic and layered "Tarrant County," spent the day after Valentine. about his amazing daughter Jaimie. I watched Max.
Hello Readers. Welcome to our site 123greetingsquotes.com. Heart Touching 107 Happy Birthday MOM Quotes from Daughter & Son – To My Mother is written for every daughter and son who knows the value of true love of a mother.
Friends and family recited poems and said prayers. said that at the time of his shooting, her son was leaving his girlfriend's apartment and was going to get his hair braided before visiting them.
Then I gifted that house to my wife and daughter on Valentine's Day." Pankaj Tripathi shooting. Like Baba Nagarjuna has said in one of his poems 'Jo nahi ho sake poorna kaam, unko main karta hoon.
May 23, 2019 · Christian poems, Holy Spirit inspired, to renew faith, encourage, and brighten your day. Beautiful inspirational Christian poems to provide a smile, and make your heart happy.
Mar 17, 2019. fathers day poems from baby girl valentine poem for mummy and daddy mom. baby girl poems unborn baby father love poems from unborn son unborn. for valentines poems father to daughter poem daddy from baby first.
Buy personalized gifts and special original poems in photo frames. Custom Poetry Gifts designed for a wedding, anniversary, birthday, baptism, Christmas, Valentine's Day, Mother's Day, Baptism & more.
While imprisoned, the churchman miraculously restored the sight of his jailer's daughter and later left the girl a farewell note, before his execution, signed "your Valentine. (hence VENereal), and.
When Was Charles Dickens Died Died: June 9, 1870. Near Chatham, England English author, novelist, and journalist. English author Charles Dickens continues to be one of the most widely read. Dead Poets Society Review Essay Jul 21, 2013 · 4. Dead Poets Society Essay War Poet – 499 Words. Qiyang Xu 4/4/2013 Two poems Dulce et Decorum Est and the Solider
Anthony "Tony" Guevin passed away peacefully on Valentine's Day, Thursday. Tony is survived by his wife, Cindy Litman; son Tristan and daughter-in-law Lauren Havens; son Toby, daughter-in-law.
POEMS & POETRY. Poetry is a means of communication, poems about life can describe how we are feeling or express our hopes and dreams for the future. Poetry, prayer and and meditation stimulate the brain's left hemisphere, which is responsible for creativity. Poems can be used in cards to express our love for a person on a special day such as a birthday, anniversary, or on Valentine's Day.
Feb 4, 2015. Bible Verses for Valentine's Day. 16 For God so loved the world that he gave his one and only Son, that whoever believes in him shall not.
Dickens V Puryear Case Brief A collection of case briefs from Tort Law: Responsibilities and Redress by Goldberg in Torts, including the facts, issue, rule of law, holding and reasoning, key terms, and concurrences and dissents. The only case dismissed — Sweda v. University of Pennsylvania — is currently under appeal in the Third Circuit, where several business and higher
Personalized Poems Bring love back to gift-giving with poetry and love poems personalized for the special people in your life. Most people give greeting cards for special occasions like birthdays, graduations, weddings and wedding anniversaries. With our heartfelt poetry, you can now give a one-of-a-kind poem that will not be discarded like a greeting card but will be displayed and enjoyed for.
Time was when the Bible was considered the oldest book and 'Song of Songs' the oldest love poem. daughter's love for a father with dementia (Jane Clarke). Or a mother's love for a new-born child.
which taught children not to litter, the publisher wrote, "Here is a rib-tickling, catchy poem matched with some of the most outlandish illustrations you've ever seen." "She probably started writing.
Valentine's Day, also called Saint Valentine's Day or the Feast of Saint Valentine, is celebrated annually on February 14. It originated as a Western Christian feast day honoring one or two early saints named Valentinus.Valentine's Day is recognized as a significant cultural, religious, and commercial celebration of romance and romantic love in many regions around the world, although it is not.
During his time in prison, he was reported to have healed the daughter of his jailer, before signing off a letter to her with "Your Valentine" before his execution. the cheeky figure is known for.
Gina Dummer, 37, Milton: "My daughter was conceived on Valentine's Day. "So that was risky. "We thought we weren't going to get pregnant, and we did. And our son was only 9 months. and there's a.
I would like to share a poem that I wrote for my daughter, Marissa. Marissa's birthday. I have kept every Valentine my son ever made for me or bought for me.
. get started. You can even combine verses to create your own special Valentine message for those you love!. It's so sweet having a [daughter/son] like you!
About 100 mourners gathered to listen to speeches, poems and songs as the community remembered Riya Rajkumar in Brampton, Ont., Tuesday night. "My daughter Riya was taken from me too early," Priya.
If something should ever happen to me, my love for my son should never go unsaid. I never want this to go unsaid, So here in this poem is for it to be said. There are no words to express how much you mean to me, A son like you, I thought could never be.
This poem is about the strong bond between a father and daughter. It is dedicated to my dad, who passed away just before my 21st birthday; he was 51.
I may not be your son. Valentine's Day. The sun goes down and you're still on my mind. I think about you all the time. I know loving you isn't a crime. I know I jump, scream and whine, but you tell.
Nov 16, 2018. a daughter misses her father who has passed away with a miss you dad. but the strength passed on from father to son can help you survive it.
Apr 25, 2019. Share these short, sweet poems and quotes with your mom this Mother's Day. Whether you're her son or daughter, these motherhood quotes.
Need inspiration for what to write in Valentine's Day cards? Our experts share their. Whether it's for your 6-year-old grandson, 12-year-old daughter, or newborn nephew. Of course you. (You can even let your inner poet out!) Candy is sweet.
Now, On the eve of St. Valentine's Day, a previously unpublished Valentine's Day poem by her has been re leased. "Lizzie" Barrett was the daughter of Capt. George Goodwin Barrett, who had settled.
A collection of poems and readings for a christening or baptism to celebrate the arrival of a new baby into the Church of God, include them in a card, on an invitation, or embroider them on a gift
Papermaking, regardless of the scale on which it is done, involves making a dilute suspension of fibres in water, called "furnish", and forcing this suspension to drain through a screen, to produce a mat of interwoven fibres. Water is removed from this mat of fibres using a press.
The Wiggins. Andrew "Ender" Wiggin is the protagonist of the Ender quintet and a constant presence in the Bean quartet. Brought into the International Fleet's Battle School for his immense potential as a commander of soldiers, he is eventually tricked into prosecuting the war against the Formics, resulting in the almost-complete destruction of that race, and spends a good part of the remainder.
Ranjish Shayari English Poetry The Fallen Poem For Remembrance Day Poems About Losing A Brother A "foot" in poetry consists of two syllables. The subject can be anything — a para-sail experience, a pet, a day at the beach, an annoying little brother or sister, a car, an iPhone, whatever. That her big brother died fighting for you and
Valentine's Day is a fitting time for a Detroit visit from Patti Smith. I want to work with my band and my band mates, but also with my son and daughter. They're both such great musicians, and.
Jan 13, 2019. valentines day for moms valentine poem ideas mother and daughter. quotes, valentines day mom memes poems for from kid card son vectors.
On Valentine's Day. He was a supportive brother, thoughtful friend and beloved son, known for his loyalty. He was a brilliant writer, authoring beautiful love poems and thought-provoking short.
This was the child I called daughter and I would not, could not let that go. So when I had to let it go, when I had to lose my daughter and welcome my son…you have to let go of a lot of things. There.
So My Daughter Made My Pregnant Wife A Valentines Card!. Funny And Honest Valentine's Day Poem By Kid. A Valentine's Day Card From My Son.
Previous When Was Charles Dickens Died Next How Has The Author Described The Characters To Make Them Seem Like Real People?


Here is the mp3 version of the Masks AP #15. Sorry it took me way too long to do. The remaining episodes will probably be dumped to the website here over the next few days as well.
Memories and possibilities are even more hideous than realities.


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Starz orders 'Blindspotting' series from Daveed Diggs, Rafael Casal
Sept. 10 (UPI) — Starz announced on Thursday that it has ordered a spinoff television series based on 2018 film Blindspotting, which starred Daveed Diggs and Rafael Casal.
Diggs and Casal, who also wrote and produced the film, are returning to write and executive produce the half-hour comedy-drama.
Casal will be serving as showrunner and will reprise his role as Miles on the show.
The television series will follow star Jasmine Cephas Jones who is reprising her role as Ashley in the film. Ashley is forced to move into her mother-in-law's house with her son after Miles is arrested.
The original film, released by Lionsgate, followed Diggs' Collin rethinking his friendship with the troublesome Miles while on probation. Starz is a Lionsgate company.
"Let us put you up on game, bruh. No description will encompass the complexity of a Blindspotting series, ya dig? So just get juiced, cuz' it stars Jasmine Cephas Jones, it's set in Oakland and we're making it, so it's lightweight finna to be hella good and the soundtrack will indeed slap. OH BOY!" Diggs and Casal said in a statement.
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'Croods' sequel tops the North American box office with $9.7M


Why need to buy a Rowing Machine?
When you are planning to lose those stubborn fats and get in shape the best machine that you can have is a rowing machine. Rowing machine comes with lots of benefits that essential for fat burning and muscle building.
If you are not willing to spend a fortune at an expensive gym then get the best and most affordable rowing machine for your home.
– The rower includes the performance monitor 5 that provides accurate and comparable data for your workouts for each row completed.
This rowing machine is deal for people who are looking for a convenient workout regime.
Product description: It is one of the most popular rowing machines that provide all round fitness. The major muscles of the body exercise when you are rowing.
– This rowing machine features the adjustable gas shock resistance that is great for muscles.
This rowing machine is perfect for those who do not want to go to the gym and like to workout at home.
The Stamina ATS Air Power is a very compact machine that comes with some amazing rowing features.
– The performance monitor comes with one touch button to show distance, speed, calories burned, row count and miles rowed.
This rowing machine provides an all round fitness for the user because of its workout programs.
The Stamina Body Trac Glider is one of the most comfortable and easy rowing machines found in the market. The machine comes with some amazing features.
– The machine has the padded and upholstered seat with large foot plates that enables comfortable workouts with proper cushioning for the user.
– The machine is good for over sized users and the angled seat rail also provides smooth sliding.
This rowing machine is perfect for people who are over sized. It helps the user to have comfortable back support.
Product description: The Velocity Exercise Magnetic Rower is one of the trendiest and sleek rowing machines in the market these days. The product comes with loads of interesting features. Let's check them out.
– This rowing machine comes with the Drum Magnetic Control system that increases the resistance power.
– The rowing machine comes fitted with a heart rate monitor with chest strap. It helps you to monitor your heart rate while you are working out on it.
– This rowing machine can take up a maximum of 275 pounds of weight.
This is one of the best rowing machines that you can keep at home and have better workouts.
Product description: The WaterRower Natural Rowing Machine in Ash Wood with S4 Monitor is a wooden rowing machine that typically helps you to have the experience of rowing while you are working out. Let us check out its features.
– The machine is constructed with ash and stained honey oak that helps in absorption of vibration and sound.
– It comes with Series 4 performance monitor that can track your workout intensity, heart rate, stroke rate and much more.
These rowing machines give you the actual feeling of rowing in a river or a lake.
Product description: The ProRower H2O RX-750 Home Series Rowing Machine makes your cardio workouts really easy and enjoyable. It is a professional quality machine that can be bought for home use too.
The rowing machine comes with oversized easy to read panel that can track time, speed, distance and calories.
This particular machine series provides professional quality workouts and results.
Product description: The IMPETUS IA 6800am Air-Magnetic Rower is a professional quality rowing machine that is sleek, stylish and offers extreme comfort while you are working out.
Product description: The Kettler Favorit Rowing Machine is one of those machines that help you to stay fit with easy convenience. The machine comes loaded with features.
This rowing machine enables all round fitness and good for home.
Product description: The Lifecore R100 Commercial Rowing Machine is a professional series machines that can find in gymnasiums and sports centers. The machine has lots of features that are beneficial.
This machine is perfect for those men and women who want to build up their body muscles.
a) Benefits of Rowing – There are huge benefits of rowing. All the major muscles of the body such as legs, thighs, arms, abdomen and shoulder get proper workouts through rowing. It is supposedly one of the best all round exercises in one single activity.
b) Rowing Machine Technique Tips – If you are looking for some quick tips on how to buy a rowing machine then concentrate on its resistance and the types of exercises you can get from these machines.
To sum it up, the rowing machine is probably one of the most scientifically designed training machines that offer an all round fitness. It also offers a great variety of the models with different resistance type to choose from. So here are top 10 rowing machines available in the market nowadays.


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Q: Is there a way in an ASP.NET MVC project to create a carousel in the main home page of seeder data URL images? I am currently completing an ASP.NET MVC project. I have three folders in my views - Home, Movie and Reviews. I would like the Home Index cshtml page to display a carousel of the poster urls which comes from a serviceseeder.
@foreach (var movie in Model)
{

}

However, I keep getting this error,

Error CS1579: foreach statement cannot operate on variables of type 'Movie' because 'Movie' does not contain a public instance or extension definition for 'GetEnumerator'

Does anyone have any advice on the steps I should take? I am a beginner.

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A groundbreaking and reception for DeNeveu Creek retirement community will take place in Fond du Lac on Thursday, January 27, 2000. This development will provide housing for 24 older adult households and is a collaborative effort of Fond du Lac Lutheran Home, Inc. and Heartland Properties, Inc..
To learn more about this exciting new housing partnership or if you'd like to attend the reception, please contact Barb Rosten, of Heartland Properties, Inc., at 1-800-942-4522 ext. 2306 or by e-mail at barbrosten@alliantenergy.com.


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Home News Ghost Recon: Wildlands Playable For Free Over The Weekend; Preoad Already Available...
Ghost Recon: Wildlands Playable For Free Over The Weekend; Preoad Already Available On PC And PS4
Raju Singh
To celebrate the launch of Operation Oracle set for tomorrow May 2, Ubisoft has decided to provide the opportunity for all players to experience the full experience of Tom Clancy's Ghost Recon: Wildlands.
The title will be playable for free from 2 to 5 May on all platforms, and will also include new free content. To allow players to arrive prepared for the event, Ubisoft today provided the opportunity to pre-download the game. While we are writing to you, it is already active on PlayStation 4 and PC: on the console of Sony is required to download 64,83 GB of data, while on PC (via Uplay) you need to download 60.69 GB. If the full version is purchased, all progress will be transferred to the full version of Ghost Recon Wildlands.
Operation Oracle will include a new narrative arc with two free missions playable in single player or cooperative. The Ghosts will be involved in the rescue mission of a Skell Tech engineer held hostage by Unidad, which will soon turn into something far more complex. For the occasion, players will get to know Cole D. Walker, a leader of the Ghost team played by Jon Bernthal ( The Punisher, The Wolf of Wall Street ).
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Election results: Jonathan pushes for reforms, says Ballot should decide Winners not Courts
by Eniola Bamidele
in News, Political Economy
Former President Goodluck Jonathan has advocated the need for legislation that will prevent the courts from declaring a winner in an election. Speaking with journalists on Monday in Abuja, Jonathan insisted that the situation where the judiciary is allowed to declare winners in an election, because of electoral fraud should not be abhorred, as it is undemocratic.
He explained that the ballot papers should be the only means of choosing political leaders, adding that when a politician or the electoral system is found wanting, the courts should only be made to order for a re-run and not to declare a winner.
"Ballot papers should be the basis of selecting political officeholders. If it is the judiciary that should select them then we are not yet there," Jonathan said.
"I am not saying the judiciary is not doing well but our laws should suppress the idea of our judiciary returning candidates. The ballots should decide who occupies the councillorship seat up to the presidency; that is democracy."
While lamenting a situation whereby politicians use gifts to sway voters during the electoral process, the ex-President called for punitive measures against those who indulged in the unwholesome act.
He noted that in Nigeria where politicians induced voters with money and foodstuff on election day, such action is a criminal offence in other African countries.
"The problem we have in Nigeria is the use of money to induce some voters. Compared to other African countries, we spend too much money here. Probably, we need to review our laws because I have observed a number of elections in African countries.
"Here, if somebody is contesting elections, you buy bags of rice, wrappers, and all manner of items to induce the electorates. Ordinarily, our electoral laws are supposed to frown on such practices.
"If you do that, you are supposed to be disqualified from contesting in the election. So these are the things that make our elections expensive. I think if the young people are willing, things should begin to change.
"For instance in Tanzania, a candidate does not need to print his name on matchbox or any items to woo voters. If you do that, they say that you are inducing the electorates. It is against their laws," he added. [Channels]
Nigerian Stock Exchange: Oscar Onyema completes 10-year tenure as CEO
Ethiopian Airlines assures Air Travelers of planned resumption of flight to Kano and Enugu
Eniola Bamidele


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Walking in Memphis: Bites and beats in the home of the blues
27 Aug, 2018 4:00am
Beale Street at dusk.
By: Mike Yardley
It's a gritty riverside city, with a raffish charm, built on the back of cotton. Lustily billed as the Birthplace of the Blues and the Home of Rock 'n' Roll, I began my urban adventures in Memphis at historic Beale Street, declared the "Home of the Blues" by an Act of Congress in 1977.
As the morning heat sparked up, and the bustle of Beale Street's bars, cafes and time-honoured businesses roared into life, I made my way to BB King's Blues Club, to join Backbeat Tours' Memphis Discovery Tour.
A frolic in Beale St.
I joined my fellow sightseers on-board "Miss Clawdy," a 1959 vintage bus, where our ebullient driver/guide, Larry, enchanted us over the course of three hours, unfurling a spree of endearing city insights.
As another FedEx cargo jet lumbered overhead, Larry remarked that Memphis is home to the world's second biggest cargo airport, beaten only by Hong Kong. Rolling through the downtown district, a plethora of multi-coloured giant murals added welcome injections of vitality in a city which in places looks scruffy around the edges.
Beale St, Memphis.
A notable exception is Victorian Village, an historic area once known as "Millionaire's Row." Many of the mansions date were built in the mid to late 1800s, like Mallory Neely House, Woodruff Fontaine House, James Lee House and Mollie Fontaine. Willy regaled us with tales about the original residents and their life stories. Another housing development which is a must-see is the Lauderdale Courts apartment complex.
Developed as a public housing project, in Colonial-Revival style, Elvis Presley's family lived in one of the apartments, when he was a teenager. It's now privately owned and is available to rent for a night. Just google Elvis Lauderdale Courts!
Stay at Elvis' home before Graceland.
We also called into the Rock 'N' Soul Museum, which delivers great insights on the social currents and changes that led to the development of Rock "n" Roll in Memphis. Another indelible highlight was calling into The Peabody Hotel for the legendary "Duck March." Larry quipped that "it's more like a duck sprint, than a march!"
This kooky tradition that holds court in The Peabody fountain began in the 1930s when the hotel's boss returned from a weekend hunting trip and thought it would be funny to place some of his live duck decoys in the lobby's fountain.
Nearly 90 years after the inaugural march, the resident ducks still visit the lobby fountain at 11am and 5pm daily, drawing hundreds of camera-toting onlookers inside the lobby to witness this truly bizarre spectacle.
Beale St.
In April this year, Memphis and much of the USA paused to commemorate the 50th anniversary of the assassination of Martin Luther King, Jr. Larry led us to the Lorraine Motel, where the fateful event took place, in South Main neighbourhood.
We stopped, reflected and gazed across the frozen-in-time site, outside Room 306, where he was shot and killed, while standing on the balcony. It's a sobering, goose-bumps experience to stand there and stare up.
Beale Street by day.
Five years ago, the National Civil Rights Museum, which incorporates the Lorraine Motel was given a NZ$40 million transformation.
The museum is a multi-sensory triumph, providing you with a searing journey through the backstory of oppression and the civil rights struggles, threaded deep into the American narrative.
Lorraine Motel.
The experience begins in a replica slave hold on a ship, before passing through rooms dedicated to the Jim Crow era. There's a replica of the Montgomery bus where Rosa Parks refused to give up her seat and lunch counters where students held sit-ins in the 1960s. Transitioning to MLK and his civil rights activism, the emotional journey climaxes outside room 306, the well-preserved guestroom he occupied before he was assassinated.
After such a powerful experience, we crossed the road to a Memphis favourite: Central BBQ. Brightly clad in vivid wall murals, and brimming with cheerful school groups, Central BBQ is a multiple-award-winning restaurant, where the slow smoked Memphis-style pork is cooked for 14 hours.
Larry the tour guide.
You can't go wrong with ribs or shoulders at Central, nor their delicious sides. But it's their sensational barbecue nachos, that are the talk of the town. Backbeat Tours offer a wide variety of Memphis experiences, leaving you with an unmistakeable taste of the city's soul, like a show on wheels. www.backbeattours.com
I jaunted back to Beale Street on several occasions, for more flavourful dips. Don't miss popping into Lanksy's, Clothier to the King. The 126 Beale Street store pays tribute to their VIP customers, including Elvis, with an on-site museum within the clothing store.
In 1952, Bernard Lansky noticed a young man who would repeatedly window-shop, without ever coming inside. Eventually, Bernard beckoned the 17- year old Elvis to come in. To cut a long story short, Lansky's provided much of Elvis's attire, including his outfit for his first appearance on The Ed Sullivan Show in 1956.
Bernard once remarked, "I put Elvis in his first suit, and I put him in his last." He died in 2011, but the business still does a roaring trade including a line of reproductions of clothing that Elvis actually wore. Celebrities still flock to the clothier.
The Peabody Duck Hotel.
Get some south in your mouth! BB King's Blues Club is a fabulous dining and entertainment haunt, while other star specimens on Beale Street include breakfast at Miss Polly's Soul City Café, (fried chicken and waffles) and Jerry Lee Lewis' Café & Honky Tonk. Packed with memorabilia, his multi-level venue showcases superb Delta musicians and he's probably got the best balcony-level view across the world famous street.
Finally, pay a visit to STAX: The Stax Museum of American Soul Music. Honouring the legacy of artists like Isaac Hayes and the Staple Singers, and with feature exhibits like an authentic 100-year old Mississippi Delta church inside, you'll get a deep appreciation of the origins and traditions of soul music.
From 1959 to 1974, this movie theatre-turned-recording studio produced a parade of hits from the likes of Otis Redding, Isaac Hayes and Booker T. & the MG's. Among the huge collection of more than 2,000 artifacts and exhibits, Hayes' gold-plated, peacock-blue Superfly Cadillac El Dorado is a delight.
Air New Zealand will start operating non-stop services between from Auckland and Chicago flying up to three times a week from 30 November 2018. One way Economy fares start from $1,019 (including taxes).
Fares are also available via Los Angeles, San Francisco and Houston with onward connections to Chicago on partner airlines. Visit www.airnewzealand.co.nz to book or for more details.
I organised my American rental car through Expedia, who offer a great range of sharply-priced car hire rates, no matter where you are travelling to. Did you know on Expedia you can get discounted rates on a great range of hotels when you add on to your existing trip? It's called the Expedia Add-On Advantage.
Book your flight, car or package on Expedia and you'll instantly unlock up to 49% off select hotels until the day of your trip. Jump to https://www.expedia.co.nz/addonadvantage
NORTHERN ADVOCATE | Sport
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COMMENT: We're Lamborghini, not Toyota, and we ought to start acting like it.
Demand leaps for Northland lifestyle blocks
26 Apr, 2019 11:18am
Northland lifestyle block sale prices have increased 27 per cent in the past year.


text:
Q: Check if a character is not in a string (pyt password = "e"
if any(c in password for c in "abcd"):
   print("yes")

I am trying to write a code that allows me to check if a character "e", is not included in "abcd".
However, the code above only prints yes if there's an occurrence in the password of a, b, c, or d. I don't know how to change it to have: If there's any occurrence of a character not present in abcd, in the password, then it returns yes.
(note if the password is "ea" for example, it should print "yes" as well)

A: any takes an iterable. It expects a True. Statement so you should use not in rather than in. Because if not in you want it to say True, not in.
if any([c, if c not in 'abcd' for c in password]):
    print("yes")


London: The GSMA today released a specification that allows consumers to remotely activate the SIM embedded in a device such as a smart watch, fitness band or tablet. This new specification, which is the first output from the GSMA's industry-backed Consumer Remote SIM Provisioning initiative, will enable consumers to add a new generation of devices to a mobile subscription and connect them securely to a mobile network.
Mobile network operators, mobile device manufacturers and SIM vendors have worked closely together through the GSMA initiative to deliver a specification that will allow consumers to easily activate the SIM embedded in a range of devices with a subscription from a mobile network operator of their choice. The initiative does not aim to replace all SIM cards in the field, but is instead designed to help users connect multiple devices through the same subscription and will help mobile device manufacturers to develop a new range of smaller, lighter mobile-connected devices that are better suited for wearable technology applications.
The specification is backed by the world's largest operators including AT&T, CK Hutchison, Deutsche Telekom, Etisalat, EE, KDDI, NTT DOCOMO, Orange, Rogers, SFR, Sprint, Telefónica, Telenor, TeliaSonera, Telstra, TIM, Transatel, Verizon and Vodafone. It is also supported by SIM and chipset vendors including Gemalto, Giesecke & Devrient, Morpho (Safran), Oasis Smart SIM, Oberthur Technologies, Qualcomm, STMicroelectronics, and Valid, as well as device makers including Apple, Huawei, LG, Microsoft, Samsung and Sony. Telecom equipment provider Nokia and security provider Trustonic also support the specification.
The GSMA will also release an additional specification in June 2016 that will include remote provisioning for any mobile device.
For further information on the GSMA Remote SIM Provisioning Initiative please visit: http://www.gsma.com/remoteSIMProvisioning.


» Is Microsoft building a B2B ad network?
Is Microsoft building a B2B ad network?
News that Microsoft is to buy LinkedIn for a reported $US26.2 billion in cash has left some analysts scratching their heads.
What LinkedIn gets from the deal, the largest in Microsoft's history and a generous 50 percent above market value, is relatively clear.
LinkedIn has been struggling. It has over 430 million registered users and gets 100 million visitors to its site each month, but growth and revenues have slowed.
LinkedIn's revenue is largely dependent on the premium subscriptions which provide corporate recruiters and salespeople with greater access to its members. And, for the past two years, LinkedIn has lost money.
After missing Q4 earnings earlier this year, its share price fell by over 40 percent in a single day, market value falling by $US14 billion.
LinkedIn CEO Jeff Weiner (who will now report directly to Microsoft CEO Satya Nadella) celebrated the fact that his company gets to retain its brand and independence.
Nadella, who says that Microsoft has learnt a lot from past mistakes, has plans that could significantly increase the value of LinkedIn. He told the Wall Street Journal that the acquisition represented "…the coming together of the professional cloud and the professional network".
While Microsoft doesn't have a good track record in making good on its acquisitions (Nokia and aQuantive come to mind), those purchases were made when Steve Ballmer was CEO.
Nadella envisions LinkedIn communities seamlessly communicating and interacting with Office 365 programs (including Outlook) and related training videos (courtesy of Lynda.com, acquired by LinkedIn in 2015). Cortana (Microsoft's somewhat better than Siri speech-recognition personal assistant) and its messaging app Skype are also mentioned as in the mix. And let's not forget Microsoft's Dynamics CRM (which, in accessing LinkedIn data, could become more competitive with Salesforce, reportedly an acquisition target Nadella walked away from last year).
In the future, according to Microsoft's public announcement of the deal, Cortana will "know your entire professional network…so you stay one step ahead".
Such changes, if successful, could not only help shore up Microsoft's cloud-based software business, but also make LinkedIn a more appealing prospect for paid subscribers.
But there may be a bigger plan behind Microsoft's big play.
We're not alone in thinking this.
Sahil Jain, CEO of cross-network ad platform AdStage, posits that Microsoft is planning a product suite aimed at B2B advertisers, offering search, social, display, mobile and retargeting options.
Bing Ads has nearly a third of search market share globally (yes, its share is much lower in New Zealand, but has been growing).
Regulatory approval for the deal is not expected until year's end, but LinkedIn is coming under new management. And $US26 billion is a lot of money in anyone's language: big plans are afoot.
While it's too early to guess what will become of LinkedIn, Microsoft's acquisition could well bolster Bing Ads' fortunes, making it particularly appealing to B2B advertisers. At the same time, the acquisition further deprives Google of social data; data that might have complemented its advertising network.
Google could use stronger competition, of course. And with Microsoft's involvement, LinkedIn advertising may yet realise its promising potential (even in smaller markets like ours).
15% of all Google searches have never been seen before.


text:
Fluorescence HPLC Analysis of the in-vivo Activity of Glucosylceramide Synthase
Neelanjan Bose
Istvan Stadler
Anonymous reviewer
Kartik R. Roy Sachin K. Khiste Zhijun Liu Yong-Yu Liu
Kartik R. Roy
School of Basic Pharmaceutical and Toxicological Sciences, University of Louisiana at Monroe, Monroe, LA 71209, USA
Sachin K. Khiste
Present address: Department of Medicine, Brigham and Women Hospital, Harvard Medical School, Cambridge, MA 02139, USA
Zhijun Liu
School of Renewable Resources, Louisiana State University Agriculture Center, Baton Rouge, LA 70803, USA
Yong-Yu Liu
yliu@ulm.edu
Published: Vol 9, Iss 12, June 20, 2019
Cancer Biology > Cancer biochemistry > Drug resistance
Cell Biology > Cell metabolism > Lipid
Biochemistry > Protein > Activity
Mammalia > Human > Cell line > Lipid
Mammalia > Murine > Whole animal > Lipid
Roy, K. R., Khiste, S. K., Liu, Z. and Liu, Y. (2019). Fluorescence HPLC Analysis of the in-vivo Activity of Glucosylceramide Synthase. Bio-protocol 9(12): e3269. DOI: 10.21769/BioProtoc.3269.
Almost all functions of cells or organs rely on the activities of cellular enzymes. Indeed, the in-vivo activities that directly represent the cellular effects of enzymes in live organs are critical importance to appreciate the roles enzymes play in modulating physiological or pathological processes, although assessments of such in-vivo enzyme activity are more difficult than typical test-tube assays. Recently, we, for the first time, developed a direct and easy-handling method for HPLC analyzing the in-vivo activity of glucosylceramide synthase (GCS). GCS that converts ceramide into glucosylceramide is a limiting-enzyme in the syntheses of glycosphingolipids and is one cause of cancer drug resistance. In our method developed, rubusoside nanomicelles delivers fluorescence N-[6-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]hexanoyl]-D-erythro-sphingosine (NBD C6-ceramide) into mice, tissues uptake the cell-permeable substrate, and GCS converts it into NBD C6-glucosylceramide in all organs simultaneously. Further, HPLC analyzes the extracted NBD C6-glucosylceramide to assess alterations of the in-vivo GCS activities in tissues. This method can be broadly used to assess the in-vivo GCS activities in any kind of animal models to appreciate either the role GCS plays in diseases or the therapeutic efficacies of GCS inhibitors.
Keywords: In-vivo enzyme activity, Glucosylceramide synthase, Ceramide, High-performance liquid chromatography, Nanomicelles, Rubusoside, Fluorescence NBD, Cancer
Glucosylceramide synthase (GCS; EC22.41.80; also known as ceramide glucosyltransferase) catalyzes ceramide glycosylation, transferring glucose residues from uridine diphosphate-glucose (UDP-glucose) to ceramide (Cer) and thereby producing glucosylceramide (GlcCer) (Basu et al., 1968; Ichikawa et al., 1996). Cer glycosylation by GCS is the first and rate-limiting step for the synthesis of glycosphingolipids (GSLs), over 400 species in human (Yu et al., 2009; Merrill, 2011; D'Angelo et al., 2013). Highly localized on GSL-enriched microdomains (GEMs) of cellular membranes, GSLs play crucial roles in modulating membrane integrity and functions, including cell-cell recognition, interaction and communication (Fishman and Brady, 1976; Yoshizaki et al., 2008; D'Angelo et al., 2013). GSLs are involved in mediating cell proliferation, differentiation, immuno-response and oncogenic transformation (Hakomori, 1981; Kasahara and Sanai, 2000; Hakomori, 2002).
Aberrant GCS or GSLs highly associate with several diseases. Recent studies concordantly indicate that enhanced GCS is one cause of cancer drug resistance (Liu et al., 2001, 2008, 2010 and 2013; Chiu et al., 2015; Roh et al., 2015). Further, inhibition of GCS, thereby decreasing GlcCer accumulation, is a treatment option for Gaucher's disease (McEachern et al., 2007) and type II diabetes (Aerts et al., 2007; Zhao et al., 2007).
Quantitative assessment of GCS activity is essential for evaluating the roles Cer glycosylation playing in cell functions, as well as in therapeutic efficacies of relevant disease treatments. After Basu's work (Basu et al., 1968), several additional methods have been developed to analyze GCS activities (Basu et al., 1987; Shukla and Radin, 1990; Ichikawa et al., 1996; Hayashi et al., 2005). Besides UDP-[3H]glucose being incorporated into ceramide to generate [3H]GlcCer (Shukla and Radin, 1990; Lavie et al., 1997; Liu et al., 1999), N-[6-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]hexanoyl]-D-erythro-sphingosine (NBD C6-Cer) has been used as an acceptor to produce NBD C6-GlcCer by the prepared GCS protein in optimal in-vitro conditions (Ichikawa et al., 1996; Bourteele et al., 1998; Hayashi et al., 2005). The HPLC method based on NBD C6-Cer has proven to be a highly sensitive and reproducible assay for assessing GCS activity in-vitro assays (Ichikawa et al., 1996; Hayashi et al., 2005). Convergently, previous studies have shown that NBD C6-Cer can be used as an exogenous substrate for characterizing cellular Cer glycosylation and assessing GCS activities with thin-layer chromatography (TLC) and spectrometry (Gupta et al., 2010; Liu et al., 2010; Gupta et al., 2012). With nanoparticle-based delivery of NBD C6-Cer, we developed a rapid, efficient, and fully quantitative HPLC analysis for assessing GCS in-vivo activity in live mice (Khiste et al., 2017).
Rubusoside (RUB) has been used to improve solubility and consequently bioavailability of etoposide (Zhang et al., 2012). RUB based nanomicelles (Cer-RUB) significantly increases the aqueous solubility and bioavailability of NBD C6-Cer in vivo (Khiste et al., 2017). After intraperitoneal administration, almost all tissues efficiently uptake and use NBD C6-Cer as substrate for Cer glycosylation by GCS. Analysis of the extracted sphingolipids with HPLC allows us directly assess the in-vivo activities that GCS simultaneously conduct among tissues in live mice. This direct, specific and effective approach can be applied to measure intracellular and intra-organ GCS activities in live cells and tissues, enabling further assessments of the roles played by GCS in diseases. Notably, this methodology may have considerable value in predicting and monitoring therapeutic efficacy of various anticancer drugs, and of other therapeutic agents used in the treatments of GCS-related diseases.
Falcon 100 mm cell culture dish (Corning, catalog number: 353003)
Dishes 35-mm (Corning, catalog number: 353001)
Paper towels Scott C-fold (Uline, catalog number: 560801)
BD 1 ml insulin syringes (Fisher Scientific, catalog number: 22-253-260)
Pasteur pipettes (Fisher Scientific, catalog number: 13-678-20D)
Microcentrifuge tubes, 1.5 ml (Fisher Scientific, catalog number: 0548129)
Polypropylene centrifuge tubes, 15 ml (Fisher Scientific, catalog number: 22-010-076)
Polypropylene centrifuge tubes, 50 ml (Fisher Scientific, catalog number: 05-539-6)
Screw thread vials, glass 4 ml, amber (Thermo Fisher Scientific, catalog number: C4015-2W)
Screw thread vials, glass 4 ml, transparent (Thermo Fisher Scientific, catalog number: C4015-21)
LVITM (low volume insert) vials, 0.3 ml, amber glass (Wheaton, catalog number: 225328)
96-well black flat bottom polystyrene microplates for fluorescence measurement (Fisher Scientific, catalog number: 07-200-589)
Polystyrene disposable pipets, 10 ml, sterile (Fisher Scientific, catalog number: NC9868325)
Glass pipettes, 1 ml, sterile (Fisher Scientific, catalog number:12-724)
Pipet tips, sterile, 200 μl and 1 ml (Fisher Scientific, catalog numbers: 02-707-502, 02-707-509)
0.45-μm nylon filter (Thermo Fisher Scientific, catalog number: 151-4045)
Sterile alcohol prep pads (Dynarex, catalog number: 1114)
Athymic nude mice (Foxn1nu/Foxn1+, 4-5 weeks, female from ENVOGO, catalog number: 069(nu)/070(nu/+))
Human ovary cancer OVCAR-3 (ATCC, catalog number: ATCC® HTB-161TM) (Khiste et al., 2017)
Mice carrying OVCAR-3 tumor xenografts and controls
Inoculate OVCAR-3 cells (purchased from ATCC; ~3-5 passages, 1 x 106/20 μl per mouse) in serum-free medium subcutaneously inoculate into the left flanks of athymic nude mice (Foxn1nu/Foxn1+, 4-5 weeks, female; Harlan).
In the control group, we only inoculate 20 μl serum-free medium. Once tumors sizes reach ~0.6 cm in diameters (~24 days), we will inject NBD Cer-RUB nanomicelles intraperitoneally in these mice for ceramide glycosylation.
RPMI 1640 Medium (Thermo Fisher Scientific, catalog number: 11875-093), store at 4 °C
Fetal bovine serum (FBS), heat inactivated (Thermo Fisher Scientific, catalog number: 10082147), store at -20 °C
Phosphate buffer saline (PBS, 10x) (Thermo Fisher Scientific, catalog number: 70011-044), keeping at 4 °C for using
NBD C6-ceramide (N-[6-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]hexanol]-D-erythro-sphingosine) (Avanti Polar Lipids, catalog number: 810209), store at -20 °C
NBD C6-glucosylceramide, (N-hexanoyl-NBD-glucosylceramide) (Matreya, catalog number: 1622-001)
PierceTM BCA protein assay kit (Thermo Fisher Scientific, catalog number: 23227)
Bovine serum albumin (BSA) (Sigma-Aldrich, catalog number: 9048-46-8), store at 4 °C
NP40 Cell Lysis Buffer (Thermo Fisher Scientific, catalog number: FNN0021), store at -20 °C
ProteaseArrestTM for Mammalian (G-Bioscience, catalog number: 786-433), store at 4 °C
Methanol for HPLC, ≥ 99.9% (Sigma-Aldrich, catalog number: 67-56-1)
Chloroform for HPLC, ≥ 99.5% (Sigma-Aldrich, catalog number: 67-66-3)
Water for HPLC (Fisher Chemical, catalog number: 7732-18-5)
Ortho-phosphoric acid for HPLC (Sigma-Aldrich, catalog number: 7664-38-2)
Ethanol, anhydrous (Fisher Chemical, catalog number: A405-20)
Rubusoside (RUB) (Sigma-Aldrich, catalog number: 62933-10MG)
NBD C6-ceramide-rubusoside (NBD Cer-RUB) solution (see Recipes)
NBD Cer-RUB nanomicelles (see Recipes)
NP40 protein lysis solution (see Recipes)
Solvent system for HPLC analysis (see Recipes)
Solvent A
Solvent B
NBD C6-ceramide or NBD C6-glucosylceramide standard solutions (see Recipes)
10 μM NBD C6-ceramide
10 μM NDB C6-glucosylceramide
Note: Store or keep all reagents at room temperature, except indicated items.
Pipette aids and pipette
High precision microdissection scissors (Fisher Scientific, catalog number: 08-953-1B)
Analytical balance (Mettler Toledo, catalog number: ME54TE/00)
Freezer, -20 °C (Fisher Scientific, catalog number: 18LC-16WW-FS)
Freezer, -80 °C (Thermo Scientific, catalog number: ULT1386-5-D41)
Standard cell culture hood
Vortex mixers (Fisher Scientific, catalog number: 02-215-360)
HPLC system, Agilent 1220 Infinity II LC with Agilent 1260 fluorescence detector (Agilent Technologies)
HPLC Colum, ZORBAX Rx-SIL 4.6 x 250 mm, 5 μM (Agilent Technologies, catalog number: 880975-901)
Centrifuge (Eppendorf, model: 5804 R)
Water bath and incubator
Reacti-Vap evaporators (Thermo Fisher Scientific, catalog number: TS-18826)
Ultrasonic processor (Fisher Scientific, model: CPX130PB)
Microplate Reader (BioTek, Synergy HTX multi-mode reader)
Prism 5v for statistical analysis (GraphPad Software Inc.)
Agilent OpenLab CDS (EZChrom edition, version A.04.05) for HPLC analysis
Briefly, we inject NBD C6-Cer-RUB nanomicelles (Zhang et al., 2012; Hosain et al., 2016; Khiste et al., 2017) into tumor-bearing mice, and the live tissues of mice convert NBD C6-ceramide into NBD C6-glucosylceramide. Three hours after injection, we collect all tissues, including tumors, and then extract the lipids. Further, we apply HPLC to analyze NBD glucosylceramide and assess the in vivo GCS activities of tissues.
Injection of NBD Cer-RUB into mice and in vivo glycosylation
Prepare NBD Cer-RUB solution (NBD C6-Cer 0.25 mg/ml) in sterile condition (Recipe 1) and keep on ice (Figure 1A).
Weigh body weight of each mouse on balance and calculate the injection volume of NBD C6 Cer-RUB solution, based on 1 mg/kg body weight for each mouse bearing OVCAR-3 tumor xenografts (Materials and Reagents #20). For instance, we inject 100 μl NBD C6 Cer-RUB solution containing 25 μg NBD C6-ceramide to a 25 g mouse.
In animal operating hood, take NBD Cer-RUB solution into a U-100 insulin syringe, inject intraperitoneally into mice (Recipes 1, 5 cases/group) following wipe inject-site skin with 70% alcohol (Figure 1B).
Keep injected mice in cages for 3 h, allowing uptake of NBD C6-ceramide and ceramide glycosylation in live animals. There are approximately 10 min intervals among the injections.
Euthanize mouse with cervical dislocation and dissect tissues, including liver, lungs and tumors from three mice per group, according to the injection order.
Rinse tissues with sterile PBS in 35-mm dishes twice and absorb with sterile paper towels.
Dissect and put tissue of each mouse into microcentrifuge tubes, keep on ice or store at -80 °C.
Figure 1. Intraperitoneal injection of NBD Cer-RUB nanomicelle solution to mice. A. NBD Cer-RUB complex are nanomicelles. B. NBD Cer-RUB is solubilized in RPMI-1640 medium and intraperitoneally injected into mouse for the in vivo glycosylation by GCS.
Tissue collection and extraction of lipids and proteins
Dissect and weigh two of 50 mg pieces from the same tissue of all samples (liver, lung, tumor from five mice per group), and put in 2 microcentrifuge tubes containing 50 μl sterile PBS, respectively. Use one set of samples for lipid extraction and another one for protein extraction.
Mince tissue with micro-scissor and sonicate it with an ultrasonic processor in PBS on ice.
For lipid extraction:
Add 100 μl water and transfer the tissue suspension into a 4-ml transparent glass vial for all samples.
Add 0.85 ml of water, 1.0 ml methanol and 1.0 ml chloroform in the glass vial and vortex for 30 s to mix. Keep samples at room temperature for 20 min and vortex for 30 s after every 5 min.
Centrifuge the glass vial tube at 4,500 x g, room temperature for 10 min, to separate the organic and aqueous phases.
Gently hold glass vial and transfer of the organic phase (bottom layer) with Pasteur pipette into a new amber 4-ml glass vial (Figure 2, Video 1).
Take 5 μl organic solution and add into a fluorescence 96-well plate with 145 μl methanol in duplicate. Measure fluorescence units in a microplate reader (BioTek Synergy HT, λ excitation 470 nm, λ emission 530 nm). The average fluorescence unites will be used to normalize all samples.
Transfer 800 μl organic solution of each sample with glass pipette into an amber 4-ml glass vial, and dry with nitrogen gas completely (~20 min). Tightly close the vials and store all samples at -20 °C for further use.
Figure 2. Transferring lipid extracts after extraction. After centrifugation, the organic solution is in the lower phase of vials. Transferring the lower phase, which contains extracted fluorescence lipids with Pasteur pipette into a new amber glass vial. This step needs to practice before experiments.
Video 1. Transferring lipid extracts in the lower-phase with Pasteur pipette. Holding vials between left thumb and index fingers with ~65°, insert pipette tip into the bottom layer to the lowest corner of the sample vial. Slowly aspirate the lower-phase solution only (~900 µl) with pipette-aid, and then transfer it into the new glass vial.
For protein extraction:
Add 200 μl NP40 lysis buffer with ProteaseArrest (Recipe 3) and mix with tissue suspension (from Steps B1 and B2), and vortex for 30 s.
Incubate the tubes on ice for 40 min, with vortex for 30 s per 5 min.
Centrifuge samples at 9,400 x g, 4 °C for 10 min.
Collect the supernatant of each sample in new microcentrifuge tubes, and store at -80 °C, except the portion for further protein measurement.
Add 2 μl of protein extracts in triplicate and mix with 100 μl BCA reagents (50:1, Reagent A:B), into 96-well plates. Use bovine serum albumin (BSA, 0, 5, 10, 20, 40 μg) to generate the standard curve.
Measure absorbance in a microplate reader (λ = 562 nm; BioTek Synergy HT) and obtain protein concentrations of samples (x μg/μl) with the BSA standard. The total protein amount: Pro = x × 160 (μg) for each sample (from 50 mg tissue used).
HPLC Analysis of NBD C6-Glucosylceramide of mouse tissues
Characterize and measure NBD C6-glucosylceramide with NBD C6-ceramide in standard solutions.
Connect the Agilent 1220 Infinity LC system with column (ZORBAX Rx-SIL 5 μm, 4.6 x 250 mm, normal phase) and solvents, and equilibrate with Solvent A, at flow rate 1 ml/min for 60 min.
Set up OpenLab CDS EZChrom program: flow rate 1 ml/min, injection volume 5 μl, linear gradient for 14 min (Solvent A 100%~0, and Solvent B 0~100%), and fluorescence detection (λ excitation 470 nm, λ emission 530 nm).
Add 100 μl solutions of NBD C6-ceramide (Cer) and NBD C6-glucosylcerade (GlcCer) into the LVI vials, respectively. These solutions contain both Cer and GlcCer at increasing concentrations (0, 0.032, 0.063, 0.125, 0.25, 0.5 and 1 μM; see Recipe 5) (Figure 1A).
Load the LVI vials in the carousel of HPLC system, run analysis triplicate and save the files.
Characterize NBD glucosylceramide generated of samples.
Add 100 μl of methanol/chloroform into in the amber glass vial of liver extracts to reconstitute lipids (Step B4f).
Transfer 50 μl of reconstituted liver lipids (~2,000 fluorescence units) into the LVI vials.
Add 2 μl NBD C6-glucosylceramide (0.125 μM) into the LVI vials and mix with liver lipids.
Load in carousel of HPLC system and analyze three times.
Measure NBD glucosylceramide and ceramide of samples.
Add 100 μl of methanol/chloroform into amber glass vials to reconstitute the extracted lipids of samples.
Transfer 50 μl of reconstituted samples (~2,000 fluorescence units) into the LVI vials and load in carousel of HPLC system.
Set up OpenLab CDS EZChrom program: flow rate 1 ml/min, injection volume 5 μl, linear gradient for 14 min (Solvent A 100%-0, and Solvent B 0-100%), and fluorescence detection (λ excitation 470 nm, λ emission 530 nm).
Run HPLC analysis triplicate and save the files.
Generate standard curves of Cer and GlcCer.
Determine the average retention times for NBD C6-ceramide (Cer) and NBD C6-glucosylceramide (GlcCer) in chromatograms of standards opening in OpenLab CDS EZChrom program. Locate Cer at 3.10 ± 0.07 min and GlcCer at 3.8 ± 0.06 min in this column and solvent system.
Transfer HPLC data (rslt.) from CDS EZChrom to Prism software. Define the AUC values at Y-axis and the corresponding pmols (per 5 μl, injected values) NBD C6-glucosylceramide (GlcCer) and C6-ceramide (Cer) at X-axis (Figure 1A).
Generate the standard curves with equation (Y = MX + C) for GlcCer. Here, Y represents AUC (area under the curve, fluorescence units), X lipid amounts, M slope and C equation constant. For GlcCer, Y = 177.42X + 5.0143.
Compare chromatograms of liver to confirm GlcCer in liver samples (Figure 3B).
Figure 3. HPLC chromatogram of sphingolipids. A. Quantitation of C6-Cer and C6-GlcCer. Mixed standards of NBD C6-Cer and C6-GlcCer (1:1) was separated and quantitated by HPLC with fluorescence detection. The retention times for C6-Cer and C6-GlcCer were 3.1 and 3.8, respectively. Each data point represents mean ± SD of three independent experiments. B. Characterization of NBD C6-GlcCer in mouse liver. Lipids extracted from livers of mice after administration of NBD C6-Cer-RUB (1 mg/kg, i.p.; 180 min) were spiked with NBD C6-GlcCer (0.25 pmol per 5 μl) and analyzed by HPLC, respectively.
Define the in vivo GCS activities of samples.
Characterize Cer and GlcCer in the chromatographs of samples and calculate GlcCer amounts, with standard curves and equation. GlcCer amount (pmol/100 μl extracts), Pgc = 20 × X = 20 × (Y-5.0143)/177.42, (Figure 4A).
Determine in vivo activities of GCS. The enzyme activity (fmol/μg) is briefly defined as the generated GlcCer (fmol) in 3 h by GCS in one μg of tissue proteins. Tissue GCS enzyme activities can be Zgcs = Pgc/Pro (Pgc from Data analysis step 2a, Pro from Step B4f) (Figure 4B).
Example of calculation:
The in-vivo activities of GCS in tumors are significant higher than those in normal tissues, for instances of lung and liver (Figures 4B and 4C).
Figure 4. Intra-organ Cer glycosylation by GCS in mice. A. HPLC chromatograms. NBD Cer-RUB nanomicelles (1 mg/kg, i.p.; 4 cases/group) were administered to mice bearing an OVCAR-3 tumor xenograft; 3 h after injection, tissues were rapidly resected for lipid extraction and HPLC analysis. Approximately equal-fluorescence-unit samples of extracted sphingolipids were analyzed by HPLC. B. Intra-organ GCS activities. ***, P < 0.001 compared to lung or liver. C. Western blotting of tissue GCS. Equal amounts of proteins (50 μg protein/lane) extracted were resolved and then immunoblotted with antibodies for GCS or GAPDH. GCS protein levels are represented as ratios of GCS/GAPDH optical densities averaged from four blots. *, P < 0.05 compared to lung or liver.
We take 80% of total lipid as well as total protein extracts in analyses and calculations of enzyme activities, which are 800 μl from 1 ml organic layer containing lipids, and 160 μl from 200 μl protein lysis buffer solution containing proteins, respectively.
Handling NBD C6-ceramide or glucosylceramide as well as dissected tissues with light-proof, including using amber glass vials, aluminum covers and dim lights.
NBD C6-ceramide complexed to BSA (Thermo Fisher Scientific, catalog number: N22651) can be used to analyze GCS activity in cells, as described previously (Gupta et al., 2010; Khiste et al., 2017).
Intraparietal injection of NBD Cer-RUB (1 mg/kg) can provide sufficient substrates for ceramide glycosylation to measure in vivo GCS activities for most tissues, including tumor xenografts, except brain of mice.
The analysis of NBD C6-Cer and NBD C6-GcLcer can also be carried on similar HPLC system with fluorescence detector from Waters, Shimadzu or other vendors. The retention time and detection sensitivity for NBD C6-GlcCer or NBD C6-Cer may vary with the solvents, column and HPLC system, but are still identical.
NBD Cer-RUB solution, 0.25 mg/ml
Add 25.28 mg NBD Cer-RUB nanomicelles (containing 0.5 mg NBD C6-ceramide) and 2 ml RPMI-1640 medium (serum-free) in a sterile 4-ml glass vial, vortex for 30 s, and keep at room temperature before animal injection
NBD Cer-RUB nanomicelles
NBD C6-ceramide (NBD Cer) is complexed with rubusoside (RUB) (1:50, w/w) to generate NBD Cer-RUB nanomicelles, as described previously (Khiste et al., 2017).
a.	Dissolve NBD Cer in an ethanol solution containing RUB (140 g/100 ml)
b.	Filter this solution through a 0.45-μm nylon filter
c.	Evaporate and lyophilize to generate an NBD C6-Cer-RUB complex in powder form
d.	Store NBD Cer-RUB powder at 4 °C
NP40 protein lysis solution
Add 198 μl NP40 Cell Lysis Buffer and 2 μl ProteaseArrestTM solution in 1.5 ml microcentrifuge tube, mix and keep it on ice before use
Solvent system for HPLC analysis
Solvent A:
Mix 400 ml chloroform, 100 ml methanol and 0.5 ml ortho-phosphoric acid in a 500 ml glass bottle and close it tightly
Solvent B:
Mix 300 ml chloroform, 170 ml methanol, 30 ml HPLC water and 0.5 ml ortho-phosphoric acid in a 500 ml glass bottle and close it tightly
NBD C6-ceramide or NBD C6-glucosylceramide standard solutions
10 μM NBD C6-ceramide (stock solution, -20 °C):
Add 10 μg NBD C6-ceramide (mw 575.75) and 1.737 ml chloroform/methanol (5:1, v:v) in a 4-ml amber glass vial, vortex and close it tightly
10 μM NDB C6-glucosylceramide (stock solution, -20 °C):
Add 10 μg N-hexanoyl-NBD-glucosylceramide (mw 738) and 1.355 ml chloroform/methanol (5:1, v:v) in a 4-ml amber glass vial, vortex and close it tightly
Note: Each stock solution will be diluted two or three times with chloroform/methanol and the diluted ones will be used to generate standard curves.
This work was supported by Public Health Service/NIH grant R15CA167476 from the National Cancer Institute (Y.Y.L) and P20 GM103424-11 from the National Institute of General Medical Sciences (Y.Y.L). This work was also partially supported by the Mizutani Foundation for Glycoscience, Japan (MFRG120068 Y.Y.L).
All animal experiments were approved by the Animal Care and Use Committee of the University of Louisiana at Monroe (ULM) (Protocol# 16DEC-YYL 01) and were conducted in strict accordance with good animal practice as defined by NIH guidelines.
Aerts, J. M., Ottenhoff, R., Powlson, A. S., Grefhorst, A., van Eijk, M., Dubbelhuis, P. F., Aten, J., Kuipers, F., Serlie, M. J., Wennekes, T., Sethi, J. K., O'Rahilly, S. and Overkleeft, H. S. (2007). Pharmacological inhibition of glucosylceramide synthase enhances insulin sensitivity. Diabetes 56(5): 1341-1349.
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Chiu, W. H., Su, W. C., Li, C. L., Chen, C. L. and Lin, C. F. (2015). An increase in glucosylceramide synthase induces Bcl-xL-mediated cell survival in vinorelbine-resistant lung adenocarcinoma cells. Oncotarget 6(24): 20513-20524.
D'Angelo, G., Capasso, S., Sticco, L. and Russo, D. (2013). Glycosphingolipids: synthesis and functions. FEBS J 280(24): 6338-6353.
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Gupta, V., Bhinge, K. N., Hosain, S. B., Xiong, K., Gu, X., Shi, R., Ho, M. Y., Khoo, K. H., Li, S. C., Li, Y. T., Ambudkar, S. V., Jazwinski, S. M. and Liu, Y. Y. (2012). Ceramide glycosylation by glucosylceramide synthase selectively maintains the properties of breast cancer stem cells. J Biol Chem 287(44): 37195-37205.
Gupta, V., Patwardhan, G. A., Zhang, Q. J., Cabot, M. C., Jazwinski, S. M. and Liu, Y. Y. (2010). Direct quantitative determination of ceramide glycosylation in vivo: a new approach to evaluate cellular enzyme activity of glucosylceramide synthase. J Lipid Res 51(4): 866-874.
Hakomori, S. (1981). Glycosphingolipids in cellular interaction, differentiation, and oncogenesis. Annu Rev Biochem 50: 733-764.
Hakomori, S. (2002). The glycosynapse. Proc Natl Acad Sci U S A 99(1): 225-232.
Hayashi, Y., Horibata, Y., Sakaguchi, K., Okino, N. and Ito, M. (2005). A sensitive and reproducible assay to measure the activity of glucosylceramide synthase and lactosylceramide synthase using HPLC and fluorescent substrates. Anal Biochem 345(2): 181-186.
Hosain, S. B., Khiste, S. K., Uddin, M. B., Vorubindi, V., Ingram, C., Zhang, S., Hill, R. A., Gu, X. and Liu, Y. Y. (2016). Inhibition of glucosylceramide synthase eliminates the oncogenic function of p53 R273H mutant in the epithelial-mesenchymal transition and induced pluripotency of colon cancer cells. Oncotarget 7(37): 60575-60592.
Ichikawa, S., Sakiyama, H., Suzuki, G., Hidari, K. I. and Hirabayashi, Y. (1996). Expression cloning of a cDNA for human ceramide glucosyltransferase that catalyzes the first glycosylation step of glycosphingolipid synthesis. Proc Natl Acad Sci U S A 93(10): 4638-4643.
Kasahara, K. and Sanai, Y. (2000). Functional roles of glycosphingolipids in signal transduction via lipid rafts. Glycoconj J 17(3 -4): 153-162.
Khiste, S. K., Hosain, S. B., Dong, Y., Uddin, M. B., Roy, K. R., Hill, R. A., Liu, Z. and Liu, Y. Y. (2017). Incorporation of fluorescence ceramide-based HPLC assay for rapidly and efficiently assessing glucosylceramide synthase in vivo. Sci Rep 7(1): 2976.
Lavie, Y., Cao, H., Volner, A., Lucci, A., Han, T. Y., Geffen, V., Giuliano, A. E. and Cabot, M. C. (1997). Agents that reverse multidrug resistance, tamoxifen, verapamil, and cyclosporin A, block glycosphingolipid metabolism by inhibiting ceramide glycosylation in human cancer cells. J Biol Chem 272(3): 1682-1687.
Liu, Y. Y., Gupta, V., Patwardhan, G. A., Bhinge, K., Zhao, Y., Bao, J., Mehendale, H., Cabot, M. C., Li, Y. T. and Jazwinski, S. M. (2010). Glucosylceramide synthase upregulates MDR1 expression in the regulation of cancer drug resistance through cSrc and β-catenin signaling. Mol Cancer 9: 145.
Liu, Y. Y., Han, T. Y., Giuliano, A. E. and Cabot, M. C. (1999). Expression of glucosylceramide synthase, converting ceramide to glucosylceramide, confers adriamycin resistance in human breast cancer cells. J Biol Chem 274(2): 1140-1146.
Liu, Y. Y., Han, T. Y., Giuliano, A. E. and Cabot, M. C. (2001). Ceramide glycosylation potentiates cellular multidrug resistance. FASEB J 15(3): 719-730.
Liu, Y. Y., Hill, R. A. and Li, Y. T. (2013). Ceramide glycosylation catalyzed by glucosylceramide synthase and cancer drug resistance. Adv Cancer Res 117: 59-89.
Liu, Y. Y., Yu, J. Y., Yin, D., Patwardhan, G. A., Gupta, V., Hirabayashi, Y., Holleran, W. M., Giuliano, A. E., Jazwinski, S. M., Gouaze-Andersson, V., Consoli, D. P. and Cabot, M. C. (2008). A role for ceramide in driving cancer cell resistance to doxorubicin. FASEB J 22(7): 2541-2551.
McEachern, K. A., Fung, J., Komarnitsky, S., Siegel, C. S., Chuang, W. L., Hutto, E., Shayman, J. A., Grabowski, G. A., Aerts, J. M., Cheng, S. H., Copeland, D. P. and Marshall, J. (2007). A specific and potent inhibitor of glucosylceramide synthase for substrate inhibition therapy of Gaucher disease. Mol Genet Metab 91(3): 259-267.
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How to cite: Roy, K. R., Khiste, S. K., Liu, Z. and Liu, Y. (2019). Fluorescence HPLC Analysis of the in-vivo Activity of Glucosylceramide Synthase. Bio-protocol 9(12): e3269. DOI: 10.21769/BioProtoc.3269.
All Field Materials and Reagents Equipment Software Procedure Data analysis Notes Recipes Acknowledgments Competing interests Ethics References


It all started in a small blacksmith workshop in central Stockholm back in 1892, when F.W. Lindqvist and J.V. Svenson created the world's first soot-free kerosene stove! As it was the first, they decided to call it Primus (meaning first). The name became synonymous with reliable stoves and the stoves became the primary choice for people heading off on both first ascents and demanding expeditions. One of the milestone adventures was when Roald Amundsen became the first person to reach the South Pole in 1911. Another milestone is the one of Tenzing Norgay and Edmund Hillary at Mount Everest in 1953, making them the first people to reach the summit.


General Contractor services for the Rio Grande Valley, with its home office on South Padre Island. Specializing in Residential, Light Commercial and Restoration.
Willis Development has been general contractor and developer of properties on South Padre Island, Brownsville, McAllen, and Edinburg, Texas since 2003. General contractor projects have included single family residences and single family attached, midrise condominiums, gated community, large scale marinas, public projects for local municipalities, exterior restoration of high and mid-rise condominiums and remodel of single family residences.


The South Lake Hospital Robotic Surgery Program is made up of leading surgeons in their respective fields who perform minimally invasive surgery through state-of-the-art robotic technology for improved precision and patient outcomes. We offer the highest-quality robotic procedures based on quality measures that are used to continually monitor and improve our doctors' surgical performance. South Lake Hospital has created an environment that fosters collaboration in the practice of robotic surgical services, research and teaching.
Accredited by Surgical Review Corporation as a Center of Excellence in Robotic Surgery, Orlando Health South Lake Hospital is set apart from other outpatient facilities by providing the highest quality of care through an advanced offering of robotic surgeries utilizing the latest proven techniques, equipment, and processes.


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Accueil (EMEA) » Resources » Case Studies » iPERL International Customer Stories
iPERL International Customer Stories
Meet iPERL™—the data-driven smart water meter that ensures your networks run efficiently and accurately whilst increasing your business intelligence in near-real time.
Does your network run with unique, cutting edge technology that provides industry-leading accurate and near-real-time data?
iPERL™ is a next generation, solid-state water meter with integrated bi-directional communications capabilities ensuring near-real-time control anywhere in your network. The invaluable benefit of having a meter with no moving parts is seen with the flexibility it affords when considering installation locations; trapped air, dirty water and sand do not affect the high-performance of the iPERL at any stage of its service life.
How many other meters use innovative technology that guarantees sustained accurate data reads over their service life? This ensures you have up-to-date business intelligence, enabling optimum operational efficiency across your entire network and a wealth of data allowing you to conserve water, provide accurate billing to customers and to inform them about leaks, fraud or unusual usage patterns. Sounds pretty smart to us…
iPERL's key features:
Low-flow measurement: With a flow measurement down to 1 litre per hour, iPERL helps utilities to reduce undetected leakages due to next-generation sensors embedded in the endpoint. This technology also includes remnant magnetic field technology, which differs from solid-state meters and provides a constant linear working flow range
A wealth of data: The endpoint can be programmed to provide highly granular data at 15-second intervals. The embedded innovative communication enables the possibility of obtaining very detailed data from the meters, including the log of up to 2880 data points
and alarms. The data collected helps water utilities to manage distribution networks more efficiently, to conserve water, to provide accurate billing to customers and to inform them about leaks, fraud or unusual usage patterns
Solid-state meter with no moving parts: Unaffected by trapped air, sand or particles within the water, which ensures accurate readings from the beginning until the end of its service life, and subsequently preventing unplanned maintenance costs
15-year service life: There's no need for meter re-calibration or replacement as the measurement accuracy of the meter is certified for life
Read below to find out how companies around the world have benefited from the industry leading technology Sensus offers.
The city of Satu Mare, Romania, installs 30,000 iPERL smart water meters and experiences increased operational efficiency, valuable business intelligence and a reduction in non-revenue water.
The city of Satu Mare, Romania, had a water network that was riddled with problems, one of which was a discrepancy between sub-meters and the main meters in a house or apartment building. This was commonly seen when using mechanical meters, resulting in a 20-25% difference in readings from the sub-meters and the main meters. This left the city with the challenge of having to either bill the customer individually or not at all.
Another issue the city suffered was that of meter fraud with consumers turning the meter the wrong way to create a reverse flow. Overall, the city's levels of sustainability.
After conducting research into the types of technologies that could help, Satu Mare discovered that the iPERL smart meter was well suited to put an end to inaccurate readings, lack of data and fraud as the meters come fitted with alarms to alert the utility of any tampering.
The city decided to install 30,000 iPERL meters in 7,040 homes and 22,960 in apartment buildings. The devices were rolled out in a very short time frame and set up for walk-by/drive-by readout, which would help increase the utility's operational efficiency and costs.
The drive-by readings were so efficient that the water utility was able to reduce meter read-out times of 2,500 devices that would usually require 10 people over seven days, to two people over a mere two hours and 30 minutes. With the high-performance meters, they were able to detect an increase in consumption, with peaks over time which lead the utility to investigate and discover a consumer leak.
The city of Satu Mare was pleased with the results that the data from the smart meters provided, as well as the benefits that having a meter reading at a distance provided the employees with. Now the utility can look forward to increasing operational efficiency, accurate data and conserving water.
How can your smart water grid benefit from being connected at every point?
Reduce non-revenue water through accurate data
Improve operational efficiency and customer service with bi-directional radio communication
Performance uninterrupted by installation due to no moving parts
Flexible meter reading solutions with either handheld devices or walk-by/drive-by or fixed network options
The market town of Dießen, Germany, relies on economical, ecological and future-proof iPERL metering solution from Sensus to install regulation-compliant lead-free meters.
Dießen, Germany, needed to future-proof their water meter network after the German Drinking Water Ordinance introduced a stricter threshold for the amount of lead allowed in drinking water. It was changed to a maximum of 10 μg/l which serves to encourage water utilities to modernise their drinking water installations.
To ensure they comply with the regulations in the future, the town of Dießen has made a conscious decision to install lead-free meters like the iPERL. Michael Deininger, the Technical Director of the Dießen water utility, said: "When choosing our new water meters, it was important for the materials to comply with the permissible values, but also ensure our decision is future-proof."
The iPERL smart water meter consists of composite materials and is 100% lead-free, so water utilities can confidently rely on iPERL to ensure they are on the safe side even if threshold values for lead are lowered or if environmental safety regulations become stricter.
"The exceptionally long service life was yet another reason for us to choose iPERL", Deininger explains. "Longer deployment cycles mean we can reduce the costs which arise from exchanging meters and we can then invest the money this saves in additional state-of-the-art, durable meters." The added features the meter brings like contactless data reads and magnetic-inductive flow measurements ensure reliable meter performance throughout iPERL's long and unrivaled service life. Also, process steps such as venting or flushing meters can be dispensed with which eliminates the risk of faulty installation or implementation. These are important economic advantages and have a positive effect on a utility's operations.
The iPERL has the Widest working-flow range on the market
It can withstand temperatures as low as -20°C
The iPERL is made from sustainable endpoint material
It has a unique power supply design
Only the water in the supply is measured so air does not add to the volume recorded
The Bad König municipal utility company in Germany installed 2,700 iPERL smart water meters in 2014, increasing their operational efficiency and reducing water loss.
The Bad König municipal utility company wanted to be the pioneering water utility in Germany moving away from traditional mechanical meters to smart meters, in order to function in a sustainable manner and increase operational efficiency.
They investigated several suppliers to see who would be able to supply them with the best solution: "When we went to see the Sensus plant in Ludwigshafen with its highly sophisticated test labs, we were totally convinced by the manufacturer's expertise," says Markus Kunkelmann, Head of Technical Operations at Bad König's municipal utilities.
They began their pioneering project by installing quality metrology with its low start-up threshold of just one litre per hour and noted: "We were really surprised with the difference iPERL makes in comparison to the more traditional impeller wheels and oscillating piston meters," says Kunkelmann. The technical support provided by Sensus played a key role in achieving such positive results in Bad König.
Following the installation of 2,700 meters, the time of data transfer was reduced from a staggering 960 hours to 51. Not to mention, one million litres of drinking water saved in the first year following the installation of iPERL meters.
The economic benefits of iPERL also become apparent when reading the consumption data. Before they deployed iPERL, four members of staff needed 30 days to send the water consumption details to the accounts department. However, with walk-by readings, the complete supply network can be read and all the data sent to the billing department without any errors in just over two days. They have optimised their operating results, reduced water loss, improved their customer relationships and are helping to protect the environment as well.
If you can't measure, you can't manage
The data captured by a Sensus iPERL smart water meter, which is often used in conjunction with the FlexNet communications platform, provides your business with the accurate information to identify and help manage network issues proactively and efficiently. It also helps you address regulatory and environmental challenges whilst improving customer service, security of supply and operational efficiencies.
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Logging onto the Ordnance Survey website to check this month's map reference, I noticed that they're suggesting you should use the "green gym". Rather than spending money on gym or slimming club subscriptions, they recommend going walking to shed those excess calories from the seasonal celebrations. So here's a local walk you can shorten or lengthen to suit you and start off your new regime.
Blackwater Valley is on the borders of three counties; Berkshire, Hampshire and Surrey. The Blackwater Valley Path is 23 miles in total, following the blue coot sign along the River Blackwater, from its source between Farnham and Aldershot, flowing through a shallow valley before eventually joining the River Loddon near Reading.
This walk follows part of the Valley Path, starting at the car park off Mill Lane between Yateley and the A321 to Sandhurst. From here you can walk all around Horseshoe Lake, a route of about 1½ miles. Alternatively, turn left along the Three Counties Path to find a map describing a 3½ mile circular route, (or 5 miles including a walk through Finchampstead Ridges), which we'll follow in a clockwise direction.
The route follows along the southern edge of Horseshoe Lake first, on the Berkshire side of the river, beyond the wooden bridge which crosses into Hampshire, and past the two lakes of Moor Green Lakes Nature Reserve. The Reserve was created from former gravel extraction sites. After the Second World War the demand for building materials for roads and houses meant that many fields in the Valley were extracted for gravel and sand in the fifties and sixties, resulting in the lakes that remain today.
The three lakes with their gravel islands do attract wildfowl, making it an important site for breeding and wintering birds and an ideal place to go twitching at this time of year. So remember to pack your bins and bird ID books, and definitely your wellies.
Over 200 species of bird have been recorded in the area. Sightings in the last month include Little Egrets, Heron, Cormorant, Pintail, Smew, a Green Sandpiper, Water Rails, Snipe, Redwing, Fieldfare, Siskins, Lesser Redpoll, Buzzard and Sparrowhawk. Little Owls have been spotted and a Lesser-spotted Woodpecker visited in December.
Other wildlife you may see around are Roe deer, foxes and Pipistrelle and Daubenton's bats. Willow, Ash, Hazel, Maple and scrub fringe the lakes.
There are two hide screens on the route (the hides themselves are locked), one on the south east corner of Grover Lake and the other after the path turns right at the south west corner of Colebrook Lake. From these you can look out for birds on the gravel islands and beaches and specially created scrapes (very shallow water areas).
From here, count how many of the nine species of duck (Wigeon, Gadwall, Teal, Mallard, Pochard, Shoveler, Tufted Duck, Goldeneye and Goosander), five kinds of geese (Greylag, Canada, Egyptian, Barnacle, and for the really lucky Snow Geese) and five types of gulls (Black-headed, Lesser Black-backed, Common, Herring and Yellow-legged), which are expected to visit the area in January you can spot.
Stay on the path north from the hide to the other Car Park by Moor Green Farm (which closes at 4pm until March). Turn right onto Lower Sandhurst Road, then left along the footpath by Hall Farm, which will take you to the corner of Finchampstead Ridges, with great views over the three counties, but on a definite gradient. Sixty acres of the Ridge are now owned and looked after by the National Trust, but it once was part of a royal hunting forest, offering deer and wild boar hunting.
There is a legend that Prince Arthur, King Henry VII's son was hunting on the Ridges when he heard that Catherine of Aragon, his fiancée, had arrived in England. He and his party rode to meet her, but met a party of Spaniards who warned the Prince not to see his bride before they married. He ignored their advice and had Catherine's coach ride along the Ridges, but as they met, Catherine raised her veil. This was thought to have put a curse on the marriage, as Arthur died soon after the wedding.
Follow any path on the right in Finchampstead Ridges to come out on the stunning Wellingtonia Avenue, built by John Walters, who was MP of Bear Wood Estate in 1863. In 1869 it was planted with Wellingtonia trees, which are the world's largest tree. They can grow to over 350ft, weigh 1000 tons and live for 4000 years in their native habitat and the cones stay on the trees for about 20 years before they drop.
Opposite the National Trust Car Park at Simons Wood, the path leaves Wellingtonia Avenue on the right to run south back to Fisher's Copse, where you can pick up a footpath back on the left. Alternatively, to shorten the walk at the Ridges, turn right at Spout Pond and follow this path directly to Fisher's Copse (from this direction, the footpath back will be on your right). Walk south east past Beech Hill to meet Ambrose Lane by Coalpit Copse. Walk a little way east along the Lane to where it bends to the left at Ambarrow Wood and then take the Three Counties Path on the right past the sailing centre and Ambarrow Farm back to the car park.


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