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AP-MALDI Mass Spectrometry Imaging of Gangliosides Using 2,6-Dihydroxyacetophenone
Shelley N. Jackson ORCID: orcid.org/0000-0002-3296-69131,
Ludovic Muller1,
Aurelie Roux1,
Berk Oktem2,
Eugene Moskovets2,
Vladimir M. Doroshenko2 &
Amina S. Woods1
Journal of The American Society for Mass Spectrometry volume 29, pages1463–1472(2018)Cite this article
Matrix-assisted laser/desorption ionization (MALDI) mass spectrometry imaging (MSI) is widely used as a unique tool to record the distribution of a large range of biomolecules in tissues. 2,6-Dihydroxyacetophenone (DHA) matrix has been shown to provide efficient ionization of lipids, especially gangliosides. The major drawback for DHA as it applies to MS imaging is that it sublimes under vacuum (low pressure) at the extended time necessary to complete both high spatial and mass resolution MSI studies of whole organs. To overcome the problem of sublimation, we used an atmospheric pressure (AP)-MALDI source to obtain high spatial resolution images of lipids in the brain using a high mass resolution mass spectrometer. Additionally, the advantages of atmospheric pressure and DHA for imaging gangliosides are highlighted. The imaging of [M–H]− and [M–H2O–H]− mass peaks for GD1 gangliosides showed different distribution, most likely reflecting the different spatial distribution of GD1a and GD1b species in the brain.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is the most common ionization source for mass spectrometry imaging (MSI) of biological tissues. Its development has enabled the detection and spatial localization of compounds directly from tissue sections [1,2,3]. The use of MALDI-MSI in lipidomic studies has greatly increased and has been used to image all major lipid classes in several tissue types [4,5,6]. Due to the diverse structures of lipids, the selection of an appropriate matrix is key to a successful analysis of the desired lipid class [7, 8]. 2,6-Dihydroxyacetophenone (DHA) is an excellent MALDI matrix for direct tissue analysis of a wide range of lipid classes in both positive and negative ion modes [9,10,11]. DHA matrix has been used to obtain rat brain tissue images illustrating the distribution of all major ganglioside species, glycerophosphocholines (PC), and sphingomyelins (SM) using a MALDI-linear ion trap mass spectrometer with an intermediate-pressure (IP) ion source at 0.07 Torr [12, 13]. In these studies, the laser step size was 75 [12] and 80 [13] μm and the sample run time was under 5 h. Despite this success, the major limitation of DHA as a matrix for MSI is that it sublimes under vacuum or intermediate-pressure conditions and thus is not applicable for high mass resolution MSI on a MALDI Orbitrap due to the long run times. The development of MS systems with high mass resolution greatly improved the assignment of lipids and the clarity of generated images by reducing the amount of overlap between mass peaks [14,15,16,17,18]. Recently, DHA has been used to image lipids in mouse brain on a high-throughput MALDI time-of-flight (TOF) MS system [19]. In this study, lipids were imaged in both positive and negative ion modes at a spatial resolution of 50 μm for an entire sagittal mouse brain section in under 40 min. This allowed for the imaging run to be completed before DHA sublimed in the low-pressure environment of the ion source.
Another way to prevent DHA sublimation is to use an atmospheric pressure (AP) MALDI ion source [20,21,22,23,24]. Since an AP-MALDI source operates at atmospheric pressure, volatile solid organic matrices, like DHA, and liquid matrices can be used without any time constraints relating to sublimation of the matrix. Furthermore, since the tissue section is kept at atmospheric pressure, dehydration of the sample is reduced compared to vacuum MALDI. Additionally, AP-MALDI is a softer ionization technique than vacuum MALDI and the fragmentations that are produced in-source are more reproducible [20, 25, 26]. An AP-MALDI source equipped with an infrared laser has been used successfully to image small carbohydrates in fruit tissue using the native water content as a matrix [27]. MSI of phospholipids in rat brain tissue with AP-MALDI with DHA matrix has been conducted but the data was collected on an ion trap with low mass resolution [28]. Recently, an AP-MALDI source has been coupled with an Orbitrap mass spectrometer and used to obtain high spatial and mass resolution of lipids in different tissues and cells using DHB or DHB/CHCA matrix [29,30,31]. In this work, we combined the AP-MALDI source, the volatile DHA matrix, and an Orbitrap mass spectrometer for MSI analysis. Due to the stability of DHA at atmospheric pressure, the duration of the MSI run was no longer a limiting factor. This permitted very long MSI runs utilizing smaller pixels and MS instrumentation with higher mass resolution. This work demonstrates the benefits of using the DHA matrix for imaging a large variety of lipid classes in tissues. Particular emphasis is placed on the analysis of gangliosides with DHA matrix at atmospheric pressure. Finally, we demonstrated the potential of in-source fragmentation to map the different distribution patterns observed for structural isomers of GD1 in tissue sections.
Mass Spectrometry Imaging
An AP-MALDI ion source (AP/MALDI(ng)UHR source, Mass Tech, Columbia, MD) was coupled to a Q Exactive and a LTQ Orbitrap Velos (Thermo Fisher Scientific, San Jose, CA) for mass analysis. The AP-MALDI ion source was equipped with a diode-pumped solid-state laser (λ = 355 nm) operating at a 0.1–10 kHz repetition rate. In the current work, the repetition rate was set at 1 kHz. This rate was chosen to allow for the best signal intensity without signal degradation in accordance with the sample stage motor speed that yielded 50/60 μm spatial resolution on the Orbitrap mass spectrometer. Higher repetition rates could be used on faster mass analyzers such as time-of-flight mass spectrometer. Maximum laser pulse energy was 3 μJ at 1 kHz repetition rate and a beam attenuator was used to adjust laser energy. The voltage applied between the MALDI plate and MS inlet was 5 kV. The distance between the MALDI plate and the face of inlet capillary was 3 mm. A continuous laser raster sampling was used to acquire MS images. Data was acquired in either negative or positive ion modes with mass resolution up to 140,000 at m/z 200 and between 531 to 2020 laser shots per pixel depending upon the instrumental settings (sample stage velocity and spatial resolution). A MALDI-LTQ-XL Orbitrap (Thermo Fisher Scientific) was used for mass analysis at intermediate pressure (IP) as a comparison to the AP-MALDI source. MS images were acquired in positive ion mode with a mass resolution of 60,000 at m/z 400 and a spatial resolution of 50 μm with three laser shots per scan. The measured pressure for the IP-MALDI source was 0.07 Torr. Thermo's ImageQuest software was used to generate MS images. The number of pixels and pixels per seconds is reported with each MALDI-MS image. However, the AP-MALDI acquisition software does not permit a free draw option when programming the area to be imaged (i.e., a rectangle shape is used when the brain is not a rectangle shape thus many pixels are not associated with tissue image); thus, the total number of pixels will vary when compared to the IP-MALDI data for serial sections, in which Thermo's Xcalibur software allows for a free draw option and better reflects the shape of the tissue being imaged. To give a better estimate of number of pixels on the actual tissue section, we used a custom IDL software developed by Ionwerk's Inc. (Houston, TX) to extract the number of pixels from a region of interest. For AP-MALDI images, we also provide the number of pixels for the tissue section image.
Lipid Standards
Ganglioside GD1a (disialo, diammonium salt, bovine brain, no. 1062) and GD1b (disialo, diammonium salt, bovine brain, no. 1501) were purchased from Matreya (Pleasant Gap, PA). Stock solutions of the lipids were prepared in chloroform to methanol ratio (2:1 v/v) at a concentration of 1 mg/mL. Ganglioside standards were diluted in matrix solution prior to being spotted on the sample target.
Sample Preparation for Lipid Standards
The MALDI matrices, 2,6-dihydroxyacetophenone (DHA, no. 37468, Sigma-Aldrich, St. Louis, MO), 1,5-diaminonaphthalene (DAN, no. 56451, Sigma-Aldrich), and 9-aminoacridine (9-AA, 92817, Sigma-Aldrich) were used to analyze the GD1 standards. DHA was prepared at 10 mg/mL concentration in 70% methanol with 125 mM ammonium sulfate (AmSulf) and 0.05% heptafluorobutyric (HFBA). DAN and 9-AA were prepared at a concentration of 10 mg/mL in 70/30 v/v methanol to water ratio. GD1 standards were diluted 1:4 v/v in the matrix solution, vortexed, and 1 μL was spotted onto the sample plate.
Tissue Preparation
The animal work in this study follows the Guide for the Care and Use of Laboratory Animals (NIH). Adult male Sprague-Dawley rats and C57BL/6 mice were sacrificed and their brains were immediately removed and frozen in dry ice-chilled isopentane. Frozen brain tissue was cut into thin sections (18-μm thickness) using a cryostat (Leica Microsystems CM3050S, Bannockburn, IL) at – 21 °C (cryochamber temperature) and −18 °C (specimen cooling temperature). Tissue sections were directly deposited on stainless steel sample plates or glass microscope slides for MALDI imaging.
Sample Preparation for Tissue Imaging
DHA was prepared at 10 mg/mL concentration in 50% ethanol with 125 mM AmSulf and 0.05% HFBA. DAN was prepared at 10 mg/mL concentration in 90% acetonitrile. A TM-sprayer (HTX Technologies, Chapel Hill, NC) was used to coat mice brain tissue sections with DHA and DAN. The following parameters were employed for coating tissue: flow rate = 0.1 mL/min, velocity = 1200 mm/min, track spacing = 2.5 mm, nozzle height = 40 mm, temp = 80 °C for DHA and 30 °C for DAN. For the analysis of rat brain sections, DHA matrix solution was sprayed on the tissue sections with an artistic airbrush [12]. A total of 4 mL of matrix solution was sprayed to coat an entire rat brain section.
Lipid Assignment
Assignment of lipid species was based upon accurate mass with mass error ± 2 ppm in positive ion mode with a m/z range of 700–900 and with mass error ± 6 ppm in negative ion mode with a m/z range of 1400–2300. Mass peaks for glycerophosphocholine (PC) were labeled so that species equal the total length and number of both radyl chains with a representing diacyl species and p representing plasmalogen species. For example, a mass peak labeled as [PC 32a:0+H]+ includes all possible structural isomers ([PC 16:0/16:0+H]+, [PC 14:0/18:0+H]+, etc.). The mass peaks for gangliosides were labeled so that species assignments correspond to the length and number of double bonds of the acyl chain and the sphingoid base with d representing a 1,3-dihydroxy sphingoid base.
The major drawback to using DHA as a matrix for lipid imaging is its volatility under low-pressure conditions. To demonstrate the advantages of an AP-MALDI source for volatile matrices, like DHA, serial sections of mouse cerebellum were analyzed at AP (760 Torr) and IP (0.07 Torr) at varying matrix thicknesses (e.g., number of matrix layers sprayed onto the tissue section). For these experiments, the AP-MALDI source was coupled to a Q Exactive mass spectrometer, while the IP experiments were conducted on a MALDI Orbitrap XL. The HTX sprayer was used to coat the tissue sections to ensure reproducibility from section to section and to control the amount of matrix coated onto the sections. The experimental parameters for these MSI runs were as follows: 50-μm pixel size, m/z range 700–900 in positive ion mode, and FT resolution setting of 140,000 for the Q Exactive and 60,000 for the Orbitrap XL. The higher resolution was used on the Q Exactive so that the scan time would more closely match the scan time on the Orbitrap XL. Total time of acquisition varied from 4 to 6 h for the samples. Initial MSI experiments were conducted on tissue sections that were coated with four layers of DHA matrix (0.0013 mg deposited per mm2 of tissue) and analyzed under both AP and IP conditions. Figure 1 shows the results of these experiments. Initially, there is enough matrix to get excellent lipid signal and generate high-quality images showing the cell layers of the cerebellum. This trend continues for the analysis at AP. However, at IP, the signal quickly decreases as the matrix sublimates. To overcome this, higher amount of matrix (that is, more layers) was sprayed onto the tissue sections. A matrix coverage of 0.0333 mg/mm2 was found to be sufficient for the analysis time (~ 4.5 h) for the sample at 0.07 Torr. However, this would change if the analysis time increased due to the size of the sample or an increase in lateral resolution. Additionally, the image quality decreased at a matrix density of 0.0333 mg/mm2 and showed several pixels of no signal. This was most likely due to the AmSulf in the matrix solution forming uneven clusters on the surface of the tissue section. However, it could also just reflect the challenges of adding thicker coatings of matrix. This step was only necessary to increase the lifetime of the matrix under vacuum. This is a major advantage of an AP-MALDI source because the amount of matrix added in this case is only based upon efficient ionization of analytes not upon sublimation rate. Thus, the use of an AP-MALDI source greatly enhances the flexibility and usefulness of volatile matrices, such as DHA. Furthermore, the sample planning/preparation is much easier at AP conditions when compared to lower pressure (high vacuum) conditions for volatile matrices because parameters such as matrix thickness and instrumental settings (mass resolution and spatial resolution) do not have to be adjusted due to matrix sublimation.
MALDI-MSI of serial mouse cerebellum sections with varying DHA matrix coating at AP (760 Torr) and IP (0.07 Torr) pressure. An average mass spectrum in the m/z range 780–840 and a combination plot (red/green/blue false color image) for three PC species, [PC 36a:1+H]+, 788.6164 Da (blue); [PC 38a:6+H]+, 806.5694 Da (red); [PC 40a:6+H]+, 834.6007 Da (green) are shown at different source pressures and matrix density. The number of pixels and pixels/s for the MSI runs shown in this figure are as follows: AP-MALDI—4 layers = 28,000 pixels, 1.59 pixels/s, 578 laser shots/pixel, number of pixels representing tissue image = 12,355 pixels; IP-MALDI—4 layers = 18,725 pixels, 0.98 pixels/s; IP-MALDI—20 layers = 15,617 pixels, 0.88 pixels/s; IP-MALDI—50 layers = 15,423 pixels, 1.06 pixels/s; IP-MALDI—100 layers = 16,005 pixels, 1.06 pixels/s
Gangliosides are complex glycosphingolipids that contain one or more negatively charged sialic acids (SA) and have been implicated in brain development, neuritogenesis, memory formation, synaptic transmission, and aging [32]. Disruptions in ganglioside metabolism has been linked to GM1 and GM2 gangliosidosis, Niemann-Pick C, and Gaucher disease types II and III [33], and gangliosides play a role in Alzheimer's disease and Guillain-Barré syndrome [34]. The main gangliosides in the central nervous system of higher vertebrates are monosialogangliosides (GM1), disialogangliosides (GD1 with two structural isomers: GD1a, GD1b), and trisialogangliosides (GT1b), which account for approximately 80–90% of the total gangliosides [35]; of these GD1s are the most abundant ganglioside species in the mammalian brain. GD1s consist of two structural isomers (see Fig. 2), GD1a in which the two sialic acids are attached, one to the terminal galactose residue and one to the first galactose residue in the oligosaccharide head, and GD1b where the two sialic acids are attached to the first galactose residue in the oligosaccharide head. Analyses of gangliosides, especially mixtures such as tissue sections, are difficult due to in source fragmentation, in which GD1 and GT1 species generate GM1 species through the loss of sialic acid [36, 37]. A previous study has shown that by increasing the pressure in the MALDI ion source, you can reduce this fragmentation and loss of sialic acid in gangliosides [38]. Figure 2 contains mass spectra of GD1a and GD1b standards with DHA matrix in negative ion mode with MALDI sources at AP and IP conditions. To minimize fragmentation in these experiments, the laser fluence was tuned to just above (~ 10–15%) MALDI signal threshold, yet ions were detected. The two major species that were observed for both GD1a and GD1b standards had ceramide backbones of d36:1 and d38:1. The d36:1 species was more abundant for GD1a while d38:1 was more abundant for GD1b. For both GD1a and GD1b, the amount of fragmentation was greatly reduced at atmospheric pressure. Additionally, GD1a and GD1b can be easily distinguished based upon the amount and type of fragmentation. This result has been observed previously [39]. Overall, GD1a species were more fragile compared to GD1b species and its major fragment corresponded to the loss of a sialic acid. The most distinguishing fragments for GD1b species was the [M–H2O–H]− mass peaks, which were either not detected or only observed at trace levels for GD1a species. The higher water loss from GD1b compared to GD1a has been observed previously in MALDI-MS studies [40, 41]. Furthermore, lactonization of GD1b under acidic conditions (DHA matrix solution is acidic) is known to occur and is the most likely reason for the appearance of the intense [M–H2O–H]− mass peak observed for GD1b [40, 42].
MALDI mass spectra of GD1a and GD1b standards with DHA matrix in negative ion mode with an AP and IP source. Mass spectra are the average of ten scans
In order to test the AP-MALDI source and DHA matrix further, we conducted additional experiments where we compared the metastable fragmentation of GD1a and GD1b for DHA versus two other MALDI matrices, 9-AA and DAN. 9-AA [43] and DAN [44] were selected because they have been used in previous MSI studies of gangliosides. To compare the amount of fragmentation of gangliosides for each matrix, we collected mass spectra at low laser fluence (~ 10–15% above threshold) and high laser fluence (twofold above threshold). Figure 3 shows graphs that plot the signal intensity for three major fragment peaks ([M–SA–H]−, [M–CO2–H]−, [M–H2O–H]−) normalized by the signal intensity of the corresponding [M–H]− parent peak for the ganglioside species. As expected, as the fluence was increased, the degree of fragmentation increased for all three of the matrices tested. DHA showed the least amount of fragmentation for both GD1a and GD1b. Additionally for GD1b, DHA produced the highest percentage of the [M–H2O–H]− mass peak, which as we noted earlier is a distinguishing peak for GD1b compared to GD1a. Based upon this result, we used DHA matrix to image this GD1b-specific peak in the studies described below.
Bar graphs showing the signal intensity for fragment peaks ([M–SA–H]−, [M–CO2–H]−, [M–H2O–H]−) produced by GD1a and GD1b standards at low and high laser fluence. The signal intensity of the fragment peaks was normalized against the intact [M–H]− mass peak for the GD1 parent species. Values graphed are the mean of three mass spectra (five mass scans) and the error bars represent one standard deviation. SA, sialic acid
Next, MSI analysis was conducted on serial mouse coronal sections (− 1.94 mm re: bregma, Franklin and Paxinos mouse atlas) with DHA and DAN with the AP-MALDI source on the Q Exactive MS instrument. The experimental parameters for the MSI runs were as follows: spatial resolution of 50 μm for DHA matrix and 60 μm for DAN matrix, m/z range 1400–2300 in negative ion mode, and FT resolution setting of 140,000. Figure 4a shows the summed average mass spectra for the whole tissue section with DHA and DAN matrix. For DHA matrix, six ganglioside species (GM1 d36:1, GM1 d38:1, GD1 d36:1, GD1 d38:1, GT1 d36:1, GT1 d38:1) were detected as [M–H]− mass peaks and imaged. Four ganglioside species (GM1 d36:1, GM1 d38:1, GD1 d36:1, GD1 d38:1) were recorded using DAN as a matrix. All species were detected as [M–H]− mass peaks. However, the dominant mass peaks associated with GD1 species were [M+Na–2H]− and [M+K–2H]−. GT1 species did not have sufficient ion counts to produce images with DAN. Comparing the average mass spectra for the whole tissue section for DHA versus DAN clearly shows the high amount of fragmentation of GD1 and GT1 species that is observed with DAN. The spectrum produced using DAN is dominated by GM1 and only minor mass peaks are observed for GD1, while the spectrum recorded using DHA has strong mass peaks observed for all three major brain gangliosides in the following order GD1 > GM1 > GT1. A previous study found the abundance of mouse brain gangliosides as follows: GD1 ~ 49%, GM1 ~ 23%, GT1 ~ 21% [45]. Clearly, a significant amount of the GM1 that was detected using DAN matrix is from GD1 and GT1 fragmentation and not from native GM1 species. All the images and ganglioside assignments obtained for theses MSI runs are supplied in the Supplemental Data section. MSn analysis was also conducted on the gangliosides species to confirm their assignments and this data is included in the Supplemental Data section.
(a) Average mass spectra for brain tissue sections with DHA and DAN matrix in the m/z range 1500–2200. (b) Combo plot (red/green false color image) of [GM1 d36:1–H]− (1544, red) and [GD1 d36:1–H]− (1835, green) with DHA matrix and combo plot (red/green false color image) of [GM1 d36:1–H]− (1544, red) and [GD1 d36:1 + K–2H]− (1873, green) with DAN matrix. (c) Average mass spectra of areas in the cortex (Cx) and corpus callosum (Cc) region with DHA and DAN matrix. The number of pixels and pixels/s for the MSI runs shown in this figure are as follows: AP-MALDI with DHA matrix = 59,800 pixels, 1.73 pixels/s, 531 laser shots/ pixel, number of pixels representing tissue image = 19,499 pixels; AP-MALDI with DAN matrix = 28,800 pixels, 1.75 pixels/s, 545 laser shots/pixel, number of pixels representing tissue image = 14,309 pixels
GM1 species can be quite difficult to image due to the fragmentation of GD1 and GT1, which can produce erroneous results where GM1 appears to have the same distribution in the brain as GD1 species, i.e., high amounts in gray matter with little to none observed in white matter. Previous studies using immunostaining techniques have shown GM1 to be highly localized in white matter regions throughout the brain [46,47,48]. Figure 4b contains two combo plots (red/green false color images): one from the DHA MSI run that combines images of [GM1 d36:1–H]−, 1544.861, red and [GD1 d36:1–H]−, 1835.959, green; and the second from the DAN MSI run that combines images of [GM1 d36:1–H]−, 1544.869, red and [GD1 d36:1+K–2H]−, 1873.921, green. Despite the high amount of fragmentation observed with DAN, both matrices detected strong GM1 signal in white matter regions such as the corpus callosum (Cc). However, when compared to GD1, the signal for GM1 when using DAN was still very high in gray matter regions such as the cortex (Cx). Thus, the combo plot image using DHA matrix shows the expected distribution for GD1 and GM1 while the combo plot image using DAN matrix is incorrect when looking at the gray matter regions. Figure 4c contains average mass spectra from the Cx and Cc regions of the brain and confirms the results observed in Fig. 4b.
As noted above, when using DHA matrix, we were able to easily distinguish GD1a and GD1b isomers based upon the presence of the [M–H2O–H]− peak observed with GD1b. Next, a MSI run with the AP-MALDI source on the LTQ Orbitrap Velos was set up to maximize the fragmentation of GD1 species to yield the [M–H2O–H]− peak. The experimental parameters for the MSI runs were as follows: 60 μm pixel size, m/z range 1000–3000 in negative ion mode, and FT resolution setting of 60,000. Figure 5(A) contains the summed mass spectrum in the m/z range of GD1 for a MALDI imaging run on coronal rat brain (− 5.88 mm re: bregma, Paxinos and Watson rat atlas) in negative ion mode with DHA matrix. Over this narrow m/z range, images were obtained for five mass peaks corresponding to GD1 gangliosides ([GD1 d36:1–CO2–H]−, [GD1 d36:1–H2O–H]−, [GD1 d36:1–H]−, [GD1 d38:1–H2O–H]−, [GD1 d38:1–H]−). Similar MS images have been obtained for [GD1 d36:1–H]− and [GD1 d38:1–H]− with DHA matrix [12]. These images reflect the combined distribution of GD1a and GD1b isomers that are both known to be present in the brain. However, based on the parameters of our AP-MALDI source and matrix solution, the MS images for [GD1 d36:1–H2O–H]− and [GD1 d38:1–H2O–H]− should almost exclusively reflect the distribution of the GD1b isomer. Figure 5(B) contains overlay combo plots (red/green false color image) for the MS images of [GD1 d36:1–H2O–H]− + [GD1 d36:1–H]− and [GD1 d38:1–H2O–H]− + [GD1 d38:1–H]−. These combo plots show the [M–H2O–H]− mass peaks, mostly representing the GD1b isomer, to be more concentrated in the brain stem and periaqueductal gray area compared to the [M–H]− mass peaks, that mostly represents the GD1a isomer. Thus, our results suggest that GD1b is present in higher amounts than GD1a in the brain stem and periaqueductal gray area. This result agrees with a previous study using immunostaining techniques, in which GD1a was shown to be largely absent in the brain stem region [48]. The possibility to image GD1b separately is important because the GD1 isomers are produced by different biosynthetic pathways with GD1a being a-series and GD1b being b-series [49]. Thus, observed difference and changes, especially in disease models, may represent changes in one specific biosynthetic pathway. Recently, it has been suggested that changes to the fatty acid chain length of GD1b gangliosides affect Alzheimer amyloid deposition in certain brain regions, while no changes were observed for GD1a gangliosides [50].
(A) Average mass spectra for brain tissue section and MALDI images for five major mass peaks in the GD1 m/z range. (B) Two combo plots (red/green false color images) for [GD1 d36:1–H2O–H]− (red) + [GD1 d36:1–H]− (green) and [GD1 d38:1–H2O–H]− (red) and [GD1 d38:1–H]− (green). Abbreviations for anatomical regions: Aq, aqueduct; BrSt, brain stem; Cc, corpus callosum; Cx, cerebral cortex; IP, interpeduncular nucleus; PAG, periaqueductal gray; SN, substantia nigra; SuG, superficial gray. The number of pixels and pixels/s for the MSI run shown in this figure are as follows: 41,250 pixels, 0.48 pixels/s, 2020 laser shots/pixel
The coupling of an AP-MALDI source with an Orbitrap mass spectrometer eliminated the problem of DHA matrix sublimation that prevented completing long MSI runs in vacuum, intermediate, and low-pressure MALDI sources. The combination of the AP-MALDI source with DHA matrix significantly reduced the amount of fragmentation observed for gangliosides when compared to other MALDI matrices used with any MALDI source and to DHA used with an intermediate-pressure MALDI source. However, it was also demonstrated that by increasing the metastable fragments, it was possible to distinguish GD1a and GD1b isomers in our studies. The mapping of the metastable fragment of the loss of water from mostly GD1b ganglioside permitted the comparison of its localization in the brain compared to the [M–H]− GD1 mass peak that consists mostly of GD1a isomers.
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Potocnik, N.O., Porta, T., Becker, M., Heeren, R.M.A., Ellis, S.R.: Use of advantageous, volatile matrices enabled by next-generation high-speed matrix-assisted laser desorption/ionization time-of-flight imaging employing a scanning laser beam. Rapid Commun. Mass Spectrom. 29, 2195–2203 (2015)
Laiko, V.V., Baldwin, M.A., Burlingame, A.L.: Atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry. Anal. Chem. 72, 652–657 (2000)
Laiko, V.V., Moyer, S.C., Cotter, R.J.: Atmospheric pressure MALDI/ion trap mass spectrometry. Anal. Chem. 72, 5239–5243 (2000)
Doroshenko, V.M., Laiko, V.V., Taranenko, N.I., Berkout, V.D., Lee, H.S.: Recent developments in atmospheric pressure MALDI mass spectrometry. Int. J. Mass Spectrom. 221, 39–58 (2002)
Gallicia, M.C., Vertes, A., Callahan, J.H.: Atmospheric pressure matrix-assisted laser desorption/ionization in transmission geometry. Anal. Chem. 74, 1891–1895 (2002)
Tan, P.T., Laiko, V.V., Doroshenko, V.M.: Atmospheric pressure MALDI with pulsed dynamic focusing for high-efficiency transmission of ions into a mass spectrometer. Anal. Chem. 76, 2462–2469 (2004)
Gabelica, V., Schulz, E., Karas, M.: Internal energy build-up in matrix-assisted laser desorption/ionization. J. Mass Spectrom. 39, 579–593 (2004)
Schulz, E., Karas, M., Rosu, F., Gabelica, V.: Influence of the matrix on analyte fragmentation in atmospheric pressure MALDI. J. Am. Soc. Mass Spectrom. 17, 1005–1013 (2006)
Li, Y., Shrestha, B., Vertes, A.: Atmospheric pressure molecular imaging by infrared MALDI mass spectrometry. Anal. Chem. 79, 523–532 (2007)
Sundaram, A.K., Oktem, B., Razumovskaya, J., Jackson, S.N., Woods, A.S., Doroshenko, V.M.: In: Ivanov, A.R., Lazarev, A.V. (eds.) Sample Preparation in Biological Mass Spectrometry, p. 749. Springer Science, New York, NY (2011)
Khalil, S.M., Rompp, A., Pretzel, J., Becker, K., Spengler, B.: Phospholipid topography of whole-body sections of the Anopheles stephensi mosquito, characterized by high-resolution atmospheric-pressure scanning microprobe matrix-assisted laser desorption/ionization mass spectrometry imaging. Anal. Chem. 87, 11309–11316 (2015)
Rompp, A., Both, J.-P., Brunelle, A., Heeren, R.M.A., Laprevote, O.B., Prideaux, B., Seyer, A., Spengler, B., Stoeckli, M., Smith, D.F.: Mass spectrometry imaging of biological tissue: an approach for multicenter studies. Anal. Bioanal. Chem. 407, 2329–2335 (2015)
Kompauer, M., Heiles, S., Spengler, B.: Atmospheric pressure MALDI mass spectrometry imaging of tissues and cells at 1.4 μm lateral resolution. Nat. Methods. 14, 90–96 (2017)
Sonnino, S., Chigorno, V.: Ganglioside molecular species containing C18- and C20-sphingosine in mammalian nervous tissues and neuronal cell cultures. Biochim. Biophys. Acta. 1469, 63–77 (2000)
Kolter, T., Sandhoff, K.: Principles of lysosomal membrane digestion: stimulation of sphingolipid degradation by sphingo-lipid activator proteins and anionic lysosomal lipids. Annu. Rev. Cell Dev. Biol. 21, 81–103 (2005)
Ariga, T., McDonald, M.P., Yu, R.K.: Role of ganglioside metabolism in the pathogenesis of Alzheimer's disease-a review. J. Lipid Res. 2008, 1157–1175 (2008)
Schwarz, A., Futerman, A.H.: The localization of gangliosides in neurons of the central nervous system: the use of anti-ganglioside antibodies. Biochim. Biophys. Acta. 1286, 247–267 (1996)
Costello, C.E., Juhasz, P., Perreault, H.: New mass spectral approaches to ganglioside structure determinations. Prog. Brain Res. 101, 45–61 (1994)
Harvey, D.J.: Matrix-assisted laser desorption/ionization mass spectrometry of carbohydrates. Mass Spectrom. Rev. 18, 349–450 (1999)
O'Connor, P.B., Mirgorodskaya, E., Costello, C.E.: High pressure matrix-assisted laser desorption/ionization Fournier transform mass spectrometry for minimization of ganglioside fragmentation. J. Am. Soc. Mass Spectrom. 13, 402–407 (2002)
Juhasz, P., Costello, C.E.: Matrix-assisted laser desorption ionization time-of-flight mass spectrometry of underivatized and permethylated gangliosides. J. Am. Soc. Mass Spectrom. 3, 785–796 (1992)
Ivleva, V.B., Sapp, L.M., O'Connor, P.B., Costello, C.E.: Ganglioside analysis by thin-layer chromatography matrix-assisted laser desorption/ionization orthogonal time-of-flight mass spectrometry. J. Am. Soc. Mass Spectrom. 16, 1552–1560 (2005)
Ito, E., Tominaga, A., Waki, H., Miseki, K., Tomioka, A., Nakajima, K., Kakehi, K., Suzuki, M., Taniguchi, N., Suzuki, A.: Structural characterization of monosialo-, disialo-, and trisialo-gangliosides by negative ion AP-MALDI-QIT-TOF mass spectrometry with MSn switching. Neurochem. Res. 37, 1315–1324 (2012)
Bassi, R., Riboni, L., Sonnino, S., Tettamanti, G.: Lactonization of GD1b ganglioside under acidic conditions. Carbohydr. Res. 193, 141–146 (1989)
Ermini, L., Morganti, E., Post, A., Yeganeh, B., Caniggia, I., Leadley, M., Faria, C.C., Rutka, J.T., Post, M.: Imaging mass spectrometry identifies prognostic ganglioside species in rodent intracranial transplants of glioma and medulloblastoma. PLoS One. 12, e0176254 (2017)
Caughlin, S., Park, D.H., Yeung, K.K.-C., Cechetto, D.F., Whitehead, S.N.: Sublimation of DAN matrix for the detection and visualization of gangliosides in rat brain tissue for MALDI imaging mass spectrometry. J. Vis. Exp. 121, e55254 (2017)
Zarei, M., Bindila, L., Souady, J., Dreisewerd, K., Berkenkamp, S., Muthing, J., Peter-Katalinic, J.: A sialylation study of mouse brain gangliosides by MALDI a-TOF and o-TOF mass spectrometry. J. Mass Spectrom. 43, 716–725 (2008)
Kotani, M., Kawashima, I., Ozawa, H., Terashima, T., Tai, T.: Differential distribution of major gangliosides in rat central nervous system detected by specific monoclonal antibodies. Glycobiology. 3, 137–146 (1993)
Heffer-Lauc, M., Lauc, G., Nimrichter, L., Fromholt, S.E., Schnaar, R.L.: Membrane redistribution of gangliosides and glycosylphosphatidylinositol-anchored proteins in brain tissue sections under conditions of lipid raft isolation. Biochim. Biophys. Acta. 1686, 200–208 (2005)
Vajn, K., Viljetic, B., Degmecic, I.V., Schnaar, R.L., Heffer, M.: Differential distribution of major brain gangliosides in the adult mouse central nervous system. PLoS One. 8, e75720 (2013)
Merrill Jr., A.H.: Sphingolipid and glycosphingolipid metabolic pathways in the era of sphingolipidomics. Chem. Rev. 111, 6387–6422 (2011)
Oikawa, N., Matsubara, T., Fukuda, R., Yasumori, H., Hatsuta, H., Murayama, S., Sato, T., Suzuki, A., Yanagisawa, K.: Imbalance in fatty-acid-chain length of gangliosides triggers Alzheimer amyloid deposition in the precuneus. PLoS One. 10, e.0121356 (2015)
This research was supported in part by the Intramural Research Program of the National Institute on Drug Abuse, NIH.
Integrative Neuroscience, NIDA IRP, NIH, 333 Cassell Drive, Room 1119, Baltimore, MD, 21224, USA
Shelley N. Jackson, Ludovic Muller, Aurelie Roux & Amina S. Woods
Mass Tech, Inc., Columbia, MD, USA
Berk Oktem, Eugene Moskovets & Vladimir M. Doroshenko
Shelley N. Jackson
Ludovic Muller
Aurelie Roux
Berk Oktem
Eugene Moskovets
Vladimir M. Doroshenko
Amina S. Woods
Correspondence to Shelley N. Jackson.
ESM 1
(DOCX 6729 kb)
Jackson, S.N., Muller, L., Roux, A. et al. AP-MALDI Mass Spectrometry Imaging of Gangliosides Using 2,6-Dihydroxyacetophenone. J. Am. Soc. Mass Spectrom. 29, 1463–1472 (2018). https://doi.org/10.1007/s13361-018-1928-8
Revised: 02 February 2018
Issue Date: July 2018
AP-MALDI


text:
Nous allons maintenant aborder un chapitre très important pour le langage __Rust__. Ceux ayant déjà utilisé des langages fonctionnels n'y verront qu'une révision (mais ça ne fait jamais de mal après tout !).

Pour ceux qui n'ont jamais utilisé de closures, on peut les définir comme des fonctions anonymes qui capturent leur environnement.

"Une fonction "anonyme" ? Elle "capture" son environnement ?"

Ne vous inquiétez pas, vous allez très vite comprendre, prenons un exemple simple :

```Rust
let multiplication = |nombre: i32, multiplicateur: i32| nombre * multiplicateur;

println!("{}", multiplication(2, 2));
```

Pour le moment, vous vous dites sans doute qu'en fait, ce n'est qu'une fonction. Maintenant ajoutons un élément :

```Rust
let nombre = 2i32;
let multiplication = |multiplicateur: i32| nombre * multiplicateur;

println!("{}", multiplication(2));
```

Là je pense que vous vous demandez comment il fait pour trouver la variable __nombre__ puisqu'elle n'est pas dans le scope de la "fonction". Comme je vous l'ai dit, une closure __capture__ son environnement, elle a donc accès à toutes les variables présentes __dans le scope de la fonction qui l'appelle__.

Mais à quoi ça peut bien servir ? Imaginons que vous avez une super interface graphique et que vous voulez effectuer une action lors d'un événement, disons lorsque le bouton est cliqué. Hé bien cela donnerait quelque chose dans ce genre :

```Rust
let mut bouton = Bouton::new();
let mut clicked = false;

bouton.clicked(|titre| {
    clicked = true;
    println!("On a cliqué sur le bouton {} !", titre);
});
```

Facile, non ? Cela dit, vous pourriez aussi vous servir des closures pour trier un vecteur d'objets ou d'autres choses similaires.

Si jamais vous souhaitez écrire une fonction recevant une closure en paramètre, voici à quoi cela va ressembler :

```Rust
fn fonction_avec_closure<F>(closure: F) -> i32
    where F: Fn(i32) -> i32
{
    closure(1)
}
```

Ici, la closure prend un [__i32__] comme paramètre et renvoie un [__i32__]. Vous remarquerez que la syntaxe est proche de celle d'une fonction générique, la seule différence venant du mot-clé __where__ qui permet de définir à quoi doit ressembler la closure. À noter qu'on aurait aussi pu écrire la fonction de cette façon :

```Rust
fn fonction_avec_closure<F: Fn(i32) -> i32>(closure: F) -> i32 {
    closure(1)
}

```

Nous avons donc vu les bases des closures. C'est une partie importante, je vous conseille donc de bien vous entraîner dessus jusqu'à être sûr de bien les maîtriser !

Après ça, il est temps d'attaquer un chapitre un peu plus "tranquille".

[__i32__]: https://doc.rust-lang.org/stable/std/primitive.i32.html

redpajama_set_name: RedPajamaGithub


Hey everyone we are nearing the end of this Disney series which is a bitter sweet experience for me. Bitter because obviously I want to keep producing posts that are solely Disney related but sweet because it means I can post different posts for you guys. So for today's blog post I am going to be sharing with you various youtubers who either solely upload videos relating to Disney or have done in the past.
If you are like me and are obsessed with anything Disney related then I am sure you would have probably watched a number of Disney related Vlogs or videos. I really love the concept of a vlog as it makes you feel like you are actually there. I have discovered some amazing Disney vloggers who I will continue to watch and re-watch all the time.
The first youtuber I'm going to be sharing with you is Adam Ruck who is a youtuber that posts videos mostly related to Disney. I especially like his Disney vlogs however I do enjoy his other videos too. He has great enthusiasm and I'd say I find the vlogs like a show as he really makes each and everyone entertaining. I definitely think he should be recognised more and should have way more subscribers. He's also from Texas which means he has a pretty cool accent make sure to go & check Adam out!!
The second youtuber I'm going to introduce you two is the lovely Em. First of all Em is a really pretty girl who loves Disney just as much as me if not even more. I really liked Em's Disney vlogs as they were so jammed pack I love how she included her family in them and she just seemed like a really down to earth girl. What I also like is that she's herself on the vlogs she's not trying to be something she's not which some youtubers do. So after watching a few of her vlogs I just had to subscribe to her and you should too!
Faith only started her Youtube about a month ago however she's already doing really well on it. I can't quite remember how I discovered her vlogs but I'm so happy that I did. They're quite long ones which are great for a Sunday afternoon or while you're cooking. I also really liked seeing the relationship that Faith has with her family especially with her dad it was just such a lovely thing to see. Faith's also from New York so she has a cool accent which I'd love to master (no really).
This particular youtube channel is run by the lovely Jen who also has many other youtube channels as well as a blog. However recently Jen has decided to combine all of her channels and post just on her main one which will include aspects from these other channels which you can find here. Although Jen won't be uploading any videos to this channel any more you can still go back and watch all of the videos that she has posted. I really like her Disney vlogs in particular because it shows that people who are older can have fun and enjoy the Disney parks too. Jen's vlogs provide a different perspective of going to Disney world which I like as it makes them more interesting.
The second to last vloggers that I am going to introduce you two are this lovely family who consist of Phil, Emma and Ben. They have many vlogs which are taken in Disney World each time they go they usually stay in a different hotel which I like as it gives you the opportunity as the viewer to see what each one is like. I also like that they go at different times of the year such as Christmas and New Years. Their vlogs are always full of everything Disney whether it's the food, the rides or the music which is included in the vlogs. I really like watching their vlogs as they seem like such a down to earth family so I'm happy that I discovered them.
The final Disney youtubers that I am going to sharing with you today are these two lovely ladies Megan and Rhiannon who are sisters from New Zealand. As you can probably tell by the name of their youtube channel they are pretty big fan's of Disney. Although they don't go to Disney that much I really like their vlogs when they do as they usually consist of staying on site within Disney world as well as going on one of the Disney cruises which I've always wanted to go on as well.
"7 Days of Disney | Day 5 Disney Pins"
"7 Days of Disney | Day 7 – 3 Reasons why I love Disney"


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Guides successful completion of major engineering activities related to the development and certification of the Network Security Systems and Information Management System (IMS) on the aircraft.
Prepares and reviews necessary documents such as, but not limited to: system level requirements, equipment specifications, source control drawings, electrical interface control documents, wiring diagrams, certification plans, ground and flight test procedures, etc.
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redpajama_set_name: RedPajamaCommonCrawl


The Scientologist contains critiques, an ongoing analysis, interesting and informative articles on the subject of the scientology religion and philosophy of Lafayette Ron Hubbard and related subjects.
This site is operated by independent scientologists and is neither endorsed by, nor associated or affiliated with the corporate company known as the Church of Spiritual Technology or its companies & churches such as the Religious Technology Centre, Church of Scientology International or any other of its affiliates, corporations, management organisations. groups or members.
"If you want peace, you won't get it with violence."
Imagine a world in which everyone understood they were a spiritual being who occupied a succession of bodies. They did not get to choose the body they occupy and each time it was 'pot luck' if they got a body of a particular color, race or culture each time.
Imagine if they were aware of this. How would that affect racism? How would that affect attitudes to colour and race? Would the white supremist be so intolerant knowing that in some past lives he was NOT white and quite possible in his next life he would not be white? Would he not BE a white supremist after all? How about the attitude of a black or brown person? Or Arab, or Jew, or Palestinian?
"I'm not afraid of death because I don't believe in it. It's just getting out of one car, and into another."
Each car started out new and got old. Each car eventually had parts wear out and had to be replaced. Eventually each car died and had to be replaced. In each car I opened the door got in and sat in the seat. When I was travelling people saw the car, not me.
When I was driving a Mitsubishi I did not complain about the Toyota or Nissan or their drivers. I did not hold my nose in the air when I saw a Mercedes. Nor did I look in disgust when I saw a beat up old Datsun. And as for the Rolls Royce, well! It is a car I would LIKE to have regardless of the color or age. But the price tag is too high for me.
But no one goes to war over a car. No one drops bombs on cars that are of a different color or comfort level or size. No one goes into a parking lot and shoots up all the cars there or bombs themselves into oblivion because they disagree with the cars present. Just as no one singles out all the ginger headed people for extermination or all those over certain weight or age. No. That would be insanity.
Understanding that one is a being who moves from one body to another would totally change one's viewpoint on life. Many people, I am sure, would poo poo this idea as they consider themselves a body. This is the present culture in the western world. 'One is a brain, one is a body. A piece of meat.' This, of course does not reconcile with the religions of ther world which, for the most part, consider an individual a spiritual being of one sort or another occupying a body. One can test this quite easily (we will come to that later) and for those that appreciate they are a being and not a physical body the next step is not a hard one to take. It is also possible to take a person back to re-experience a life before the present one. This has been done so many times that it is now an established fact. Apart from establishing the authenticity of past lives it gives the individual a reality of what they are as distinct to the culture of being a meat body the present society fosters one people. But you cannot keep a good person down and probably one of the simplest test an individual can make (and this would apply to around 50 percent of the population) is this.
Be three feet back of your head.
Now if you can do that, no further 'proof' is required. For those that have difficulty with this further actions can be done but they will take more time and it them becomes a case of how willing is one to undergo such activities to find out something one has a disagreement with to find the truth. The truth is there. You just have to look.
"Your potentialities are a great deal better than anyone ever permitted you to believe."
The western media harps on and focuses on the person being a body. A brain with synapses and electrical currents being the thinking processes of an individual. Nothing could be further from the truth. But look. If you convince a population they are a brain, they are much easier to manage and control. People who know they are like the will o' the wisp*, cannot be controlled. Only their bodies can be controlled. Where's your ID?' 'Show me your passport.' A being does not have or need a passport. Only a body does. There are no borders or barriers for a being. How would the 'authorities' feel if they had no control over where people went without restrictions? How would it be if someone convinced you that you were a car? And had to behave a certain way, was restricted in your movements, had to be licenced to be on the road, had to justify your driving or choice of car at every turn? Sound familiar?
All the violence, prejudice, narrow thinking based on the culture of the 'one lifers' does not detract from the truth. You are a being who is a whole lot better than a car. You are the driver of the car. You cannot measure a dog by his biscuits and you cannot measure a man by the car he inhabits and drives.
Time to wake up! Regardless of what your beliefs are, Christian, Islamic, Buddhist or any other, you are a spiritual being. And for some people, it is just three steps to finding out who you are. Others may have a road to travel. But it is a road to freedom and you will never be the same again.
*Will-o'-the-wisp - Wikipedia. In folklore, a will-o-wisp is an atmospheric ghost light seen by travellers at night, especially over bogs, swamps or marshes. It goes where it wants when it wants.


text:
Prof. Dr. Vasilis Ntziachristos
Chair of Biological Imaging, School of Medicine, Technical University of Munich
bioimaging.translatum@tum.de
Professor of Medicine, Professor of Electrical Engineering and Director of the Chair for Biological Imaging (CBI) at the Technical University of Munich, Director of the Institute for Biological and Medical Imaging (IBMI) at Helmholtz Munich and Director of Bioengineering at the Helmholtz Pioneer Campus.
Prof. Ntziachristos has received the Diploma in Electrical Engineering and Computer Science from the Aristotle University of Thessaloniki, Greece and the M.Sc and Ph.D. degrees in Bioengineering from the University of Pennsylvania in Philadelphia PA. Prior to his current appointment, he was faculty at Harvard University and the Massachusetts General Hospital. Professor Ntziachristos is the founder of the journal Photoacoustics, regularly Chairs in international meetings and councils and has received numerous awards and distinctions, including the Karl Heinz Beckurts prize (2021), the Chaire Blaise Pascal (2019) from the Region Ile-de-France, the Gold Medal from the Society for Molecular Imaging (2015), the Gottfried Leibnitz prize from the German Research Foundation (2013), the Erwin Schrödinger Award (2012) and was named one of the world's top innovators by the Massachusetts Institute of Technology (MIT) Technology Review in 2004.


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Fan-owned pages on YouTube are parading obscene, cartoon-like characters and packaging them as 'real supporters', and they need to be kicked off the internet.
Social media platforms are centred on instant, fleeting gratification. One of the biggest, Arsenal Fan TV (now known as AFTV media due to the club wanting nothing to do with them) is close to reaching 1 million subscribers.
That's one million Spurs, Liverpool and Chelsea fans that swarm to the comments sections to ridicule these 'personalities' when Arsenal lose.
They have made z-list 'celebrities' out of the average supporter and the formula couldn't be simpler: the more outrageous the personality and rhetoric, the more successful they are.
In the internet world, numbers are everything, so AFTV cherry-pick their most controversial pawns and prod them with a stick. They wind up the fans at their most vulnerable – a heavy defeat – until they embark on another angry tirade, which draws in millions of viewers.
It's easy money for these guys; all it takes is a camera, and microphone and they're making pure profit from the advertisement revenue on their clickbait videos.
Certain fans on AFTV led the witch hunt to get Arsene Wenger sacked, with constant tirades, and even attacking the character of the man, which was then broadcasted to millions of people. Players have come out and said they watch it, so there's no reason why Arsene or members of the board would've watched. Surely the upper echelons would've realised how damaging it is to the brand when these videos go viral. Some have already laid into Unai Emery – before his first season is even over.
Unfortunately, we have to live in a world where some of these people are going on to forge careers out of fan channel appearances. One loudmouthed buffoon who screams "BOOM" after every other word, landed roles in Storage Hunters UK and Celebrity Big Brother.
Granted the term 'celebrity' is used as loosely as his grip on reality.
IS IT REALLY FREEDOM OF SPEECH?
The fan channels argue that they're giving the regular fan a voice, but at what cost? If all this supposed 'voice' does is open the fan contributors up to swathes of ridicule, it isn't really benefitting anyone. The culture it has spawned is particularly demonising for regular fans too. Channels like AFTV give their supporters a bad name.
What they don't realise is that 'real' fans may have something more worthwhile to add to the debate, other than screaming 'BOOM' at the top of their lungs.
The landscape of football content is changing, with newspapers' revenues dropping faster than Mo Salah in the box. These 'influencers' who command armies of followers have more of a draw than some legitimate news outlets. But the way it's packaged needs to be better.
There are endless opportunities for fans to express themselves in a more civilised, constructive fashion, and that is being done by podcasters/YouTubers who don't need to attack people to succeed. To be fair to AFTV, there are instances where they sit down one-on-one with fans, away from the testosterone-fuelled environment of a football game, but these videos are far less successful. And it does feel like that is filler content until the next 'I'D RATHER PLAY SOMEONE WITH ONE LEG THAN MUSTAFI' rant releases.
But who is to blame – the business savvy fan channels that are tapping into our primitive desires to watch people making fools of themselves? Or is it just the human race – the reality TV junkies – that need their own heads checked?


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The qualification covers both safety at work in the preparation of food, healthy eating and special diets, costing and menu planning and the preparation, cooking and finishing of a wide range of commodities and dishes.
All our programmes consists of 50% academic study (kitchen practical and classroom theory) and 50% work integrated learning which takes place in leading hospitality establishments.
This exciting new programme will provide the knowledge and experience to the move effortlessly between the most basic puff pastry to complex plated desserts. Covered in the programme are cake baking, decorative cake icing, desserts and basic bread making. As well as Sugarcraft, which is the use of sugar and chocolate as a creative medium.
All our programmes' structure includes theory and practical cookery lesson, with work integrated learning spent in bakeries, restaurant or hotel patisserie kitchen, gaining knowledge and work integrated experience from the industry experts.
A Traineeship is a "work while you learn" programme, comprising of work integrated learning and theoretical study by way of distance learning through International Hotel School Online learning platform and supported by tutorials. Trainees will be required to have access to a computer with internet access.
Trainees spend the duration of their studies in a work environment, receiving on the job training at the hospitality establishment from professionals in the workplace at the hospitality establishment where they are placed. Whilst they are there they will also be supported by Student Facilitators who will conduct visits in the workplace. Trainees are subject to the same terms and conditions as full time employees.
Our Open Days are the very best way to find out more. Come and speak to students who are doing the course, speak to lecturers and watch a cooking demo.
You are also welcome to contact us and a consultant will help you plan your journey to becoming a chef.
If you can't get to our campus right now, feel free to browse our website for all the information, or check out our Facebook Page or Instagram pics for inspiration.


At The First Fall of Snow | All things considered.
We're getting ready for the usual busy days in the run up to a new exhibition at the gallery.
Opening on Saturday 24th November "At The First Fall of Snow" features the work of both Jonny Hannah and Mark Hearld - they'll be exhibiting a range of paintings, prints and collages.
Born & bred in Dumfermline, Jonny studied illustration at Liverpool Art School and then the Royal College of Art.
unique paintings, collages and hand-painted ceramics.
We've also been working with Mark on a range of textiles - the first of which, Doveflight, is available now.
If you're anywhere near Norfolk on Saturday 24th November, pop in between 11am and 4pm for a glass of wine and a natter with Jonny and Mark.


Build spaceships and rockets with Magformers Space WOW 22Pc Set. Beginner builders can blast into space with the new astronaut character! Add antenna and wheels to create space buggy or search for extra-terrestrial life by building a space station. Great for young minds to learn about constructing cube nets, patterns, shapes and colors!


The methodology employed is based on non parametric statistics, allowing for the use of small sample sizes without impacting the high confidence intervals clients demand in their structured problem solving efforts. The TPS progressive convergence method will help to reduce the disruptions to plant operations that typical problem solving approaches generate and will save valuable engineering resources and time. TPS will provide an expert problem solver to work side by side with your resources to quickly develop and execute a strategy to converge on the root cause of your technical problem, confirm the root cause with statistical confidence, and apply countermeasures with proposed corrective actions. The statistics and sample sizes used for convergence are based on a 95% confidence interval. Confirmation testing can be performed at confidence levels higher than 95% when required. We will also provide support to leadership to ensure that the problem solving team is working effectively and efficiently. Most Statistical Engineering projects can be completed in less than 10 days and will consume less than 50 parts or final assemblies.


Wales McLelland uses SiteMax daily to organize and complete all their projects more efficiently. Construction professionals love using SiteMax to increase field productivity and save valuable time and money on the job site. Consolidate your daily field management processes with our simple and powerful construction app. So your teams can focus on what really matters, building.
"As the CSO of Fusion Projects, SiteMax has been integral in the ability to be Core Certified. The system has enabled us to have a repeatable and scalable process that has been very easy for the field to adopt."
"The team at SiteMax has met or exceeded every challenge we have presented to them in order to make their software work for us. SiteMax is highly customizable but still very user friendly and has allowed us to simplify many processes and procedures within our company. Keep up the great work!"
"SiteMax has streamlined our site paperwork into an easy digital experience that our people took well to and our clients love the real-time updates"
"SiteMax has allowed me to get my business organized with my field crews. Before using SiteMax my project information was all over the place. Now my clients can tell we are a well oiled machine "
"We have been working with SiteMax for many years and would recommend their system and service to anyone looking optimize their site workflows. We can always count on them to help us with our needs."


Sasson Ancient Art - Classical Art - Greek Art - Greek Hellenistic Bronze Herakles Roundel. Portrait of a ruler.
Missing inlayed eyes, otherwise perfect. Fine dark green patina.
Ex. German privet collection 1990's.
The bust of Herakles with the head turned to the left, adorned with a wreath of ivy leaves, with the straps falling on his shoulders. The Nemean lion skin legs are falling from the shoulders and tied in "Herakles Tie" over his chest.
The head is modeled with characteristics of a portrait, possibly of a Ptolemaic king.


BHS School Store open for business – W.A.M.P.U.S.
On Feb. 5, our very own Boonton High School opened up a school store, very out of the blue, offering items such as sweatpants and T-shirts, in a grand show of school spirit.
Under the direction of Mr. Nash and Mr. Engelberger, the store, located in the media center, was an attraction its first week, although it is questionable whether or not it will stay that way. The first two customers were students Farhana Elias, a sophomore, and Mariana Lozano, a freshman.
Farhana Elias and Mariana Lozano were the first customers at the Boonton High School school store.
Because the store opened very abruptly and without fanfare, the W.A.M.P.U.S. staff decided to do some investigating, and interviewed Mr. Nash.
According to Mr. Nash, the store came about because of several Gateway students, now graduated. Current Gateway students will run the store, open during the common lunch block, and all profits will go to purchasing more merchandise.
Mr. Nash hopes the store will foster school pride. In the future, the media center will be more akin to a coffee shop, a perfect area for studying but also enjoying your time.
With the coming of this store, new ideas are being brought to reinvent the school in many great ways, and, honestly, it's really pumping a lot of the students up. The relatively fair prices and new ideas have benefited the school and hopefully will continue to do so as we observe the rise in school spirit and new traditions.


text:
Q: Converting grouped tibble to named list I feel like there is probably a better way to do this in tidyverse than a for-loop. Start with a standard tibble/dataframe, and make a list where the name of the list elements are the unique values of one column (group_by?) and the list elements are all the values of another column.
  my_data <- tibble(list_names = c("Ford", "Chevy", "Ford", "Dodge", "Dodge", "Ford"),
                    list_values = c("Ranger", "Equinox", "F150", "Caravan", "Ram", "Explorer"))

# A tibble: 6 × 2
  list_names list_values
  <chr>      <chr>
1 Ford       Ranger
2 Chevy      Equinox
3 Ford       F150
4 Dodge      Caravan
5 Dodge      Ram
6 Ford       Explorer

This is the desired output:
  desired_output <- list(Ford = c("Ranger", "F150", "Explorer"),
       Chevy = c("Equinox"),
       Dodge = c("Caravan", "Ram"))

$Ford
[1] "Ranger"   "F150"     "Explorer"

$Chevy
[1] "Equinox"

$Dodge
[1] "Caravan" "Ram"

It can be accomplished with a for-loop but I bet there is a tidyverse function that makes it more simple/faster, etc.
  desired_output <- list()
  for(i in seq_along(my_data$list_names)) {
    entry <- my_data %>%
      filter(list_names == my_data$list_names[i]) %>%
      pull(list_values)
    desired_output[[my_data$list_names[i]]] <- entry
  }

A: We can use split
with(my_data, split(list_values,
     factor(list_names, levels = unique(list_names))))
$Ford
[1] "Ranger"   "F150"     "Explorer"

$Chevy
[1] "Equinox"

$Dodge
[1] "Caravan" "Ram"

Or with unstack
unstack(my_data, list_values ~ list_names)
$Chevy
[1] "Equinox"

$Dodge
[1] "Caravan" "Ram"

$Ford
[1] "Ranger"   "F150"     "Explorer"

A: Here is another option (though a little more verbose) using group_modify and deframe from tidyverse.
library(tidyverse)

my_data |>
  group_by(list_names)  |>
  group_modify(\(x, ...) tibble(res = list(deframe(x)))) |>
  deframe()

Or another option could be to use summarise then again use deframe:
my_data %>%
  group_by(list_names) %>%
  summarise(named_vec = list(list_values)) %>%
  deframe()

Output
$Chevy
[1] "Equinox"

$Dodge
[1] "Caravan" "Ram"

$Ford
[1] "Ranger"   "F150"     "Explorer"

Benchmark
I was curious to see what is fastest of the answers here, and definitely looks like split by @akrun is by far the fastest, followed by unstack.

bm <- microbenchmark::microbenchmark(
  akrun_split = with(my_data, split(list_values,
                                    factor(list_names, levels = unique(list_names)))),
  akrun_unstack = unstack(my_data, list_values ~ list_names),
  andrew_deframe1 = my_data |>
    group_by(list_names)  |>
    group_modify(\(x, ...) tibble(res = list(deframe(x)))) |>
    deframe(),
  andrew_deframe2 = my_data %>%
    group_by(list_names) %>%
    summarise(named_vec = list(list_values)) %>%
    deframe(),
  paulsmith = my_data %>%
    group_by(list_names) %>%
    summarise(list_values = list(list_values)) %>%
    {set_names(.$list_values, .$list_names)},
  times=1000L
)

A: Another possible solution:
library(tidyverse)

my_data <- tibble(list_names = c("Ford", "Chevy", "Ford", "Dodge", "Dodge", "Ford"),
                  list_values = c("Ranger", "Equinox", "F150", "Caravan", "Ram", "Explorer"))

my_data %>%
  group_by(list_names) %>%
  summarise(list_values = list(list_values)) %>%
  {set_names(.$list_values, .$list_names)}

#> $Chevy
#> [1] "Equinox"
#>
#> $Dodge
#> [1] "Caravan" "Ram"
#>
#> $Ford
#> [1] "Ranger"   "F150"     "Explorer"

A: Update II: Changed tibble output to vector outouput. Thanks to Jeff Parker!
Update due to Jeff Parker's comment (please see comments). I now updated the code. The issue was setting Names as unsorted, after using sort we can use setNames correctly. Then I added map to apply dplyrs select to remove first column in each dataframe:
library(dplyr)
library(purrr)

my_data %>%
  group_by(list_names) %>%
  mutate(list_values= paste(list_values, collapse = ", ")) %>%
  slice(1) %>%
  group_split() %>%
  setNames(sort(unique(my_data$list_names))) %>%
  map(., dplyr::pull, -list_names) %>%
  map(., ~str_split(.x, ", ")[[1]] )

$Chevy
[1] "Equinox"

$Dodge
[1] "Caravan" "Ram"

$Ford
[1] "Ranger"   "F150"     "Explorer"

A: Just for fun, added benchmarks with my for-loop and TarJae's answer.
bm <- microbenchmark::microbenchmark(
  akrun_split = with(my_data, split(list_values,
                                    factor(list_names, levels = unique(list_names)))),
  akrun_unstack = unstack(my_data, list_values ~ list_names),
  andrew_deframe1 = my_data |>
    group_by(list_names)  |>
    group_modify(\(x, ...) tibble(res = list(deframe(x)))) |>
    deframe(),
  andrew_deframe2 = my_data %>%
    group_by(list_names) %>%
    summarise(named_vec = list(list_values)) %>%
    deframe(),
  paulsmith = my_data %>%
    group_by(list_names) %>%
    summarise(list_values = list(list_values)) %>%
    {set_names(.$list_values, .$list_names)},
  jeffs = {
    desired_output <- list()
    for(i in seq_along(my_data$list_names)) {
      entry <- my_data %>%
        filter(list_names == my_data$list_names[i]) %>%
        pull(list_values)
      desired_output[[my_data$list_names[i]]] <- entry
    }
    desired_output},
    TarJae = my_data %>%
    group_by(list_names) %>%
    mutate(list_values= paste(list_values, collapse = ", ")) %>%
    slice(1) %>%
    group_split() %>%
    setNames(sort(unique(my_data$list_names))) %>%
    map(., dplyr::pull, -list_names) %>%
    map(., ~str_split(.x, ", ")[[1]] ),
  times=100L
)

I also ran benchmarks with a larger data set on the two fastest options (from Akrun).
library(nycflights13)
my_data <- nycflights13::flights %>%
  select(list_names = carrier, list_values = flight)

A: Another way of doing it:
library(tidyverse)

my_data %>%
  group_split(list_names) %>%
  map(~ lst(!!unique(pull(.x, list_names)) := unique(pull(.x, list_values)))) %>%
  flatten()
#> $Chevy
#> [1] "Equinox"
#>
#> $Dodge
#> [1] "Caravan" "Ram"
#>
#> $Ford
#> [1] "Ranger"   "F150"     "Explorer"

Created on 2022-01-14 by the reprex package (v2.0.1)
Data:
my_data <- tibble(
  list_names = c("Ford", "Chevy", "Ford", "Dodge", "Dodge", "Ford"),
  list_values = c("Ranger", "Equinox", "F150", "Caravan", "Ram", "Explorer")
)

redpajama_set_name: RedPajamaStackExchange


text:
HBO Orders 'The Outsider' Drama Series Based On Stephen King Novel Starring Ben Mendelsohn From Jason Bateman & MRC
Photo courtesy of CAA
HBO has given a series order to supernatural drama The Outsider, based on Stephen King's bestselling novel, with Ben Mendelsohn set to star and produce. The project, which marks Mendelsohn's return to TV following his Emmy-winning turn on Bloodline, hails from The Night Of co-creator/executive producer Richard Price, director-producer Jack Bender (Mr. Mercedes) Jason Bateman's Aggregate Films, Temple Hill Entertainment and MRC. Ozark star/director/executive producer Bateman is set to direct the first two episodes of The Outsider and I hear he may play the co-lear opposite Mendelsohn in the two-header.
In The Outsider, written by Price, a seemingly straightforward investigation into the gruesome murder of a local boy leads a seasoned cop and an unorthodox investigator to question everything they believe to be real, as an insidious supernatural force edges its way into the case. The book was published in May by Charles Scribners and Sons.
Price executive produces with Bateman and Michael Costigan via Aggregate Films, Marty Bowen for Temple Hill Entertainment, and Bender. MRC is the studio on the project, which is being produced by Bateman's Aggregate Films and Temple Hill Entertainment.
MRC optioned King's The Outsider earlier this year for a 10-episode limited series adaptation and in June tapped Richard Price to write/executive produce it and Jack Bender and Marty Bowen to exec produce. Mendelsohn has since been tapped to star and Bateman and his Aggregate Films boarded the project, which MRC developed internally with Price writing the pilot script before the package was taken out.
This marks Aggregate's second collaboration with MRC following the Netflix series Ozark, recently renewed for a third season, on which Bateman serves as director, producer and male lead. Bateman also helmed the first two episodes of that series.
REX/Shutterstock/Scribner
Australian-born Mendelsohn won a best supporting actor Emmy for his performance in Netflix series Bloodline. He was recently seen as Director Orson Krennic in Stars Wars: Rogue One and also starred in Steven Spielberg's Ready Player One. He currently headlines the Nicole Holofcener film The Land of Steady Habits for Netflix, just wrapped a role in The King for director Joel Edgerton and appears in Disney's upcoming Captain Marvel. Mendelsohn is repped by CAA in the U.S. and United Management in Australia.
Bateman's Aggregate Films in July inked a first-look deal with Netflix for film and TV. Bateman is repped by CAA, Lighthouse Management + Media and HJTH law firm.
Price co-created and executive produced HBO's acclaimed limited series The Night Of. He's currently co-executive producer on the network's period drama The Deuce. He is repped by UTA.
Bender and Bowen previously teamed on the Audience series Mr. Mercedes.
King is repped by Paradigm and Gang, Tyre, Ramer, Brown & Passman.
This article was printed from	https://deadline.com/2018/12/hbo-orders-the-outsider-drama-series-based-on-stephen-king-novel-starring-ben-mendelsohn-from-jason-bateman-mrc-1202512665/


This lovely orchid arrangement will appeal to naturalists with a glossy earth tone pot, charming succulent, and natural birch accent. Its bright white orchids lend effortless elegance, too! Place this arrangement near a window or right in the middle of a coffee table.


Known Relatives: Raphael (Brother), Leonardo (Brother), Michaelangelo (Brother), Splinter ("Father").
Donatello possesses several of the species wide adaptions that one would find in reptiles classified as turtles (chelonians).
Presumably the mutagenic ooze converted Donatello into a warm-blooded organism (unlike normal turtles).
A skilled practitioner of the Bojutsu martial arts style.
Outside of his physical attributes, Donatello possesses a genius level intellect. Despite his not receiving any formal education, he is capable of understanding complex concepts with ease (especially those within the field of software development and hardware engineering). He's also adept at repairing conventional technology when given the right materials.
Donatello uses a long sturdy bo staff in combat. Like all of his brothers, he also keeps a reserve of shuriken on his person at all times.
Like all of his brothers, Donatello appears to be a large bipedal turtle. His body is hairless, has no visible ears or nose and is a deep green colour. His hands and feet are three fingered/toed, and his eyes are jet black.
Unlike a normal turtle, his eyes point directly forward and are distinctly human in appearance (if not unusually large). Donatello is the tallest and leanest of his brothers.
Donatello has a large and pronounced shell situated in the centre of his back. It is clearly a part of his body, and extends across the entirety of his torso. The portion of the shell that covers his chest, stomach and crotch area is a light brown in colour.
With his shell protecting his modesty as well as his vitals, Donatello doesn't normally wear cloths unless he is attempting to hide his mutated features from humans.
A battered leather belt that he uses as a harness/holster for his weapons.
Light strips of cloths around his joints (wrists, elbows, knees, and ankles).
A red strip of cloth that he wears around his eyes when expecting combat. The cloth resembles a domino mask while on, but in truth does little to protect his identity.
Of the four brothers, Donatello is the quietest and most pensive. He doesn't share their love of combat. While he takes his lessons seriously, if given a choice he will spend his time working out an interesting mental puzzle or playing with a computer instead of practicing his ninjutsu.
He constantly searches for a better (and often more modern) way of dealing with the issue at hand. He embraces technology that his siblings and father would not.
Donatello scientific mind and practical manner has made him the "odd duck" of the family. The others have similar (if still varied) interests that he simply doesn't share.
Skin Armor: : Partial Armor, Can be bypassed by a Trick Shot (- 2FC).
Acrobatics (Athletics, Climbing)*: 05, Gadgetry: 07, Martial Artist: 05, Military Science (Camouflage, Danger Recognition): 08, Scientist: 07, Thief (Security Systems, Stealth): 09, Weaponry (Ninja weapons): 05, Weaponry (Staves): 06.
Familiarity (Automotive Mechanics, Japanese History, Ninja Lore), Languages (French, Japanese), Scholar (Computer Repair, Mathematics, Mechanical Engineering, Software Development).
Raphael (High), Michelangelo (High), Leonardo (High), Splinter (High), April O'Neil (High), Casey Jones (High).
Bo Staff [BODY 04, EV 04 (06 w/Martial artist)].


I love eating while watching TV, but I don't like struggling with my plate or making a mess. That is usually what happens if I balance my plate on my lap, especially if I have to use a knife. That's why if I have anything that is more than a snack, I bring out my TV table. It may not be the sturdiest thing in the world, but it does the job fine. Then I fold it up and store it away.
There are so many uses for folding tables, that it makes a lot of sense to own one. They can be used for parties, or as a temporary desk, or even as patio furniture, depending on the materials it is made of. They can even be used as permanent furniture if the rather functional look of them is not an issue for you. They are available in several different sizes and shapes.


text:
Indiana high school football state finals Class 3A preview: Brebeuf Jesuit vs. Gibson Southern
Kyle Neddenriep
As the seconds ticked down on Gibson Southern's Class 3A semistate win over Tri-West on Saturday night at Tri-West, coach Nick Hart got a little sentimental with his senior quarterback, Brady Allen.
"He has all the records in the world and he would throw them all in the trash can for a state championship," Hart said of the Purdue-bound Allen, a four-year starter for Gibson Southern. "He's talked about that since he was in sixth or seventh grade. But he reminded me as I was getting sentimental with him that I'm happy we get to end it here (at Lucas Oil Stadium) that, 'Hey, we have one more left.'"
What we learned from semistate:Brady Allen is the real deal and more
Finals preview:What you need to know for this weekend's games
The one that is left looks, on paper, like it might be the best game of state finals weekend. Second-ranked Gibson Southern clinched its first state finals appearance with a 44-7 win over Tri-West, playing in front of hundreds of Titans' fans who made the three-hour drive and will do so again on Friday. Third-ranked Brebeuf also had a long road trip, rallying for a 17-16 win at Mishawaka Marian.
Both teams are well-tested, led by seniors, and have unusual ties and commonalties. One is the connection between Allen and fellow Purdue recruit Joe Strickland, a standout Brebeuf senior defensive end who will be roommates with the Gibson Southern quarterback as early enrollees at Purdue in just a few weeks.
Not only is a state title on the line, but also future roommate bragging rights. But the similarities in Allen and Strickland as leaders extends beyond the numbers.
"Everyone can see how big and physical he is and the way he plays," Brebeuf Jesuit coach Matt Geske said. "But what Joe has meant most to us is the growth and leadership and the tone he's set for us. It wasn't just about being a talented guy, it was about the best player needs to be at the 6 a.m. workouts. We need to lift. We need to get bigger and stronger. I think that piece has really echoed throughout our leadership and been the most important part. More than just having talented players, you have talented players who are bought in."
Another similarity is that both coaches have family members also coaching in the state finals in other states. Nick's brother, Derek Hart, is coaching Kankakee in the Illinois Class 5A state title game on Saturday morning on the Northern Illinois campus. Geske's father, Frank, will be coaching Marist Catholic in the Class 4A state championship in Oregon on Saturday night.
But both coaches, while happy for their family members, have enough to worry about on their own for Friday's 3A showdown.
"No disrespect to any other quarterbacks that we played, but it's tough to mimic what (Allen) does," Geske said. "We've played quality opponents so we've seen that piece. But I don't know if we've seen a quarterback who can do what Brady Allen does."
Here is a closer look at the Class 3A title game:
Brebeuf Jesuit (12-2) vs. Gibson Southern (13-1)
Brebeuf Jesuit No. 3; Gibson Southern No. 2
3:30 p.m. Friday, at Lucas Oil Stadium
Bally Sports Indiana and IHSAAtv.org
Brebeuf is attempting to win its first state title in its third appearance. This would be the fourth state championship this fall for the school, which won state titles in boys soccer, boys cross-country and volleyball. Brebeuf could become the second school to win four state titles in the same season (Carmel won girls cross-country, girls soccer, boys tennis and football in the fall of 2011). Gibson Southern is looking for its first state title in football in its first state finals appearance.
Brebeuf coach Matt Geske, 45, is in his second season with a 17-7 record. Geske coached at Zionsville as an assistant for five years prior to coming to Brebeuf. He is a native of Portland, Ore., who played at Portland State. He coached at Lake Oswego (Ore.) prior to coming to Zionsville.
Gibson Southern coach Nick Hart, 36, is in his 10th season as coach with a record of 104-20. Hart, the athletic director at the school, is the son of Brownsburg coach John Hart. He was a three-sport athlete at Evansville Reitz and graduated from the University of Southern Indiana in 2012.
A closer look at Brebeuf Jesuit
Brebeuf lost only to 4A top-ranked Roncalli (28-10) and 5A No. 1 Cathedral (45-21) during the regular season. During a difficult tournament road, Brebeuf has knocked off three top-10 teams in No. 1 West Lafayette (24-14 in the sectional first round), No. 7 Norwell in the regional (38-9) and No. 9 Mishawaka Marian (17-16 in the semistate).
Brebeuf is led on offense by junior quarterback Nolan Buckman (2,993 passing yards, 37 TDs) and senior receivers Max Walters (84 catches, 1,298 yards, 16 TDs) and Anthony Annee (66 catches, 1,128 yards, 20 TDs). The defense is led by senior defensive end and Purdue recruit Joe Strickland (54 tackles, eight sacks), junior linebacker Cole Williams (92 tackles, seven tackles for loss), senior linebacker Jeremiah Jackson (81 tackles, 12 ½ tackles for loss) and senior lineman Jayden Childers (80 tackles, four forced fumbles).
A closer look at Gibson Southern
Gibson Southern's lone loss came in Week 4 — a 42-7 loss at South Warren (Ky.). The Titans, who are third in the state in scoring (46.7 points per game) and sixth in average margin per victory (32.2 points), have top-10 wins in the tournament over No. 10 Mt. Vernon (Posey) in the sectional first round (49-16), No. 5 Lawrenceburg in the regional (31-29) and No. 8 Tri-West in the semistate (44-7).
The high-octane Titans are led by Purdue recruit and senior quarterback Brady Allen, the state's all-time leader in passing touchdowns. Allen has thrown for 3,855 yards and 53 TDs with just three interceptions. His top targets are senior receiver Rory Heltsley (86 catches, 1,533 yards, 21 TDs) and senior receiver Brayden Malone (37 catches, 638 yards, nine TDs). Sean DeLong has 826 rushing yards and nine TDs and 30 catches for 406 yards and three TDs. Levi Dickerson, a senior linebacker, has a team-leading 149 tackles, including 21 for a loss.
How Brebeuf Jesuit can win
Put pressure on Allen. Tri-West hoped to do more of that in Saturday's semistate, but it did not happen often enough in Gibson Southern's runaway 44-7 win. Brebeuf only averages 116 rushing yards per game and probably won't have much success there against the Titans, who are allowing just 106 rushing yards per outing. Brebeuf will need Walters and Annee to make plays consistently in the pass game. It will probably take 30-something points to win.
How Gibson Southern can win
Same thing applies. The Titans have a physical defense and will look to pressure Buckman and force mistakes. It seems almost impossible to believe Allen will not have some success. He is difficult to pressure because he finds the quick reads and his receivers can run.
Brebeuf Jesuit 35, Gibson Southern 34
Call Star reporter Kyle Neddenriep at (317) 444-6649.


text:
"""
Puts a question/answer/comment onto the dullu queue for testing purposes.
"""

import json
import os
import linkbot
import pika
import os.path as osp
from dullu import Dullu

if __name__ == "__main__":
    with open(osp.join(osp.dirname(osp.abspath(__file__)), "..", "tests", "data", "test_question.json"), "r") as f:
        sample_question_json = json.load(f)

    """
    TODO - commit this current batch of work, then uncomment the following code. Going to implement some enums based on
    the stack exchange API
    """
    json_dict = {Dullu.JSON_KEY__ENTITYQ__ID: sample_question_json[Dullu.JSON_KEY__ENTITYQ__ID],
                 Dullu.JSON_KEY__ENTITYQ__TYPE: sample_question_json[Dullu.JSON_KEY__ENTITYQ__TYPE],
                 Dullu.JSON_KEY__ENTITYQ__BODY: sample_question_json[Dullu.JSON_KEY__ENTITYQ__BODY]}

    jdump = json.dumps(json_dict)

    connection = pika.BlockingConnection(pika.ConnectionParameters(host='localhost'))
    channel = connection.channel()
    channel.queue_declare(queue=Dullu.RABBITMQ_QUEUE_ENTITY_NAME, durable=True)
    channel.basic_publish(exchange='',
                          routing_key=Dullu.RABBITMQ_QUEUE_ENTITY_NAME,
                          body=jdump)
    print(" [x] Sent!")
    connection.close()


Our monthly hands-on lab series continues on Wednesday nights at Microsoft. The next session will be held at the Microsoft Mid-Atlantic Office in Malvern, PA on July 8 from 5:30-8:00. Refreshments are provided courtesy of TEKSystems.
Over eight months we will cover ASP.NET 5 and Azure Apps in depth. This is month four of the series. These Hands-On Labs will use a mix of popular tools and Visual Studio. Follow the instruction at https://github.com/aspnet/Home to add the new web framework to your laptop. This will not interfere with your existing .NET and Visual Studio work. This lab will require the new Visual Studio Code tool that was releases at the Build conference. Please install this before you arrive.
The lab details code editing of an existing MVC 6 site that was created in Visual Studio 2015. We cover basic code editing techniques, power keystrokes, window management, search, commands, file navigation, intellisense, snippets, version control, debugging, tasks, and language options.


While China's inroads into Africa and Latin America have been primarily about procuring natural resources, Chinese companies are now targeting large-scale infrastructure projects in Europe. In tandem with those big contracts, they are increasingly buying political clout in Europe and even contending with the International Monetary Fund (IMF), which as a rule ties aid loans to consolidation of the borrowers' national budgets.
Chinese state-owned corporations are assailing Europe from its outskirts: to wit, the government in Beijing has just clinched so-called strategic agreements with Moldova and Serbia. Under the arrangement with Moldova, China will grant the country a $1 billion soft loan in return for enormous building contracts to be awarded to the China Overseas Engineering Group (Covec), its largest construction group.
China has already shelled out billions in development aid to Africa, Asia and Latin America to buy access to natural resources and big infrastructure projects there. Development aid policymakers decry that strategy as "Chinese neo-colonialism", chiefly because, by lending money without any mandatory reform strings attached, Beijing is blatantly disregarding the urgent need for economic and social change in those countries, say many Western politicians. Instead of reforms, countries like Macedonia and Costa Rica have had to break off relations with Taiwan in exchange for Chinese aid.
Europe is the latest target of Chinese expansionism. Now that the Serb and Chinese presidents Boris Tadic and Hu Jintao have signed a strategic cooperation agreement, the two nations are negotiating a Chinese "soft loan" for the building of a bridge across the Danube and roads to the tune of €200 million, plus harbour extension projects, the development of special economic zones, and even defence projects. Under the terms of the cooperation deal, China must refrain from recognising the secessionist Serb province of Kosovo, and Serbia, for its part, from recognising Taiwan, which China still regards as part of its sovereign territory. "China is becoming the pre-eminent power in the world economy, so a strategic partnership with a country like that makes sense," reasons Tadic. Dušan Prorokovi of Serb ex-premier Vojislav Kostunica's opposition party, on the other hand, decries the colossal debt Serbia is incurring under the agreement.
Over in Poland, Covec is offering dumping rates in their bid to corner the new roadworks market, worth close to €40 billion in the runup to the 2012 EUFA European Football Championship there. Covec have just landed their very first two EU motorway construction contracts – viz. for parts of the new Warsaw-Lodz A2 expressway. The state-owned enterprise out-tendered big-name Western contenders by offering to build the roads at €6.3 to €6.5 million per kilometre, 60 per cent under the bid price and well below the quotes from Poland's PBG group and Ireland's SRB Civil Engineering. Since then, Warsaw has been wondering how Covec intend to maintain that price and whether cheap Chinese manpower will be flown in for the job – as for Chinese projects in Africa.
China is hammering out aid loans on easy terms in the neighbouring Ukraine as well, where prime minister Yulia Tymoshenko's "Fatherland" party is mulling cooperation with China's Communist party. What is particularly nettlesome about China's soft loans to Serbia and the Ukraine is that these very countries have signed on for billion-dollar IMF aid programmes, in exchange for which they are to submit sweeping austerity programmes in the national budget.


text:
Boundary changes would see Burnley and Nelson share an MP
Dominic Collis
Published: 16:01 Tuesday 28 February 2017
The next stage of a national review that could change the shape of Burnley's Parliamentary boundary has started.
Last year people were invited to take part in a public consultation as part of the national review of Parliamentary areas – those which an MP represents rather than borough or other boundaries.
There were almost 20,000 responses to that consultation across the country. The next stage of the 2018 Review is the publication of all those representations so that others may comment on them.
Interested parties have until Monday, March 27th to comment. All representations will be published on the Boundary Commission website www.bce2018.org.uk.
In Lancashire it is proposed to reduce the number of constituencies by two, to 14.
In the borough of Burnley the proposal is to move Gawthorpe, Hapton with Park (covering Padiham) and Coalclough with Deerplay wards into what is now Hyndburn, but change the constituency name to Accrington.
The Burnley constituency would expand northwards, taking in eight wards currently in Pendle and stretching up to, and including, Brierfield and Nelson.
The changes would not affect the borough boundaries or representation on Burnley Council.
The Boundary Commission for England is considering constituency areas across England as part of a periodic review of the boundaries of all Parliamentary seats. The overall aim is to have constituencies that have between 71,031 and 78,507 people eligible to vote living in them.
The Boundary Commission is due to make its final report in September 2018. Any changes would come into effect at the next scheduled general election in 2020.