1. Field of the Invention
The present invention relates generally to ultraviolet radiation devices. More particularly, the invention concerns an apparatus for use in genomic or proteomic research to visualize fluorescent labeled DNA, RNA or protein samples that have been separated for documentation and analysis.
2. Discussion of the Prior Art
By way of brief background, ultraviolet light (UV), which is electromagnetic radiation in the region of the spectrum located between X-rays and visible light, is typically divided into three principal ranges, namely long wave, mid-range, and short wave. For each of these UV ranges specific applications have been developed.
As a general rule, the desired ultraviolet wavelength is obtained from a fluorescent style tube that is an electric discharge device that uses a low-pressure mercury vapor arc to generate ultraviolet energy. The ultraviolet energy released in typical, commercially available fluorescent tubes is primarily at the wavelength of about 254 nanometers.
However, the fluorescent tubes can be modified to release other ultraviolet wavelengths by the use of phosphors, which have the ability to absorb the ultraviolet energy and re-radiate it in other wavelengths. For example, long wave ultraviolet of about 365 nanometers and mid-range ultraviolet of about 300 nanometers are created by coating the inside of the fluorescent tubes with the proper phosphors which convert the short wave ultraviolet.
In the past ultraviolet irradiation of selected articles has been accomplished using a single UV range fluorescent tube mounted within a suitable enclosure. In order to eliminate white light generated by the UV tube, some prior art devices make use of a UV transmitting ambient or visible light blocking filter that is typically mounted in front of the UV tube.
By way of example, U.S. Pat. No. 5,175,347 issued to the present inventor describes an apparatus for irradiating an object such as a specimen of material with ultraviolet radiation at a selected long, short or mid-wave length. Similarly, U.S. Pat. No. 3,936,186 issued to Boland et al. discloses an apparatus for exposing diazo printing plates and the like of the character that are used in the graphic arts field. In like manner, U.S. Pat. No. 5,288,647 issued Zimlich, Jr. et al. relates to a method by which polynucleotide specimens can be irradiated particularly for the purpose of fixing them to a substrate. Similarly, U.S. Pat. No. 5,736,744 issued to Johannsen et al. in which the present inventor is named as a coinventor, discloses a wave length shifting filter separate and apart from a transilluminator. The wavelength shifting filter uses phosphors in a flat array to provide a selection of visible wavelengths.
U.S. Pat. No. 5,951,838 issued to Heffelfinger et al., concerns a method and apparatus for achieving uniform illumination of an electrophoresis apparatus. In the Heffelfinger et al. method, uniform illumination is achieved by scanning the light source across the sample gel in a direction perpendicular to the axis of the source. The light source is comprised of one or more light bulbs placed in a light tray. Variations in light intensity near the source end portions is minimized using a variety of techniques including extended light bulbs, filters, reflectors, and diffusers, or supplemental sources.
The standard prior art method for separating, identifying and purifying biological samples is electrophoresis through a gel. The electrophoresis process is simple and well understood today. It is commonly used in one dimension separation where distinct bands of distinct biologicals are formed, or in two dimension separation where distinct spots or bands are formed.
Generally, following the process of electrophoretic separation, the separated biological samples are stained with a fluorescent dye, such as ethidium bromide. A set of multiple visible fluorescing dyes can be utilized that are capable of identifying specifically separated biological samples. These dyes have the ability to specifically attach (tag) themselves to specific biological samples and fluoresce in different visible wavelengths.
After the sample is dyed it is exposed to an ultraviolet radiation source, normally within the spectral bandwidth of mid-range ultraviolet (280 nm–320 nm). This range generally provides for the best and brightest wave shift conversion of the dye. During exposure, the dye labeled, separated biological sample is visible for viewing, documentation and further analysis. It is to be noted that other wavelengths of ultraviolet, such as short wave ultraviolet (generally considered as 254 nm), long wave ultraviolet (320–400 nm), broadband ultraviolet and a combination of short wave, mid-range and long wave can also be used to generate the fluorescent wave shift action of the dyes.
Although excitation of the fluorescent labeled biological sample is at times possible with visible wavelengths and light boxes that generate visible wavelengths, such as 420 nm or 480 nm, it is generally understood that UV excitation allows larger stoke shifts (that is the discrimination between excitation and emission wavelengths), enables higher signal to noise ratios and provides greater sensitivity.
A commonly used prior art tool for illuminating electrophoretically separated gels is the ultraviolet transilluminator (light box). These light boxes, generally comprise a single wavelength set of ultraviolet producing fluorescent lamps. These lamps are generally horizontally mounted within the light box behind a window upon which the dye labeled sample rests. The window typically comprises an ultraviolet transmitting, ambient (visible) light blocking filter material. Other ultraviolet light boxes are commercially available that provide dual UV wavelength combinations of 254 nm/365 nm, 254 nm/1302 nm and 365 nm/1302 nm. In this regard, commercially available mid-range ultraviolet light boxes interchangeably use the wavelength designations 300 nm, 302 nm, 310 nm or 312 nm, since the UV bandwidth output of these wavelength designations is substantially the same. Additionally, UV light boxes are commercially available that provide all three UV wavelengths of 254 nm, 302 nm and 365 nm. However, substantially all presently commercially available ultraviolet transilluminators (light boxes) use commercially available ultraviolet producing lamps that singly provide UV wavelengths in 365 (UV-A bandwidth), 302 nm (UV-B bandwidth) and 254 nm (UV-C bandwidth).
Another device used to capture fluorescent labeled biological samples is commercially available from Bio-Rad, Inc. of Hercules, Calif. under the name and style FLUOR S MULTIMAGER. This device uses a single broadband (290 nm–365 nm) ultraviolet fluorescent lamp. This ultraviolet fluorescent style tube lamp is horizontally mounted below the sample holding window and is typically scanned across the sample permitting the acquisition of the fluorescent signal via a charge coupled device (CCD) based camera system. This configuration limits the actual viewing of the fluorescent labeled sample by the researcher in real-time. The previously mentioned U.S. Pat. No. 5,951,838 issued to Heffelfinger, et al. and entitled “Method and Apparatus for Correcting Illumination Non-Uniformities” describes this method in greater detail.
As a general rule, all commercially available ultraviolet light boxes use 4, 5, or 6 fluorescent style UV generating lamps. These UV fluorescent lamps (254 nm, 302 nm, 365nm or broadband) are typically commercially available in 4 watt, 6 watt, 8 watt, 15 watt and 25 watt styles and in varying lengths. The lamps are normally configured in a horizontal pattern and are generally superimposed over a reflective aluminum reflector. Typically, a UV transmitting-ambient visible light blocking filter is positioned above the lamps.
It is well understood that conventional ultraviolet generating fluorescent style tube lamps of the type described in the preceding paragraph generate ultraviolet radiation in an arc formed between the electrodes in the lamp. However it is not well known that the intensity or output of this type of lamp diminishes from the center point of the arc towards the arc origination points. Accordingly, in virtually all wattages and configurations, the presently commercially available lamps provide a sample illumination surface that is substantially non-uniform. This problem of non-uniform UV illumination of fluorescent biologically labeled samples has been addressed in the past by the development of data manipulation and correction software that is specially designed to account for UV background on a fluorescent labeled sample. A description of such software and of its use is discussed in detail in U.S. Pat. Nos. 5,951,838 and 5,897,760 issued to Heffelfinger, et al.
Other prior art devices suggest the use of a cold cathode type serpentine grid to generate a more uniform visible light for use in LCD and photographic film viewing background lighting. A description of such prior art devices can be found in U.S. Pat. Nos. 5,731,658 and 6,069,441 issued to Lengyel et al.
Commercially available alternatives to the ultraviolet light box are available in devices that use lasers to illuminate the fluorescent labeled biological samples. Typically, these devices rely on laser light sources to illuminate the fluorescent “tagged” samples to excite the samples. In such devices, the laser source is scanned serially to excite each sample.
As will be better understood from the discussion that follows, the present invention overcomes many of the drawbacks of the prior art devices.