Gel documentation (gel doc) and analysis FAQs
Please click a question to view the relevant answer.
If you are unable to find the answer to your query here, please let us know. Our expert development and sales teams will be able to answer your questions directly, and we are more than happy to listen to your views and take on any suggestions concerning the information we present on our website.
Our Support Telephone Numbers are:
U.S. 800-686-4407 (Toll Free) or 301-662-2863
Europe +44 (0)1223 727123
Rest of World +44 (0)1223 727123
or contact us at support@syngene.com
FAQ 1:How do I know which excitation wavelength and which emission filter I should use?
Syngene provides full flexibility throughout the range of imaging sysems, with excitation energy sources available for all fluorescent, colorimetric and chemifluorescent applications.
Enter this web based facility to optimise your filter and lighting combination for an extensive range of dye applications.
FAQ 2:I can see UV tubes on my images. How can I avoid this?
Always check that the correct filter is selected for the chosen application. It is very easy to have the wrong one selected, especially when using a motorised wheel. For UV applications use the UV/IR filter. It is also possible to see tubes by using inferior light boxes where the illumination is uneven. Check the transilluminator (using a face shield) to see if you can see the tube pattern. With the Syngene GelVue transilluminator, you should be almost unable to detect the tube pattern by eye as illumination is so even. If you compare this to competitor light boxes you will see a remarkable difference. The tubes are often highly visible with dark banding patterns between them. This makes a huge difference to results obtained during quantification analysis. A band positioned directly over a tube may appear quite bright, whilst another positioned over a gap may appear fainter.
FAQ 3:How can I use the UV light box to illuminate protein gels?
By use of a white light / UV phosphor converter screen, you can convert UV to white light for the illumination of protein gels, autoradiographs, ELISA and colony plates. This method is more efficient than many normal light boxes, particularly combined UV and white light boxes. You will find the illumination is far more even when using the converter screen.
FAQ 4:When capturing images of protein gels and blots, what is the best way to illuminate them?
This depends on how the sample has been stored. The normal way would be to use the UV / white light converter screen and illuminate from below. This is fine for wet gels or those stored in bags and even dried onto film. However for gels dried on filter paper it is far more difficult. In this case you have to illuminate from above - if you illuminate from below you see the filter paper fibres as well as the gel. To avoid reflections from the surface you need to have even epi-illumination and avoid the use of saran wrap.
FAQ 5:When capturing images for chemiluminescence what difference does the signal to noise ratio make?
The longer the exposure time used to capture an image, the more background noise builds up in the system. This noise will often be visible on the captured image. Cameras used for chemiluminescence applications should be electronically or Peltier cooled. Cooling drastically reduces inherent background noise. The best results are therefore obtained by having the lowest signal to noise ratio possible.
FAQ 6:There are a number of cameras with different specialist applications. Which type should be used for a particular application?
When you are looking for an imaging system you should consider all the applications you are using. Protein gels, DNA & RNA gels, Zyma gels, colony counting, spot blots, ELISA plates and other images that require short exposure times, can all be done using a standard CCD camera system.
For chemiluminescence, spot blots or antibody assays which require much longer exposure times, a cooled CCD camera system should be selected.
Please bear in mind that at a later date your research interests may change, resulting in a requirement to upgrade your system. Always ensure that you can upgrade your system with minimal cost.
FAQ 7:My images are very dark. How can I improve them?
Firstly, check that the aperture on your lens is fully open. In addition, all SYNGENE CCD cameras have the ability to integrate (extend the exposure time), thus improving the brightness of the image. Using the integration control, increase the camera exposure - this will make the image brighter. This is especially relevant to faint bands on DNA gels. To get the correct aperture/integration simply and quickly whilst avoiding over exposure, use the Auto-Exposure function. The software then captures the correct integration time for the set aperture as well as assessing the saturation of the image during the process.
FAQ 8:What is on-chip integration and how does it affect my image?
On-chip integration is a method of adjusting the exposure time of the camera. This has the same effect as adjusting the shutter speed on any normal (35mm) camera. The higher the integration level you set, the brighter your image becomes.
FAQ 9:I am very interested in quantifying my images but am worried how accurate it will be. How do I avoid introducing inaccuracies into the analysis?
There are several points to be considered here. To obtain the most accurate analysis you must first have the best possible image This means the lighting must be as even as possible, preferably by using RTD (Reflective Tube Doubler) technology. The integration time should be set to a level that clearly shows all bands but does not saturate them. By using the Auto-Exposure function this can be done very simply. This will avoid most of the common inaccuracies that may be introduced into the image.
Neutral Field correction normalises the images by correcting for any non uniformities in either the lens optics or the lighting whilst maintaining GLP compliance.
The other area of concern is analysis of gels. It is often best to set the area of interest for a band to be the same for all bands being analysed. GeneTools allows you to set this automatically. You may also have smiling bands that are close together. These are very difficult to quantify, as many software packages do not allow for grimacing. Other packages find the centre of a band and draw horizontal lines to the edge of the track to define area. If smiling bands are close together the area defined will take in some of the bands above and below, often to the extent of ‘losing’ some of the bands of interest in the process. Any software at the very least should allow manual compensation for this. GeneTools will perform grimace correction automatically, thus eliminating any inaccuracy in quantification.
FAQ 10:What is saturation and how do I avoid it?
Saturation occurs when an image is over exposed. This means that the part or all of the image exceeds the dynamic range of the camera. Once parts of an image are saturated they are unsuitable for quantification as it is impossible to measure genuine differences in intensity.
Syngene systems using the GeneSnap interface have a live Saturation Detection function. When activated over-exposed white bands are viewed as “RED” areas. To avoid these, manually decrease the integration time until the red areas disappear. Another method is to use the Auto-Exposure function. The software increases the exposure time until saturation is reached on part of the image. It then takes off one frame (1/30 of a second) from this time and captures, thus giving the brightest image possible without saturation.
FAQ 11:I want to follow Good Laboratory Practice (GLP) in my work. How can I use my system to help me do this?
GLP is achieved by adhering to strict protocols or procedures. Using any Syngene system these procedures can be followed absolutely. The image captured can be saved in a private .SGD file format. This file cannot be changed or modified. The software records all system parameters from the moment of capture.
FAQ 12:I work with Green Fluorescent Protein. It is often difficult to visualise clearly, how do I get the best image/print for my labbook?
Illumination of the sample and filtering of the camera are the important issues here. Make sure that illumination is as even as possible, whether from above or below the sample. To increase contrast of the image, use the short band pass filter. This will give more definition between the protein and the background. You may find that transillumination by blue light improves your results, this option is available from Syngene
FAQ 13:Syngene systems have a motorised zoom lens. What is the significance of this?
Syngene selected the motorised zoom lens so that the user would not need to touch the camera lens assembly. Using a manual zoom lens means that the user can easily contaminate the camera/lens with some very nasty chemicals. Some chemicals crystallise upon drying and potentially prevent free movement of parts. Having a motorised lens prevents this from happening.
FAQ 14:In a lot of the brochures I read about “Dynamic Range” and “Dynamic Integration” What does this mean?
Dynamic Range and Dynamic Integration are two different parameters. Dynamic Integration is the term used for the integration of the camera i.e. its on-chip exposure time. The Dynamic range of the image is the number of grey levels to which the image is digitised in the camera. The Dynamic Range is directly related to the number of bits of the camera. The more bits the camera then the greater the number of grey levels or grey scale it has.
FAQ 15:I do chemiluminescence work. I cannot see my samples by eye so I need long exposures. How long an exposure can I set and how do I cope with the noise build-up on the image?
Most chemiluminescent exposure times will depend on the strength of the secondary antibody and the substrate used. Some chemiluminescent substrates work more effectively with CCD cameras than others, those that are poor may be improved by adjusting the secondary antibody. Exposure times vary from around 30 seconds to 8 -10 minutes. On average you should be looking at 1-2 minutes. The cameras we use in all our chemiluminescence systems are Peltier (electronically) cooled. This results in a reduction of noise to almost undetectable levels.
FAQ 16:What is the resolution of the camera in pixels?
Each camera is different - see specifications for each product.
FAQ 17:Can I buy a Syngene system locally and get local service?
Yes. SYNGENE has representation in over 65 countries worldwide. Please contact us to identify your local distributor.
FAQ 18:What does the Series Capture function do?
In this mode the software will capture a series of images which may or may not be additive. This function is especially useful when measuring chemiluminescence. The ideal exposure time is often unknown. By capturing a series of images you watch the image develop over time i.e.: you may capture 6 images with an exposure time of 10 seconds each; the third image will be equivalent to a 30 second single capture and the sixth will be equivalent to a one minute capture. The real beauty of series capture is that the user can select the best image in terms of appearance and saturation and save only that one. It is rather like developing a Western blot by using numerous pieces of x-ray film but without the disadvantages of time and money wasting.
FAQ 19:Can I use my images for publication?
Yes the images can be used for all the leading journals. Images in TIFF file format are usually used.
FAQ 20:Which types of printers can I use to print my images?
Any printer can be used so long as it has Windows drivers. The most common are thermal, dye sublimation, inkjet or laser printers.
FAQ 21:Can the software analyse colour images taken from scanners?
The software can analyse images up to 24 bit colour.
FAQ 22:Which operating systems can I use with my Syngene systems?
Syngene systems and their PC Windows operating system requirements:
|
System
|
98 SE
|
NT
|
2000
|
XP Pro/Home
|
ME
|
Vista
|
|
DigiGenius
|
|
|
|
|
|
|
|
InGenius L range
|
|
|
|
|
|
|
|
G:BOX and G:BOX Chemi
|
|
|
|
|
|
|
|
GeneGnome
*earlier versions of GeneGnome were supplied with Windows 2000 and should not be upgraded to Windows XP
|
|
|
|
|
|
|
|
GeneGnome HR
|
|
|
|
|
|
|
|
InGenius
|
|
|
|
|
|
ND
|
|
GeneGenius
|
|
|
|
|
|
ND
|
|
GeneGenius Super 12
|
|
|
|
|
|
ND
|
|
ChemiGenius 16
|
|
|
|
|
|
ND
|
|
ChemiGeniusQ
|
|
|
|
|
|
ND
|
|
ChemiGenius2 & XE
|
|
|
|
|
|
|
N.B.If you have an existing system please call us with the serial number for confirmation of compatibility.
FAQ 23: What are the minimum requirements for computers used with Syngene systems?
If you are buying a new system then the following PC specifications are suitable. If you are upgrading or replacing a PC on an existing system please contact Syngene.
Operating System: Windows XP Professional SP2
Processor: Intel Processor (Pentium 4, Pentium D, Celeron D, Pentium M, Core, Core 2 or Xeon) (we recommend 2GHz or higher Core 2 processor)
Memory: 256MB RAM (Standard G:Box or InGenius) 512MB RAM (G:BOX Chemi systems) (we recommended using 1GB or more if possible)
OpenGL compatible graphics card with a resolution of at least 1024 x 768 with true colour (1280 x 1024 resolution preferred)
Hi-Speed (480 Mbps) USB 2.0 port available* (note: must not be shared with a Lo-Speed device)
RS232 serial port available - if your PC does not have a serial port you might be able to use a USB to Serial RS232 conversion cable, but it must show in device manager as a standard COM port in the range COM1 to COM9
If you are using a laptop you may need a powered PCMCIA (Cardbus) Hi-Speed USB 2.0 hub for connection to the camera.
* You need to check that the system has at least one available high-speed USB 2.0 port
Note that there are three possible port speeds with USB 2.0: Lo-Speed (1.5Mbps) , Full-Speed (12Mbps) and Hi-Speed (up to 480Mbps).
Not all USB 2.0 compatible systems implement the Hi-Speed mode.
|
