To keep track of information generated in Western blots, most scientists use a documentation system. These tools turn images of protein analysis—the data captured from chemiluminescent, colorimetic, or fluorescent readers—into digital files. “Western blotting documentation has improved dramatically in the last several years,” says Raymond Miller, product manager for protein quantitation at Bio-Rad Laboratories. With a documentation tool, the data can be analyzed more quantitatively. Also, the saved files can be searched.
Still, the way that scientists use Western blots continues to evolve. “Researchers running Western blots have been slowly moving away from the film and dark room paradigm with many labs adopting increasingly popular fluorescent Western blotting techniques,” explains Austin Hallgren, technical product specialist at Azure Biosystems. “There are two main advantages of fluorescent detection: quantitative data and the ability to multiplex.” Nonetheless, Hallgren notes that chemiluminescence is more sensitive, and some researchers revert to that approach where the highest sensitivity is needed.
In addition to various techniques for collecting data from Western blots, most scientists also require a system that can handle multiple targets. “Most researchers will start a project with one fluorophore or dye but during the project will likely have the need to expand beyond a single or a couple of fluorophores,” according to Miller.
The available features—various imaging and target-labeling options—already indicate that a Western blot documentation system does not stand alone.
Make it multifunctional
Various modern imaging systems provide a complete solution for Western blotting documentation. That’s what Bio-Rad had in mind with its ChemiDoc MP Imaging System. “Traditionally, researchers needed multiple imaging systems—for example, x-ray film developer and laser scanner—for different applications and even for different sensitivities,” Miller says. “The ChemiDoc MP matches and exceeds these traditional solutions with the convenience of having one system for everything.”
Various modern imaging systems provide a complete solution for Western blotting documentation.
A variety of vendors aim at easy multifunctional use. “We continue on the path of developing innovative products that streamline and modernize Western blotting,” says Paul Haney, senior product manager at Thermo Fisher Scientific. “With the iBright Imaging Systems, we aim to improve the convenience, speed, and quality of Western blot experimentation and circumvent the limitations associated with the capture of Western blot images using x-ray film.” He adds, “Our iBright FL1000 model is designed to work seamlessly with our SuperSignal chemiluminescent substrates or our Alexa Fluor Plus reagents for maximum multiplex abilities with up to four proteins detected on a single blot.”
Depending on a lab’s need, some kinds of imaging matter more than others. To accommodate that, LI-COR Biosciences offers three products for Western blot imaging: the C-DiGit for chemiluminescence, the CLx for infrared, and the Fc for chemiluminescence and infrared. In describing these platforms, Jeffrey Harford, the company’s senior product marketing manager, says, “Our primary goal is to provide our customers with not just a product, but a complete solution to get publishable Western blotting data.” He adds, “This is a huge area right now, as publishers are beginning to tighten the guidelines on Western blot submissions due to reproducibility issues.”
Other vendors also make multifunctional systems. For example, platforms from Azure Biosystems combine laser-based fluorescence Western blot detection with chemiluminescence detection. “Instead of being relegated to the detection chemistry dictated by capabilities of their imager, Azure imagers enable researchers the power to choose their detection chemistry based on the specific needs of their experiment,” explains Hallgren.
The Syngene G:BOX mini Gel and Blot Imaging System from Synoptics also supports fluorescent and chemiluminescent detection. They have optimized high resolution and sensitivity by incorporating a moving platform instead of digital zooming. “Owning the camera company allows us to also optimize a CCD camera for this purpose,” says Simon Leath, vice president of sales and marketing at Synoptics. “Plus, the software for these systems includes an application that allows stain-free gel capture and analysis.”
Depending on the research, some scientists want more than just the LED-CCD based camera systems for quantitative fluorescence imaging. Joe Hirano, product manager at GE Healthcare Life Sciences, explains that “GE, with its Amersham brand of imaging systems, provides two different platforms.” For sensitive chemiluminescence detection and for basic fluorescence work, researchers can use the AI600 range of LED-CCD based cameras. “The very popular Typhoon laser scanners are used for advanced quantitative fluorescent work,” Hirano says. “The Typhoon Laser Scanner provides researchers with a great combination of high sensitivity and a very high dynamic range with more than five orders of magnitude.”
Hirano also points out that GE recently added the Typhoon NIR Plus Scanner to the Typhoon platform. The Typhoon NIR Plus is a near infrared scanner that has a three-laser configuration for tri-plex Western blotting that can, for example, detect two target proteins and one housekeeping gene for normalization on the same membrane. “A platform like that along with products working together with it, expand a researcher’s options,” Hirano explains. “As an example, the Amersham Quickstain and Typhoon NIR Plus combination enables researchers a more accurate and reproducible Western blot normalization by supporting total lane normalization.”
As you think about a documentation system, remember that it is a system. Consequently, Harford points out: “Our solution includes not just instruments, but also critical tools, including handbooks, protocols, and reagents, especially ones geared toward publication requirements; and finally training in order to meet new Western blot guidelines.”
Imaging in an instant
These systems often need to handle high workflows. “We know that many researchers want to take the best images quickly and with a minimal amount of training so they can spend more time analyzing their data,” Miller points out. So, Bio-Rad developed its Image Lab Touch software, which acquires images on the ChemiDoc MP. He says this software “was designed and tested to ensure users can learn in minutes.” It also includes help windows to easily explain options.
Western-blot platforms must be easy to use, and vendors make that a priority. In describing the Invitrogen iBright FL1000 and CL1000 Imaging Systems, Jessica Slack, associate product manager at Thermo Fisher Scientific, says that they “deliver high resolution and sensitivity, combined with advanced automated features—simply unbox, setup, and capture stunning images within minutes.” These instruments also help scientists capture and catalog information, because they can “rapidly acquire and automatically analyze data from up to four mini blots or gels at a time,” Slack says.
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No doubt, scientists look for speed from these tools. At the Protein Facility of the Iowa State University Office of Biotechnology, manager Joel Nott says that their documentation systems “work sufficiently for what they are used for.” When asked what advances that he’d like to see, he says, “Faster scan times.”
In brief, tomorrow’s Western blot documentation systems need to be faster and smarter. As Harford concludes, “The bottom line is: There are lots of Western blot systems out there, but if they are not used in a fashion to meet new publication guidelines, it is going to lead to a lot of frustration for researchers.” Frankly, that’s exactly the sort of thing that a Western blot documentation system should prevent. Scientists use these systems to simplify processes, analyze data more accurately and to publish the most objective and complete studies.
The iBright FL1000 detects up to 4 proteins on a single blot. (Image courtesy of Thermo Fisher Scientific.)