Top 5 FAQs on Protein Analysis
Our support scientists answer the five most common questions they have about protein analysis. We explain how to study antibody-antigen interaction on the Biacore ™ system, measure fluorescence intensity on the Typhoon ™ imager, and more.
Question 1: I want to study an antibody-antigen interaction on my Biacore system. What sensor surface should I use?
If the antibody is used as a ligand, we recommend one of our ready-to-use detection chips:
- Sensor Chip Protein A: Primarily used for specific binding to orientation of human antibodies across the Fc region.
- Sensor Chip Protein G: Used for the oriented capture or binding of all human antibody subclasses and a wide variety of mammalian antibody subclasses.
- L protein: Used for directed antibody uptake. This sensor chip is ideal for studying a wide range of antibody fragments, including Fabs and domain antibodies.
To choose an appropriate sensor chip, consider your species and type of antibody. We have three different capture kits for you to choose from:
- Human Fab Capture Kit: This kit includes reagents for the capture of human antibody Fab fragments in biomolecular interaction assays on a Biacore SPR system.
- Mouse Antibody Capture Kit: Choose this kit if you need reagents to capture mouse antibodies as ligands in biomolecular interaction assays.
- Human Antibody Capture Kit: Use this kit to capture human or humanized IgG antibodies as ligands in biomolecular interaction assays.
You can also use a method such as amine coupling to attach the antibody to the CM5 sensory chip – but unlike the ready-made chips and kits we mentioned above, you will need to establish regeneration conditions. .
Question 2: I have a target protein and would like to study its interaction with other molecules. What sensor surface should I use?
Choosing a sensor surface depends a lot on the protein and other molecules you want to study. Here are some general tips:
If your target protein is tagged, we have capture kits and sensor surfaces for capture via His, GST, and biotin tags. For covalent binding, we recommend using our Biotin CAPture kit and biotinylating your protein with a friendly and protein friendly protocol. The Biotin CAPture Kit allows for the reversible uptake of biotinylated ligands and allows you to easily see multiple molecules bind to your target protein on a new unused surface each time. As with all of our kits, you do not need to establish regeneration conditions as we provide a regeneration protocol and solution in the kit. Our second recommendation is to use a covalent amine coupling with our Sensor Chip CM5. There is a downside: if you have stable interactions, you will need to establish regeneration conditions in order to be able to reuse the surface.
Question 3: I am using the Typhoon FLA 9500 imager to visualize the fluorescence of 6-FAM labeled DNA on agarose gel. I also want to visualize and measure the fluorescence of my liquid samples in a 96 well plate. I know the instrument can read and image the titration plate – can it also measure the fluorescence intensity in each well so that I can quantify my samples?
You can perform this type of application using the multistage and title plate frame. You should use titer plates designed for fluorescence applications (that is, low fluorescence titer plates). Then you can analyze your data using ImageQuant ™ TL matrix analysis, which was also created for this type of application. Please note that you cannot extract relative intensities if you perform a calibration curve on the side (i.e. steps of known concentration of your probe).
Question 4: I want to transfer my machine and my program to a newer computer (Windows ™ 10 Pro). Is it possible? Do we need to install anything on the new computer?
We have software compatible with Windows 10. Please note that you will need to transfer the calibration from the flat frame, or you can request a visit from a Cytiva support engineer who can do this for you. Please contact us with the serial number of your instrument and we will help you transfer your machine.
Question 5: Where can I find the software installation file for ImageQuant TL?
Type IQTL in the search bar of our website and click on the page of the product you are looking for. Click READ MORE in the product description and click the software package to download the installation file. Note: You can only download the latest version of IQTL from our website. To download an older version, please contact us.
To learn more about our Biacore SPR systems, please visit: www.cytiva.com/en/au/Biacore-SPR
For more information on Western blotting and imaging, please visit: www.cytiva.com/en/au/WB