I am planning to order an antibody, but for the commercially available antibody, it is written that the application is ELISA. So, if I want to order the same antibody can we use that?
If the antibody comes with the ELISA kit, better to avoid using it. Since it has probably been diluted to cater for ELISA analysis. Also, usually Western blot need a higher concentration of antibodies. But it’s possible if it’s the other way around.
Hello Tiyas Sengupta,
For an antibody to be used in experiment, it must bind as specifically as possible, meaning it must bind the correct epitope and should be clearly visible above any background. Non-specific binding can lead to false positive results or create sufficient background noise as to mask the protein of interest.
When you purchase an antibody, the specification sheet mentions the application for which it can be used. This means that the manufacturer has validated the antibody for that particular application, and it cannot be used for any other applications unless stated.
Please note that the conditions of each assay are different. For instance, a western blot depends on the denaturing of proteins. So, a western blot-validated antibody may work fine in denaturing conditions but may fail to recognize antigens in their native conformation (such as in ELISA). Likewise, an antibody validated for native protein affinity could fail to bind the same antigen following denaturation or fixation.
So, I would not recommend the use of antibody which is meant for ELISA, to be used for western blot.
Best.
Antibodies for Western blots bind to denatured proteins, which is not the case for ELISA. It does not mean that it won't work, you have to try it. I got some positive results in the past.
Hello!
You will try. We had a positive answer for detection of PVM and PVY coat proteins with antibodies from Bioreba
Tiyas Sengupta
Yes, you can often use an antibody meant for ELISA in Western blot, but it's not guaranteed to work as well. To make sure, you can test the antibody using a dot blot first. In a dot blot, you spot small amounts of your protein on a membrane and apply the antibody to see if it binds well. If it shows good binding, it’s more likely to work in a Western blot. This quick check can help you decide whether to proceed with the full experiment.
Hello! You could try. I am also successfully using the Western blot antibodies for IHC. Have a wonderful day!
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