Radioimmunoassay (RIA) is a very sensitive immunoassay used to measure antigen concentrations. RIA has many uses, including drug testing, virus screening, disease diagnosis, etc. Creative Biolabs has extensive experience in RIA. We describe specific protocols for RIA implementation, and our scientists are happy to discuss the details of your project and develop a tailored experimental strategy.
You can see below for general RIA protocols and troubleshooting tips for common problems, please contact us for details.
|Stages||Solutions and Reagents|
|Preparation||Standards, RIA buffer|
|Immunoassay||Antibodies, radioisotope ligands, dilution buffer, wash buffer|
RIA uses antibodies to detect and quantify antigens in samples. As unlabeled antigen from the sample and quantitative radiolabeled antigen reacts with quantitative antibodies, as the amount of unlabeled antigen increases, less and less of the labeled antigen binds to the antibody. Thus, the concentration of free labeled antigen is proportional to the bound unlabeled antigen.
First extract and dilute your sample to be tested. Prepare the sample concentration within the measurement range. Prepare antigen standards using dilution buffer and perform serial dilutions as directed.
Make a known amount of antigen radioactive by labeling it with radioisotopes. Then fix a known concentration and a quantitative amount of the specific antibody in a micro-titration well. Next, add a known amount of labeled antigen to the wells and incubate. Wash carefully after incubation to remove any unbound labeled antigen. At this point, the wells will be most radioactive.
Set up wells for control, sample to be tested, and standard samples. Then add unlabeled antigen to the wells for incubation. The unlabeled antigen will compete with the labeled antigen for binding to the antibody, so there will be free labeled antigen in the wells. Wash carefully again to remove the free labeled antigen.
Measure well radioactivity by gamma counter. As the concentration of unlabeled antigen increases, the radioactivity decreases. A standard curve is obtained by plotting radioactivity versus unlabeled antigen concentration. The sample to be measured is run according to the procedure. The measured radioactivity is calibrated with the standard curve to determine the concentration of the antigen.
Abnormal standard curve
These are informational resources related to RIA experiments. We hope you will browse and get support on our webpage. Or if you have questions about it, you can contact us to discuss troubleshooting methods.
For research use only. Not intended for any clinical use.