My lateral flow assay uses a gold particle linked capture and secondary surface bound antibody; both mouse Mabs. The sample is human blood and I'm looking for a disease protein marker. How can I best reduce, or better still, remove non specific binding that can give false positives?
I know this is a Broad Question to ask but I'd really appreciate your help: Thanks.
John Clarkson
Dear John, False positives are primarily due to nonspecific binding of the conjugate particles. You can overcome this problem using bigger pore size membrane, usually at the cost of sensitivity. Addition of detergents in the sample pad can also help minimize it. Furthermore, as you are using human blood samples, look for better and efficient separator presence of RBCs, even in traces, results in unfavorable result.
I am sending a link that may help somehow you.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4263565/
Dear John, False positives are primarily due to nonspecific binding of the conjugate particles. You can overcome this problem using bigger pore size membrane, usually at the cost of sensitivity. Addition of detergents in the sample pad can also help minimize it. Furthermore, as you are using human blood samples, look for better and efficient separator presence of RBCs, even in traces, results in unfavorable result.
I am sending a link that may help somehow you.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4263565/
hi
Your question is very general.
1- if the size of gold nano particle about 20 nm or more is ok.
2- If desired protein on the surface of white blood cells you can reduce the background used by RBC lysis buffer.
3- The free or unconjugated nanoparticles can be effective on testing background.
good luck
Thanks for all your answers.
The protein marker is in the blood serum
To Atul Jha,
Your answer is familiar from other sources.
I guess I could screen for types of Human anti-Animal Antibodies by, say adding animal serum to the chase buffer? If the NSB is reduced by the addition of mouse serum in the chase buffer then the NSB was caused by Human anti-Mouse Antibodies. Repeat for other animal serum in chase buffer to screen for other Human anti-Animal Antibodies.
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