Is it possible to run a sandwich ELISA using two sets of polyclonal antibodies for the same antigen? If I try and run ELISA using polyclonal antibodies, what are the major significant issues that will happen? How can I reduce background noise under such circumstances?
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AB Bayazid
Article A novel polyclonal antibody-based sandwich ELISA for detecti...
Will work well with polyclonals from 2 species.
You even can do it with one polyclonal, when you label it with e.g. biotin and use a streptavidin coated plate to immobilize the capture antibody. For the secondary, use a different label. Then, to introduce the reporter (e.g., peroxidase) use an antibody against the label.
Orthogonality is important to reduce background.
You can use the same polyclonal antibody for both capture and detection in sandwich ELISA. Coat the plates with the antibody directly (5ug/mL) then use the same antibody or the other polyclonal antibody labelled with biotin followed by HRP-streptavidin for detection. There are commercial kits available for biotin labelling of very small quantities of antibodies that take 30 min only from start to finish to carry out the labelling and have a conjugate ready to use in the ELISA. you can get them from the company ABCAM in Cambridge, UK.
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