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Non-specific binding in Lateral flow assay - (Jul/13/2010 )

HI all,
I am new to this forum and am not quite certain where I should post this, but I think this is a good starting point. We are developing a lateral flow device and I am having a lot of problems with non-specific antibody-antibody interactions. The lateral flow test is of a sandwich style (very similar to a sandwich Elisa). I have a rabbit polyclonal at the test line and the same polyclonal on the detector particle. The antigen is a protein displaying many epitopes which is why I am able to use the same antibody. I do get a relatively decent dose response, but i get very high background in the negative control (antigen absent). I have tried varying the concentrations of tween 20, BSA, and polymer.

Any suggestions other than changing around the polyclonals? Is this just the nature of some polyclonals?



Tween-20 or Triton X can be dried on/in the paper/lateral flow support in conjunction with the blocking agent (BSA etc). The sample pad should contain the dried buffer and surfactant as well. If your sample has heterophile abs you will need to block them with non-specific rabbit IgG (which you can test by pre-incubation before running the test.)

If you are using Au particles or colored latex (I suppose that is dried...) a surfactant should be in that matrix as well.

Make sure your polymer is titered to the right concentration in that it increases ab-ag interaction (and also increases background).