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Sandwich optimization and recombinant purity

Sandwich optimization and recombinant purity - ELISA Assay Forum

Sandwich optimization and recombinant purity - ELISA Forum. Discuss and post questions including optimization, troubleshooting, protocols and theory of ELISA assays using monoclonal, and polyclonal antibodies or peptides.


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Old 05-18-2010, 03:31 PM
Pipette Filler
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Default Sandwich optimization and recombinant purity



Hi all,

Can anyone help me about how to improve the sensitivity of the Elisa assay I'm trying to optimize?
I'm coating directly my mAb, saturate, then add samples or standard, after a polyclonal antibody and at last antibody labelled with HRP.
(with washing at every steps)

Do you think I need to increase the incubation time of one of the reagents? or increase the quantity of reagents?

I don't really know what to change to have a better sensitivity.

And another question is about the purity of the protein used as the standard in my test. Do you think a raw recombinant protein is enough or it is better to have a purified protein?
Thanks for your help
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Old 01-08-2011, 08:16 PM
Pipette Filler
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Default Re: Sandwich optimization and recombinant purity

10-20% below the saturation point for the capture antibody might help to decrease entrapment of detection or 2 antibody (HRP linked).
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optimization , purity , recombinant , sandwich


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