IgM competitive capture ELISA

littgj at esvx23.es.dupont.com littgj at esvx23.es.dupont.com
Wed Oct 25 06:15:43 EST 1995


In article <46heuk$itd at stork.qut.edu.au>, d.beasley at qut.edu.au (David Beasley) writes:
>
>I posted an article here a while ago asking for some help with regard to
>some IgM capture ELISAs I'm doing.  
>
>I have been trying capture ELISAs in 96-well plates from several 
>manufacturers, and it seems that the majority of the antibodies are 
>coating ok but cannot bind the antigen when it is added to the plate (maybe
>because the antibodies can't alter their conformation very well?).  So
>now I am looking for alternatives.
>
>At the moment I am setting up to try blotting the antibodies onto nitrocellulose
>and then do a capture ELISA on the membrane, but I was wondering whether
>anyone has any other suggestions as to how I could immobilise these antibodies.
>
>Thanks,
>
>David B.
There are other ways but you might try coating the wells with
a secondary Ab to the primary (e.g., goat antimouse or whatever)
at a fairly high concentration, washing and then coating with
your primary
The primary will now be immobilized through the secondary and
might have better binding characteristics. You may also be
surprised that the amount of primary needed may be a lot 
less (if you ar e luckey!



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