Hi Peter,
I have seen that phenomenon quite often during testing different batches (one
batch originate from one mouse) of ascites from the same hybridoma cells. The
background fluorescence was sometimes one decade higher (max.). I also realized
it has nothing to do with Fc receptors.
There are two additional questions:
1/ Does your mAb concentration match your isotypic control conc.? > If yes,
than we can't speak about "non-specific" binding but this is rather specific.
2/ Is your particular mAb an ascites fluid or a culture supernatant? > If it is
an ascites fluid, my interpretation would be that the higher background comes
from natural antibodies (usually low affinity Ab against carbohydrates,
cross-reactive) and their levels could be different in a particular mouse,
therefore different batches of mAb could give you various background staining.
I am not sure whether this applies also to culture supernatants (I have not
tested large number of supernatants) but I have not seen such a variability in
their background staining.
Hope this helps.
-- _______________________________________________________________ Karel Drbal Institute of Molecular Genetics Academy of Sciences of the Czech Republic Videnska 1083 142 20 PRAHA 4 Czech Republic, Europe voice: +42-2-4752589 fax: +42-2-4727979 e-mail: drbal@biomed.cas.cz WWW: http://leuko.biomed.cas.cz
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