colony screening with Amersham ECL (RPN 3000 and 3004)

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Sun Dec 20 21:52:01 EST 1992

In article <1992Dec16.151837 at> Christiane Bouchier,
bouchier at writes:
>A week ago I asked whether anyone had ever carried out tests for
>colonies  with Amersham ECL direct system. So far I received no
>that could help me overcome the problem we have of detecting peroxydase 
>specific hybridization because of a high background.
>Once again thanks for your help !

 We have had similar problems of background with the ECL system.  We
overcame the problem by including the milk powder (2%) blocking agent in
every step i.e pre-hyb, primary antibody, secondary antibody, and  during
washing of filters. The background was considerably reduced.  Another
trick we found to increase sensitivity was to use filters from other
companies.  Even though they were all marketed as 0.45um nitrocellulose,
we found different (and reproducible) levels of senistivities.
I hope this is of some help.

 *Eamonn Gormley,  E-mail, E.P.Gormley at *

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