Secondary antibody and Strepavidin incubation times

From:Gayle Callis


30 minutes for secondary antibody (depends on concentration and antibody),
also temperature, agitation i.e. rotating platform.  Have used 60 minutes. 

20 minutes for Strepavidin enzyme conjugates, 30 minutes for SA-AP
(depends on source of this reagent as some seem better than other, in
particular SA-AP substrates), temperature, and agitation on a rotating

Some kits are more sensitive than others, the DAKO LSAB+ has less time and
found it I went over DAKO's recommendation - trouble! Following kit
instructions is a good idea since companies tend to optimize kits for you.  

DakoCytomation free handbook, Immunochemical Staining Methods 3rd Edition
has a good review of quick methods versus longer methods. 

Gayle Callis
Research Histopathology Supervisor
Veterinary Molecular Biology - Marsh Lab
Montana State University - Bozeman
S. 19th and Lincoln St
Bozeman MT 59717-3610

406 994-6367 (lab with voice mail)
406 994-4303 (FAX)


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