Dear Ronnie,
I would suggest to cut off the standard lane!! and process the standard and
the test sample separately in the appropriate way. You can still compare the
two, no problem. 
Or - change from the biotinylated standard to a prestained standard (which
is sold by lots of companies)and does not need any further processing, less
work.
regards,
Angelika

-----Ursprüngliche Nachricht-----
Von: Ronnie Houston [mailto:wee_rory@hotmail.com]
Gesendet: Mittwoch, 28. Juni 2000 21:08
An: histonet@pathology.swmed.edu
Betreff: Western Blots


Not strictly a Histo question, but one of our researchers is doing some 
Western Blots for us on skeletal muscle samples from suspected Beckers 
Muscular Dystrophy.
She is using biotinylated standards. However, the primary dystrophin 
antibody is not biotin-conjugated, so she is applying a biotinylated 
secondary anti-mouse Ig to both test samples and the standard. This is 
followed by an avidin-conjugated alkaline phosphatase and visualization with

BCIP/NBT.
My question is, can you compare the results of the standard to the test when

the biotinylated standard has been subjected with IgG-biotin? Does this 
extra amount of biotin affect the result?
My initial thoughts are that one cannot directly compare results because 
differently prepared samples have been run. Also there has been no blocking 
for endogenous biotin.
Thanks for any help you can offer.

Ronnie Houston
Cytochemistry & Molecular Pathology
Texas Scottish Rite Hospital for Children
2222 Welborn Street
Dallas, TX 75219
(214) 559 7744
(214) 559 7768 - fax
rhouston@tsrh.org

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