Re: more details on "Oops wrong 2ndary"

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From:Katri Tuomala <>
To:Bret Morrow <>
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Hi Bret,
If I understand correctly, your primary antibody was raised in rabbit and
your secondary was supposed to be goat-anti-rabbit. Before your primary
antibody,  did you block with normal goat serum? If you did, you are out of
luck unless somebody knows how to detach that goat serum off your sections,
since the "wrong"secondary is tightly bound to that blocking serum and your
whole section would be staining, if you were to complete the method. Any
thoughts anyone?


Katri Tuomala
Anatomic Pathology
St.Joseph's Hospital
Hamilton, Ontario, Canada

Bret Morrow wrote:

> Hi,
> I was asked for more details about a SNAFU using the wrong 2ndary
> antibody.  The 2ndary was a complete miss; rabbit anti-goat, instead of
> goat anti-rabbit.  So the primary should be left untouched.  My main
> concern is that the tissue was stored in cyrosolution while we tried to
> figure out what had gone wrong.  The cryosolution kept it at -20 oC but
> it was not really frozen (no ice crystals).  But I have some concern
> about the
> antigen-antibody interaction during the cryopreservation.  If it
> separates, thats OK.  If it remains intact, thats OK, too.  But if the
> antigen-recognizing portion of the primary anibody remains intact and
> the rest is destroyed, we're out of luck,  I don't imagine there is much
> information about this situation out there, but I think (feel or hope,
> may be a better choice of words) that the 3rd option is unlikely.
> Any other thoughts on this???--I am planning on running the tissue
> through the primary and then the right secondary next week some time.
> Thanks for the input,
> Bret
> ____________________________________________
> Bret A. Morrow, Ph.D.
> Associate research scientist
> Associate clinical professor
> Yale University School of Medicine
> New Haven, CT 06520-8066
> voice 203.785.4515
> fax  203.785.7670
> email

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