Re: [Histonet] negative immunohistochemistry control
Peptide inhibibtion is a good control to ensure
your antibody can be adsorbed by your antigen but
it does not tell you whether or not your antibody
will non-specifically bind to your tissue. It is
a good but not a sufficient control.
The best control is to stain tissue which DOES
NOT express your antigen, it should be negative.
>I'm not in a clinical setting - but I use peptide inhibition as a negative
>control for each antibody.
>Sent by: email@example.com
>03/08/2006 08:21 AM
> To: "HISTONET" , "Rene J Buesa"
> cc: (bcc: Jackie M O'Connor/LAKE/GPRD/ABBOTT)
> Subject: Re: [Histonet] negative immunohistochemistry control
>I don't understand the idea of only one negative control per case, as
>opposed to each antibody. How does one evaluate non-specific staining of
>antibodies that do not have a negative control? What if non-specific
>staining is inherent to a certain antibody? For example, the new CAP
>question regarding staining of endogenous biotin, particularly in kidney
>liver - if you do not run a negative control for that specific antibody,
>do you know if there is non-specific staining?
>----- Original Message -----
>From: "Rene J Buesa"
>To: "cynthia haynes" ;
>Sent: Wednesday, March 08, 2006 7:51 AM
>Subject: Re: [Histonet] negative immunohistochemistry control
>> The idea of the negative control is to try to determine if any
>detected in the case is due to specific or unspecific binding.
>> You don't need to run a negative slide for each antibody you are going
>to test on the case, you only need a negative slide per case, no matter
>many antibodies you run. The only requirement is that the negative slide
>to come from the same tissue tested.
>> What I used to do was to add buffer instead of the primary Ab and all
>the other reagents the same as in the slides run for specific Abs. You
>also add non-immune globuline instead of the antibody but in this case you
>would need 1 negative control for each type Ab (1 negative for monoclonal
>Abs and 1 for polyclonal Abs).
>> I hope this will help you.
>> René J.
>> cynthia haynes wrote:
>> Good Morning everyone, I have a question about immuno
>> staining. I've been away from this type of staining
>> for a while and now I am doing them again on a regular
>> basis. Why do you run a negative control with each
>> run? Are you only suppose to put the normal serum on
>> negative control only? I've forgotten; would someone
>> please answer these questions for me. Thanks in
> > Cynthia Haynes H.T.
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