[Histonet] negative immunohistochemistry control

Andrea T. Hooper anh2006 <@t> med.cornell.edu
Wed Mar 8 12:12:06 CST 2006

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.
>"Chris Pomajzl" <cpomajzl <@t> cpllabs.com>
>Sent by: histonet-bounces <@t> lists.utsouthwestern.edu
>03/08/2006 08:21 AM
>         To:     "HISTONET" <histonet <@t> lists.utsouthwestern.edu>, "Rene J Buesa"
><rjbuesa <@t> yahoo.com>
>         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" <rjbuesa <@t> yahoo.com>
>To: "cynthia haynes" <naje1972 <@t> yahoo.com>;
><Histonet <@t> lists.utsouthwestern.edu>
>Sent: Wednesday, March 08, 2006 7:51 AM
>Subject: Re: [Histonet] negative immunohistochemistry control
>>  Cynthia:
>>    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 <naje1972 <@t> yahoo.com> 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
>>  advance.
>  > Cynthia Haynes H.T.
>  >


More information about the Histonet mailing list