[Histonet] negative IHC controls

[Hadi Yaziji]

Theoretically, I agree with you that (a) running multiple negative 
control sections to cover the different antibodies/pretreatments is 
ideal and (b) money should be well spent on QC (I can't emphasize that 
enough). However, I honestly don't remember when was the last time when 
I had a false positive staining on a patient's slide that the negative 
control was the only way that allowed me to recognize the false 
positive signal on the real slide. And we look at 100s of IHC stains 
every day.

Experienced pathologists and technologists can still recognize in > 99% 
of the times a false positive signal from a real signal on the tissue 
section of a given antibody without even having to look at negative 
controls. I don't think it should be a CAP requirement at all. In fact, 
I'd say the same thing on positive controls. All you need is one 
positive control per antibody, but not one control/per antibody/per 
patient. In many cases positive internal controls are present on the 
same slide, so you can tell whether your antibody worked simply by 
evaluating the expected positive internal controls. In fact, if your 
positive 'external' control worked and internal controls didn't, then 
you need to repeat your antibody test regardless of the positive 
external control. External controls are different tissues, fixed 
differently, processed differently and the tissue ages differently (in 
terms of its antigenicity).

Individuals who perform and interpret IHC studies must have the 
knowledge to recognize the expected sub-cellular localization of every 
antibody on every type of tissue, including aberrant signals. For 
instance, you can easily dismiss a granular cytoplasmic TTF-1 signal as 
not positive, because TTF-1 is a nuclear transcription factor and we 
know it's expressed in the nuclei. However,  hepatocellular carcinomas 
can give you a consistent and reproducible granular and cytoplasmic 
TTF-1 signal, and if you subsequently run confirmatory markers 
(HepPar1, polyclonal CEA, CD10, etc.) they will be positive. Such 
(granular cytoplasmic) signal should be recognized by the interpreter 
(tech or pathologist) as a specific signal for  tumors with hepatoid 
phenotypes. This is just one of many examples..

The bottom line: in real life, one negative control per case is more 
than sufficient. And to say the least, it's inaccurate to state this is 
poor patient care and lousy quality control.

I look forward to any constructive criticism.

Hadi Yaziji, M.D.
PhenoPath Laboratories

On Thursday, October 9, 2003, at 07:03  AM, Horn, Hazel V wrote:

> Not only should you be running a negative control for each patient 
> slide.   That negative control should be treated just as your antibody 
> is.   If the antibody is rabbit and antigen retrieved, so should your 
> control.   If another antibody on the same patient is mouse and not 
> retrieved another negative control should be run with this same 
> protocol.   In the United States, labs that are inspected by the CAP, 
> are required to run these controls.    MONEY should never be 
> considered as a reason to stop doing a part of a procedure.   It's 
> poor patient care and lousy quality control.   IMHO.
>  
> Hazel Horn, HT/HTL (ASCP)
> Histology Supervisor
> Arkansas Children's Hospital
>
> Phone - 501.364.4240
> Fax - 501.364.3912
>
>
> -----Original Message-----
> From: vermast [mailto:vermast <@t> rogers.com]
> Sent: Wednesday, October 08, 2003 3:57 PM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] negative IHC controls
>
>  
> I would like to get a feel for how many out there are running negative 
> control slides for IHC. 
>  
> In our lab we do just a handful of antibodies and initially I had been 
> running a negative control slide with each patient slide.   After much 
> discussion with our pathologists, we decided to omit these negatives 
> (which were conistently negative) and continue to just run a positive 
> control with each primary antibody for the run.  We use the Dako 
> autostainer and prediluted primaries.  The decision to stop running 
> negatives also coincided with Dako's decision to sell the negative 
> control sera separately from the primaries (they used to come packaged 
> together).  Perhaps I assumed that discontinuing to pair these 
> reagents together meant that few labs were using the negatives.
>  
> Anyhow, after having reviewed the last QMPLS (Canada) survey committee 
> comments, I believe the committe would like a negative control run 
> with each patient tissue slide in order to evaluate background  (they 
> have used NCCLS guide pages as reference).  Incidentally we weren't a 
> part of the survey due to a technicality.
> Any help or advice would be appreciated.
>  
> L. Vermast
> Stratford, Ont.
>
>
>
>
>
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