This article looks at IHC sensitivity and scoring across laboratories in 21 countries. It highlights the need for standardization, stringent quality controls and ongoing quality assurance programs.

Reagents can lose reactivity when they sit around in a refrigerator. Low-volume laboratories may have a reagent sitting in a refrigerator for six months before it passes its end date.One doesn’t know whether the reagent was left on the counter and lost some of its sensitivity.

The controls used in many laboratories are other tumors that were strongly positive. The problem with that is that strongly-positive tumors will stay strongly positive, or at least moderately positive, even if the reagent has lost some sensitivity. It’s the weakly-positive tumors, or those 2+ control cell lines, that will lose their reactivity if the reagent goes bad.

Ann D Thor, MD

In two trials, the patients with IHC 3+ and FISH-negative disease had a response rate of zero to trastuzumab-based therapy. In one trial, two patients with IHC 3+ and FISH-negative disease responded; those cases need to be reanalyzed to make sure they are indeed FISH-negative. Since blocks were never kept, they had to use stained slides, take the cover slips off, unstain the slides and then do the FISH test.

Patients with IHC 0 or 1+ and FISH-positive disease are HER2-positive. By the traditional HercepTest™, those patients would have never received trastuzumab. This problem with IHC is a function of the fixation of the tumor when it goes into formalin.

If we had frozen material from all patients at the time of diagnosis, IHC would be just fine. The problem occurs because formalin works by cross-linking proteins. HER2 is a protein that is progressively cross-linked. The longer the tissue is in the formalin, the more epitope cross-linking and masking occurs.

Cross-linking to other proteins covers up the HER2 epitope that is detected by the antibody. Dako has tried to introduce antigen retrieval to make that better. Although one can put all kinds of fancy scanners onto the tissue, if one does not control the fixation of the tissue, there is no way one can control what is tested.

Dennis Slamon, MD, PhD