Research ArticlesLevey-Jennings Analysis Uncovers Unsuspected Causes of Immunohistochemistry Stain VariabilityVani, Kodela MS*; Sompuram, Seshi R. PhD*; Naber, Stephen P. MD, PhD†; Goldsmith, Jeffrey D. MD‡; Fulton, Regan MD, PhD§; Bogen, Steven A. MD, PhD*,†Author Information *Medical Discovery Partners LLC †Department of Pathology & Laboratory Medicine, Tufts Medical Center ‡Department of Pathology and Laboratory Medicine, Beth Israel Deaconess Medical Center, Boston, MA §PhenoPath Laboratories, Seattle, WA Supported by the National Cancer Institute and the National Center for Advancing Translational Sciences, both of the National Institutes of Health, under award numbers 1R44CA183203-1 and UL1TR001064. K.V., S.R.S., and S.A.B. have a patent interest in the antigen-coated glass beads technology, which is included as a secondary quality control along with tissue controls. The remaining authors declare no conflict of interest. Reprints: Steven A. Bogen, MD, PhD, Medical Discovery Partners LLC, c/o Department of Pathology & Laboratory Medicine, Tufts Medical Center, P.O. Box 115, 800 Washington Street, Boston, MA 02111 (e-mail: email@example.com). Applied Immunohistochemistry & Molecular Morphology: November/December 2016 - Volume 24 - Issue 10 - p 688-694 doi: 10.1097/PAI.0000000000000260 Buy Metrics Abstract Almost all clinical laboratory tests use objective, quantitative measures of quality control (QC), incorporating Levey-Jennings analysis and Westgard rules. Clinical immunohistochemistry (IHC) testing, in contrast, relies on subjective, qualitative QC review. The consequences of using Levey-Jennings analysis for QC assessment in clinical IHC testing are not known. To investigate this question, we conducted a 1- to 2-month pilot test wherein the QC for either human epidermal growth factor receptor 2 (HER-2) or progesterone receptor (PR) in 3 clinical IHC laboratories was quantified and analyzed with Levey-Jennings graphs. Moreover, conventional tissue controls were supplemented with a new QC comprised of HER-2 or PR peptide antigens coupled onto 8 μm glass beads. At institution 1, this more stringent analysis identified a decrease in the HER-2 tissue control that had escaped notice by subjective evaluation. The decrement was due to heterogeneity in the tissue control itself. At institution 2, we identified a 1-day sudden drop in the PR tissue control, also undetected by subjective evaluation, due to counterstain variability. At institution 3, a QC shift was identified, but only with 1 of 2 controls mounted on each slide. The QC shift was due to use of the instrument’s selective reagent drop zones dispense feature. None of these events affected patient diagnoses. These case examples illustrate that subjective QC evaluation of tissue controls can detect gross assay failure but not subtle changes. The fact that QC issues arose from each site, and in only a pilot study, suggests that immunohistochemical stain variability may be an underappreciated problem. Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.