Potential for false-positive staining with a rabbit monoclonal antibody to progesterone receptor (sp2): findings of the uk national external quality assessment scheme for immunocytochemistry and fish highlight the need for correct validation of antibodies on introduction to the laboratory

This study focused on recent assessment results from the United Kingdom National External Quality Assessment Scheme for Immunocytochemistry and Fluorescence In-Situ Hybridisation breast hormone receptor module in which participants were asked to demonstrate progesterone receptors (PRs). The slides consisted of 3 infiltrating ductal breast carcinomas, previously classified as a high PR expresser, a moderate to low PR expresser, and a negative tumor. During this assessment, 2 commercial rabbit monoclonal antibodies, SP2 (Lab Vision/NeoMarkers, Fremont, CA), and 1E2 (Ventana, Tucson, AZ) were used by 15% of the participants. The SP2 rabbit monoclonal antibody showed false-positive and nonspecific staining on the previously established PR– tumor. This article highlights the necessity for all clinical laboratories to validate immunohistochemical methods and protocols when using newly marketed antibodies such as SP2; use composite tissue blocks with known levels of tumor expression such as a high, mid, and negative expression; and participate in internal and external quality assessment schemes, which can highlight potential technical issues in laboratory method

External quality assurance of HER2 FISH and ISH testing: three years of the uk national external quality assurance scheme

The American Society of Clinical Oncology/College of American Pathologists guidelines highlighted the critical importance of quality assurance in diagnostic testing for HER2. Unstained formalin-fixed, paraffin-embedded human breast carcinoma cell line sections were circulated to scheme participants on 9 occasions. “Reference laboratories” reported results for the HER2/chromosome 17 ratio and HER2 copy number for 3 years for each cell line, including 418 sets of results (1,671 results total). The number of participants was 62 laboratories in the final analysis. The mean and SD of results from reference laboratories demonstrated consistency during the 3-year period. The percentage of laboratories achieving “appropriate” results ranged from 45% to 88%, and the percentage achieving “inappropriate” results ranged from 5% to 29%. No consistent effect of the HER2 in situ hybridization testing method was demonstrated. Participation in external quality assurance schemes is a valuable mechanism for demonstrating and acquiring consistency for HER2 testing by in situ hybridization. Poor performance can be corrected via assistance and advice

HER2 testing in the UK: recommendations for breast and gastric in-situ hybridisation methods

These guidelines supplement existing guidelines on HER2 testing by immunohistochemistry and in-situ hybridisation(ISH) methods in the UK. They provide a specific focus on aspects of guidance relevant to HER2 ISH testing methods, both fluorescent and chromogenic. They are formulated to give advice on methodology, interpretation and quality control for ISH-based testing of HER2 status in common tumour types, including both breast and gastric tumours. The aim is to ensure that all ISH-based testing is accurate, reliable and timely

ALK Immunohistochemistry in NSCLC: Discordant Staining Can Impact Patient Treatment Regimen.

INTRODUCTION Diagnostic immunohistochemistry (IHC) is increasingly accepted as a screening method for anaplastic lymphoma receptor tyrosine kinase gene (ALK) rearrangements in NSCLC. We have sought to establish an ongoing robust external quality assessment process to gauge quality of anaplastic lymphoma kinase (ALK) IHC, which can have an impact on interpretation of patient samples. METHODS Unstained tissue and cell line samples were distributed on a quarterly basis to participating laboratories from 30 countries. Participants stained the slide using their routine diagnostic ALK IHC method and returned the slide along with their in-house control and methodology details. Slides were assessed by a team of pathologists and scientists. RESULTS Overall, there was a mean pass rate of 83% (range 71%-98%), with 38 variations in staining protocol. Methods included the following: the Roche D5F3 assay (65% of users, pass rate 93%); Novocastra 5A4 (15% of users, pass rate 65%); Cell Signaling Technology D5F3 (7% of users, pass rate 91%), and Dako ALK1 (5% of users, pass rate 50%). Choice of methodology directly affected final interpretation of distributed ALK-positive and ALK-negative NSCLC cases, which were correctly identified by 89% and 88% of participants, respectively. Antibody detection method was a contributing factor in false-negative staining results. The choice of laboratory controls was found to be unsuitable, and as such, in-house control recommendations are also provided. CONCLUSIONS ALK IHC is a robust screening technique, but there is concern that some diagnostic laboratories are using inadequate staining methods, which has a direct impact on final interpretation. External assessment helps provide laboratories with continued confidence in their ALK IHC testing

Standardization of negative controls in diagnostic immunohistochemistry: recommendations from the international Ad Hoc expert panel

Standardization of controls, both positive and negative controls, is needed for diagnostic immunohistochemistry (dIHC). The use of IHC-negative controls, irrespective of type, although well established, is not standardized. As such, the relevance and applicability of negative controls continues to challenge both pathologists and laboratory budgets. Despite the clear theoretical notion that appropriate controls serve to demonstrate the sensitivity and specificity of the dIHC test, it remains unclear which types of positive and negative controls are applicable and/or useful in day-to-day clinical practice. There is a perceived need to provide "best practice recommendations" for the use of negative controls. This perception is driven not only by logistics and cost issues, but also by increased pressure for accurate IHC testing, especially when IHC is performed for predictive markers, the number of which is rising as personalized medicine continues to develop. Herein, an international ad hoc expert panel reviews classification of negative controls relevant to clinical practice, proposes standard terminology for negative controls, considers the total evidence of IHC specificity that is available to pathologists, and develops a set of recommendations for the use of negative controls in dIHC based on "fit-for-use" principles

Standardization of positive controls in diagnostic immunohistochemistry:recommendations from the international ad hoc expert committee

Diagnostic immunohistochemistry (dIHC) has been practiced for several decades, with an ongoing expansion of applications for diagnostic use, and more recently for detection of prognostic and predictive biomarkers. However, standardization of practice has yet to be achieved, despite significant advances in methodology. An Ad Hoc Expert Committee was formed to address the standardization of controls, which is a missing link in demonstrating and assuring standardization of the various components of dIHC. This committee has also developed a concept of immunohistochemistry critical assay performance controls that are intended to facilitate methodology transfer and harmonization in dIHC. Furthermore, the committee has clarified definitions of IHC assay sensitivity and specificity, with special emphasis on how these definitions apply to positive controls. Recommendations for "best laboratory practice" regarding positive controls for dIHC are specified. The first set of immunohistochemistry critical assay performance controls for several frequently used IHC stains or tests is also developed and presented

Evolution of Quality Assurance for Clinical Immunohistochemistry in the Era of Precision Medicine. Part 3:Technical Validation of Immunohistochemistry (IHC) Assays in Clinical IHC Laboratories

Contains fulltext : 173084.pdf (Publisher’s version ) (Open Access)Validation of immunohistochemistry (IHC) assays is a subject that is of great importance to clinical practice as well as basic research and clinical trials. When applied to clinical practice and focused on patient safety, validation of IHC assays creates objective evidence that IHC assays used for patient care are "fit-for-purpose." Validation of IHC assays needs to be properly informed by and modeled to assess the purpose of the IHC assay, which will further determine what sphere of validation is required, as well as the scope, type, and tier of technical validation. These concepts will be defined in this review, part 3 of the 4-part series "Evolution of Quality Assurance for Clinical Immunohistochemistry in the Era of Precision Medicine.