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QUALITY ASSURANCE, QUALITY CONTROL AND EXTERNAL QUALITY ASSESSMENT Anthony Rhodes, PhD, FRCPath, FIBMS (UK NEQAS Manager, 1992-2003) SUMMARY • Definitions of QA, QC and EQA • Relevance to Pathology and in particular Immunohistochemistry (IHC) and In Situ Hybridisation (ISH) • QA and Laboratory Accreditation • QC for IHC and ISH • EQA for IHC and ISH • What happens when you don’t have QA? • National and International guidelines for assuring quality • Technical and Clinical Validation • Standardisation QUALITY ASSURANCE • Quality assurance (QA) encompasses all measures taken to ensure the reliability of investigations, starting from satisfactory test sample selection, analysing it appropriately, to recording the result accurately and reporting it to the clinician for appropriate action, with all procedures being documented for reference (UK NEQAS, 1998) • In the clinical laboratory, QA is usually provided by participation in a Laboratory Accreditation Programme LABORATORY ACCREDITATION • Can be national or international, or both • Involves registering with an approved accreditation programme • Documentation of all the procedures in the laboratory, to include; • Range of tests, SOP’s, turn around times, results of audits, adherence to good laboratory practice and international guide lines (e.g ASCO/CAP guidelines), details of procurement, maintenance and servicing of all equipment, records of QC, participation and performance in EQA/Performance Testing (PT) etc • Visit by inspectors that thoroughly inspect the lab, interview the staff, review the documentation and record whether compliant or noncompliant, • Laboratory is given a time interval (e.g months), to address areas of noncompliance and provide evidence that it is now compliant • Laboratory is then give accreditation for a given time period QUALITY ASSURANCE (QA) Two of the main features of QA are; • Internal Quality Control (IQC) AND • External Quality Assessment Performance Testing (PT) (EQA), sometimes named INTERNAL QUALITY CONTROL (IQC) • The set of procedures undertaken for the continual evaluation of the quality of results on a day-to-day basis, in order to decide on whether they are reliable enough to be released (WHO, 1981, Leblanc et al., 1985) INTERNAL QUALITY CONTROL Factors to consider; • Fixation (pre-analytical) • The assay (analysis) • Evaluation of the results (post analysis) INTERNAL QUALITY CONTROL Factors to consider; • Fixation (pre-analytical) • The assay (analysis) • Evaluation of the results (post analysis) ST JOHN’S, NEWFOUNDLAND, CANADA Shipguy, Public domain, via Wikimedia Commons ST JOHN’S HOSPITAL NEWFOUNDLAND BY VERNE EQUINOX AT ENGLISH WIKIPEDIA, CC BY 3.0 JNCI; VOL 100, ISSUE 12 | JUNE 18, 2008 SOME OF THE MAIN ISSUES AT THE ST JOHN’S LAB • Newfoundland had no Laboratory Accreditation or sets standards for conducting clinical laboratory tests (unlike other Canadian provinces) • Rapid turn over of staff/shortage of pathologists (32% over years) • No one taking responsibility for overall QA of ER testing i.e pathologist or technician, • Technical issues e.g Inefficient antigen retrieval • Lack of understanding/training (CME, CPD) e.g • • • • • Invasive lobular carcinomas are ER+ve High cut points (thresholds) for a positive ER result Erroneous reporting e.g of on-slide control instead of the case Lack of sensitive controls e.g negative/low +ve/high positive Use of internal controls (normal glands should have ER+ve nuclei) • No participation in EQA ASCO/CAP GUIDELINES FOR ER/PR AND HER2 • Hammond EH, Hayes DF, Dowsett M, Allred DC, Hagerty KL, Badve S, Fitzgibbons PL, Frances G, Neal S Goldstein NS, Hayes M, Hicks DG, Lester S, Love R, McShane L, Miller K, Kent Osborne CK, Paik S, Perlmutter J, Rhodes A, Sasano H, Sweep FCG, Taube S, Torlakovic EE, Valenstein P, Viale G, Visscher D, Wheeler T, Williams RB, Wittliff JL, Schwartz JN, and Wolff AC American Society of Clinical Oncology/College of American Pathologists guideline recommendations for immunohistochemical testing of estrogen/progesterone receptors in breast cancer J Clin Oncol 2010; 28: 2784-2795 • Wolff AC, Hammond, MEH, Schwartz JN, Hagerty KL, Allred DC, Cote RJ, Dowsett M, Fitzgibbons PL, PL, Hanna WM, Langer A, McShane, Paik LS, Pegram MD, Perez EA, Press MF, Rhodes A, Sturgeon C, Taube SE, Tubbs R, Vance GH, van de Vijver M, Wheeler TM, Hayes DF American Society of Clinical Oncology/College of American Pathologists guideline recommendations for HER2 testing in breast cancer J Clinical Oncol 2007; 25: 118-145 TECHNICAL & CLINICAL VALIDATION VALIDATION OF BIOMARKERS FOR CLINICAL USE • The antibody suppliers, such as DAKO (Agilant) and Ventana (Roche) instigate internal validation procedures for clinically used antibodies • The checks by suppliers specializing in a wide range of predominantly research antibodies e.g Santa Cruz, traditionaly have not been tested as stringently as antibodies produced by companies such as Roche/Dako etc, supplying clinical hospital laboratories • Antibodies/Probes and kits for use in predictive testing go through more stringent testing than those used to assist in diagnosis TECHNICAL AND CLINICAL VALIDATION • Technical validation refers to the reproducibility of the antibody (IHC) or probe This ensures batch consistency, specificity (for the antigenic epitope) and sensitivity (signal to noise ratio) • Clinical validation, refers to the specificity and sensitivity of the marker for clinical cases If a new marker in research papers suggests it to be positive for a specific cancer e.g mesotheliomas -what % of previously diagnosed mesothelioma’s are +ve (sensitivity), -what % of tumours are true positives i.e they are mesotheliomas and not adenocarcinomas (specificity) • ASCO/CAP guidelines for HER2 recommend doing cross validation of specificity and sensitivity using complimentary assays (IHC v FISH) on 100 cases –strictly speaking this would be only technical validation, • Clinical validation would refer to the predictive value of HER2 and/or ER assays in predicting a favourable response to targeted therapies • e.g clinical validation for ER would be as in the paper by Harvey et al (1999), where it was shown that a 1% threshold included women likely respond to tamoxifen STANDARDISATION OF IHC AND ISH • This has been largely achieved by the use of automated systems (though control slides are still not standardized) • ADVANTAGE – ‘Helps’ ensure reproducibility on a day-to-day basis, which is essential for markers such as HER2 and ER and any marker that assessed in quantitative fashion and where this is predictive of likely response to targeted therapy, • DISADVANTAGES • – IHC and ISH becomes a ‘black box’ where lab staff have less understanding of the basic principles of the assay This is OK until something goes wrong and have to ‘trouble shoot’ or need to work up a new antibody • -Very expensive, suppliers frequently supply in a kit form, and rely on sales of expensive consumables BIBLIOGRAPHY • • • • Rhodes A, Miller K Internal Quality Control and External Quality Assessment of Immunocytochemistry, In: Bancroft and Gamble, 5th Edn 2002 Wick MR, Swanson PE Editorial: Targeted controls in clinical immunohistochemistry; a useful approach to quality assurance Am J Clin Pathol 2002; 117: 7-8 Moskaluk CA Editorial: Standardisation of clinical immunohistochemistry: Why, how and by whom? Am J Clin Pathol 2002; 118: 669-671 Sompurama SR et al, A Novel Quality Control Slide for Quantitative Immunohistochemistry Testing Journal of Histochemistry and Cytochemistry, Vol 50, 1425-1434, November 2002 BIBLIOGRAPHY (CONTINUED) • Rhodes A Invited Review: Developing a cell line standard for HER2 Cancer Biomarkers 2005; 1: 229-232 • Rhodes A, Borthwick D, Sykes R Al-Sam S, Paradiso A The use of cell line standards to reduce HER2 assay variation in multiple European cancer centres and the potential of automated image analysis to provide for more accurate cut points for predicting clinical response to trastuzumab Am J Clin Pathol 2004; 122: 51-60 • Rhodes A, Jasani B Quality Assurance in Immunohistochemistry Am J Surg Pathol 2003; 9: 1284-1285 • Rhodes A, Jasani B, Anderson E, Dodson AR, Balaton AJ Evaluation of HER-2/neu immunohistochemical assay sensitivity and scoring on formalin fixed and paraffin processed cell lines and breast carcinomas: A comparative study involving results from laboratories in 21 countries Am J Clin Pathol 2002; 118: 408-417 • Rhodes A, Jasani B, Couturier J, McKinley MJ, Morgan JM, Dodson AR, Navabi H, Miller KD, Balaton AJ A formalin fixed and paraffin processed cell line standard for quality control of immunohistochemical assay of HER-2/neu expression in breast cancer Am J Clin Pathol 2002; 117: 81-89