In today’s era of a modern clinical laboratory, a very important step with respect to the quality of the entire process is method validation or verification. So, in the start-up phase of any new laboratory or new tests, it is fundamental to verify or validate new process flow, new techniques, and new test parameters. Thus, validation promotes an environment emphasizing patient safety and quality patient care. Additionally, method validation is a requirement of laboratory regulations (ISO 15189:2012) which state that performance of new methods be verified prior to reporting patient test results, and periodic assessment of accuracy and precision must occur. Therefore, the accreditation bodies like College of American Pathologists (CAP) & National Accreditation Board for Testing & Calibration Laboratories, India (NABL) require that laboratories validate/verify the performance of tests.
During the laboratory accreditation process, the most frequently cited deficiency involves inappropriate verification protocol. The validation process for test methods, as well as the instrumentation that is used to perform the analysis, should have a well-established system of qualification phases: IQ (installation qualification), OQ (operational qualification), and PQ (performance qualification) protocols. In particular, PQ is supposed to ensure continued satisfactory performance of an instrument under actual running conditions during the daily run. The main principles to the PQ laboratory test validation or verification protocol are accuracy, precision, reference interval, detection limit, analytical measurement range (AMR), and clinically reportable range (CRR), etc. Laboratory regulations require that performance for any new method be verified prior to reporting patient’s test results.
Reference intervals are the most common decision support tool for interpreting pathology results. One can adopt reference limits from any of the sources, i.e., manufacturer suggested, reference laboratory, published articles, neighboring laboratory, or previous reference limits in the same laboratory.
While verifying of analytic accuracy, the agreement between the test result and true result is done in mainly two ways, i.e., comparison of results between new method and reference method; and results using a new method on certified reference materials (recovery). The first approach is the most commonly used. For this, 20 samples within testing range by both new and comparative methods are run and checked whether the average bias between the two methods is within allowable limits or not. The data is then compared with the deming regression model or passing- bablok regression model. Precision implies repeatability, which means analyze repeatedly to determine variation. Precision is the closeness of agreement between independent test results obtained under stipulated conditions. It is expressed quantitatively in terms of imprecision, standard deviation (SD) or coefficient of variation (CV percent) of the set of replicate measurements.
There are two types of precision claims, within – run precision and within – laboratory precision. Limit of detection (LOD) is the smallest amount that the method can detect to determine presence or absence of analyte. This involves two steps, determination of values obtained with blank samples, and values obtained with low-level positive samples. The term analytical sensitivity is sometimes used interchangeably with LOD or ‘lower the LOD’ in many diagnostic laboratories. The range of analyte values that a method can directly measure on the specimen, without any dilution, or other pretreatment, not part of the usual assay process. AMR must be verified before a method is introduced and checked at least every 6 months (and after recalibration or major maintenance) while in use.
CRR is the range of analyte values that are reported as a quantitative result, allowing for specimen dilution or other pretreatments used to extend the actual AMR. CRR is the range of values that can be reported with sample dilution. The laboratory should define its own AMR and CRR charts.