The EPA2 protocol from CLSI. • Uses control material with assigned concentration (e g from external quality control) or certified reference materials. We are pleased to have a guest essay explaining the latest in Method Verification , specifically the newest version of the CLSI guideline EP15 on Method. CLSI document EPA2 describes the protocols that should be undertaken by the user to verify precision claims by a manufacturer. Precision claims by a.

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If this is true then using the principle of analysis of variance components: There should be at least one quality control QC sample in each run. The assessment is performed on at least two levels, as precision can differ over the analytical range of an assay.

The EPA3 committee felt that the patient comparison experiment had little value as it was, and that users who needed to perform a patient comparison experiment should consult CLSI EP9-A3 “Measurement procedure comparison and bias estimation using patient samples. Similarly the within-laboratory precision is estimated by measuring a sample 20 times over multiple days. If an outlier is found the pair should be rejected and the cause investigated and resolved before repeating the run.

Patient samples or control materials which have been repeatedly assayed with a measurement procedure felt to be substantially equivalent to the measurement procedure being evaluated may be appropriate if the user is interested in estimating bias relative to that measurement procedure. If the sample materials are appropriate, and target concentrations are available, the user can estimate the bias between the mean concentration calculated in the precision experiment pe15 to the target concentration of each of the materials.

The document includes tables to simplify the calculation clsu the verification limit. Clinical and Laboratory Standards Institute. Aa2 the calculated standard deviation is less than the verification limit, it is not statistically significantly larger than the published standard deviation, and the user has verified the published precision.

Selection and analytical evaluation of methods with statistical techniques. There is some variation in the terminology used but for the purposes of this discussion, repeatability, also known as within-run precision, is defined as the cosi of agreement between results of successive measurements obtained under identical conditions.

In order to compare the estimated repeatability to a claimed value we can calculate the critical or verification value using the equation:. For bias relative to the quality fp15 peer group, quality control materials with peer group values for the measurement procedure are appropriate.


Using the values from our example the mean of all the results is 1. If the estimated bias exceeds e;15 bias, it is not acceptable.

Journal List Clin Biochem Rev v. When using quality control samples, these should be different to those used to ensure the instrument is in control at the time of the assessment.

Statements of opinion in AACB publications are cosi of the contributors. The only requirement is that the assigned value must be available. Sometimes the calculated standard deviations may exceed the published values, and yet the true standard deviations are less than the published values.

CLSI EPA3: verification of precision and estimation of bias – Westgard

Reproducibility is at the other extreme and refers to the closeness of agreement between results of successive measurements obtained under changed conditions time, operators, calibrators, reagents, and laboratory.

Estimating Precision When evaluating the precision of an assay, the trivial approach for estimating repeatability for any given level is to perform 20 replicate analyses in a single run on a single day. If the mean concentration from the user’s experiment is beyond the verification interval, statistically significant bias exists.

Evaluation of precision performance of quantitative measurement methods. Patient samples, reference materials, proficiency testing samples, or control materials may be used as the test samples, provided there is sufficient sample material for testing each sample five times per run for five to seven runs. Part of the process of verifying or validating a method to confirm that it is suitable for use is an assessment of precision.

Here’s a brief description of the protocol. A spreadsheet for assisting with the fp15 described in this article is available from the AACB web-site. When undertaking the assessment the data must be assessed for outliers, which are considered to be present if the absolute difference between replicates exceeds 5. Evaluation of Results As alluded to above, EPA2 is generally used to verify that a method is performing as is claimed by the manufacturer.

Guest Essay

Precision claims by a manufacturer should be tested at at-least two levels, by running three replicates e15 five days. The following example relates to the verification of performance of calcium according to EPA2 using a five day protocol.


The first replicate on day 1 is 2. On day 1 the mean of the three replicates was 1. Unfortunately this approach is insufficient, as it tends to under-estimate repeatability, as the operating conditions in effect at the time may not reflect usual operating parameters. The most significant change is the creation of a relatively simple experiment that gives reliable estimates of a measurement procedure’s imprecision and its bias.

As alluded to above, EPA2 is generally used to verify that a method is performing as is claimed by the manufacturer. This article has been cited by other articles in PMC. Summing the square of the differences gives a total of 0. Calculation of the verification interval would be complicated, but the committee simplified it greatly by providing tables for the difficult-to-calculate quantities based on the number of replicate measurements per run, the number of runs, and the uncertainty of the target value.

Use of these materials is important in establishing the traceability of measurement procedures. The EPA2 protocol is similar except that the experiment is undertaken with three replicates over five days for at least two levels.

For example, on day 1 the average of the three values is 2. Thus the variance of the daily means is:. Acknowledgments Special thanks to Amanda Caswell for her careful review of the manuscript.

If the calculated standard deviations are less than the published values, the user has verified the claim. For a normal distribution the measure of imprecision is the standard deviation SD. Two or more appropriate materials should be tested in the precision experiment.

Evaluating Assay Precision

To allow for this possibility, the user calculates a “verification limit” based on the published standard deviation and the size of the user’s experiment. For the purposes of this example the results of only a single level are shown Table 1.

EPA2 should be used to validate a method against user requirements, and is generally used by reagent and instrument suppliers to demonstrate the precision of their methods. For the purposes of this discussion reproducibility will not be considered, as it involves multiple laboratories. For this, longer-term assessment is required.

Tools, Technologies and Training for Healthcare Laboratories.