Method Validation

Question 1

Why do we need method validation?

Answer 1

To demonstrate that methods are fit for purpose
Requirement of accreditation to ISO 17025 and 15189.
Provides confidence to its customers that the method can generate accurate and reliable data.

Question 2

Validation vs Verification

Answer 2

-Validation is carried out on methods that are completely new or incorporate major changes.
-Verification is undertaken by a lab adopting a method that has been previously validated.

Question 3

The method validation process

Answer 3

1. Define the analytical requirement
2. Develop/ Identify candidate method
3. Plan validation experiments
4. Carry out experiments
5. Use data to assess fitness for purpose
6. if the analytical requirement is met, produce a validation report.

Question 4

Defining the analytical requirement

Answer 4

Consider the customer requirement
Typical tests: Qualitative, Quantitative, Limit tests.

Question 5

Specificity (selectivity)

Answer 5

How discriminating is the method?
The effect of other components on the analysis of the component of interest
Determined by adding materials that may be encountered in samples

Question 6

Sensitivity

Answer 6

How much the response to an analyte changes in response to change of concentration.

Question 7

Limit of detection (LoD)

Answer 7

The smallest amount of analyte that can be detected and identified with an acceptable degree of uncertainty, but not necessarily quantified

Concentration which gives a response that is 3x the response for blank values.

Question 8

Limit of quantitation (LoQ)

Answer 8

The smallest amount of analyte that can be quantified with an acceptable degree of uncertainty.

Concentration which gives a response that is 10x the response for blank values.

Question 9

Analytical Range

Answer 9

Typically concentrations should be 120% of the specification for impure substances.

Question 10

Accuracy

Answer 10

The closeness of agreement between the value found for the analyte and the true value.

Question 11

Trueness

Answer 11

The closeness of agreement between a large number of analyte values and the true value.

Question 12

Bias

Answer 12

A measure of the total systematic error in a method.

Question 13

Determining trueness/bias

Answer 13

-Repeat analysis of a certified reference material
-Determination of recoveries from spiked samples.

Question 14

Precision

Answer 14

The closeness of agreement between a series of independent measurements obtained from multiple sampling of the same homogenous sample under the prescribed conditions.

Reflects random errors which occur in the method

Calculated by determining the standard deviation of the test results from repeat measurements.

Question 15

Repeatability

Answer 15

Same analyst & equipment; limited time span

Question 16

Intermediate of Precision

Answer 16

within laboratory variations: different days, analysts and equipment.

Question 17

Reproducibility

Answer 17

Different laboratories, analysts, samples and equipment; same method, extended time span.

Question 18

Uncertainty of measurement

Answer 18

The quantitative measure of the limits within which the true value of the analyte is expected to lie, to a specified level of confidence.

Question 19

Sources of Uncertainty

Answer 19

-Short term variability - repeatability
-Uncertainty in calibration equipment (proficiency test results)
-Uncertainty in reference materials
-Sampling effects