REGULATORY COMPLIANCE AND METHOD VALIDATION - Flow Cytometry in Drug Discovery and Development - page 248

TABLE 12.1 Method Validation

Validation Parameters Focus Value Added

Stability Biomarker stability

in assay matrix

Determine how to ship/store
clinical samples after sample
collection
Determinewhether stained samples
could be fixed and/or whether
fixed samples could be stored
short term prior to sample
acquisition

Intra-assay precision Within run

variability

Determine sample variability and
need for replicate analysis

Inter-assay precision Between run

variability

Determine the need to analyze all
samples in same analytical run
Establishes expected variation for
comparison to drug effect

Intra-subject variability Physiological

variability

Identify diurnal variations to avoid
or target in analysis

Instrument-to-instrument,
or lab-to-lab
comparison

Quality assurance
for alignment
in analysis

Ensure data generated from
different cytometers/
laboratories are comparable

Analyst-to-analyst
comparison

Training analyst(s) on
assay performance

Ensure data generated by different
analysts are comparable

well trained to consistently perform the assay to ensure consistency and high quality in
validation results. In some cases, tests for lot-to-lot variability are warranted and
accommodation would therefore be required for increased efforts that include
trending analysis.

12.3.2.2 Validation Assessment

Precision The variability in an assay can be statistically measured and can be
assessed using standard deviation and percentage coefficient of variation (%CV).
Precision of an assay is usually determined by assessing both intra- and inter-assay
variability. Intra-assay precision is the variability within an assay (i.e., variability
between replicates). Inter-assay variability is the variability between multiple assays
(i.e., variability between experiments). Apart from intra-assay and inter-assay
variability, inter-operator, inter-instrument, and inter-laboratory precision should
also be determined during assay validation. Evaluation of these variability factors
is described in the following sections.

INTRA - ASSAY PRECISION The intra-assay precision is an assessment of within-run
variability. Consideration must be given to the frequency of the cell population
being measured to determine acceptance criteria for intra-assay precision. When cell