Flow cytometry assay modifications: Recommendations for method validation based on CLSI H62 guidelines

Area of Science:

• Clinical Laboratory Science
• Biotechnology
• Analytical Chemistry

Background:

• The Clinical and Laboratory Standards Institute (CLSI) H62 guideline (2021) offers recommendations for flow cytometry assay validation.
• CLSI H62 addresses platform workflow, quality systems, instrument setup, assay development, and analytical method validation.
• The guideline also includes recommendations for validating modified flow cytometric assays.

Purpose of the Study:

• To expand on CLSI H62 recommendations for validating modified flow cytometry assays.
• To discuss the impact of various assay modifications on overall assay performance.
• To provide practical recommendations for laboratory-initiated assay revisions and validation strategies.

Main Methods:

• Building upon existing CLSI H62 recommendations.
• Analyzing the impact of different types of assay modifications on assay performance.
• Proposing validation parameter evaluations based on the nature of the modification.
• Discussing the influence of assay modification on the assay's intended use.

Main Results:

• Different assay modifications have varying impacts on flow cytometry assay performance.
• Specific validation parameters should be evaluated based on the type and extent of the assay modification.
• Expert opinion provides rationale for minor deviations from CLSI H62 for laboratory-initiated revisions.

Conclusions:

• This manuscript offers practical recommendations for validating modified flow cytometry assays, complementing CLSI H62.
• The provided guidance aims to assist laboratories in navigating assay revisions while considering performance and intended use.
• These recommendations are intended as practical guidance and not as a restrictive or universally sufficient standard for regulatory compliance.