Rare Event Analysis in Flow Cytometry: Design Control and Validation Aligned with CLSI H62

Life Sciences, Clinical Trials,
  • Wednesday, April 05, 2023

Rare event analysis by flow cytometry is relatively common in research settings, in clinical laboratories and during drug discovery and development. For example, in research and drug discovery environments, the measurement of intracellular cytokines and intracellular signaling events (i.e., protein phosphorylation) are some of the more common rare event assays. Whereas in clinical laboratories, assays for rare events often include the measurement of CD34+ hematopoietic stem cells, paroxysmal nocturnal hemoglobinuria (PNH) and minimal residual disease (MRD) of blood cancers. During drug development and clinical trial evaluation of new therapeutic modalities, rare event methods might also include the monitoring of CAR cellular therapies and the efficacy of cell-depleting therapies.

Flow cytometry is considered an ideal technology for the detection of rare populations of cells. Not only can multiple characteristics on individual cells be measured, but also a large number of cells can be evaluated in a relatively short amount of time. However, as the technology moves out of research and non-regulated settings into clinical laboratories and the regulated phases of preclinical and clinical drug development, concerns related to how to develop, validate and implement robust assays for rare event measurements arise.

This webinar will address those concerns by introducing the concept Design Control and how its application can facilitate assay development and validation. Design Control is a structured process which is applied to the conduct of product development activities. While Design Control is mandatory in regulated industries, it can be easily adopted in non-regulated settings to increase successful outcomes.


Using the Design Control framework, the webinar will address aspects to consider when designing and validating flow cytometric methods that report rare subsets. The first step will be to define the elements that contribute to the measurement and other aspects of Measurement Assurance. Factors influencing Measurement Assurance in quantitative flow cytometry include the instrument and assay, as well as the processes surrounding monitoring, reporting and documentation of both the instrument and the assay. When measuring and reporting rare events, each of these elements has unique requirements as well as enhanced importance. The next step would be to define the requirements of the assay and its context-of-use. Only then should the assay development, optimization and validation phase be initiated.

Case studies illustrating the importance of instrument set-up, panel design and assay optimization on the final assay performance will be presented. The validation parameters critical to the validation of rare event assays such as assay specificity and assay sensitivity will be highlighted. Case studies will also demonstrate how the validation and application of a lower limit of quantification (LLOQ) can improve the translatability of the assay. In addition, considerations for reporting data near, or below, the LLOQ will be discussed.

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Register today and learn about rare event analysis in flow cytometry using design control and validation approaches aligned with CLSI H62.


Virginia Litwin, Eurofins BioPharma Services

Virginia Litwin, PhD, Director of Scientific Affairs, Eurofins BioPharma Services

Virginia Litwin is a thought-leader in validation and standardization for flow cytometry. Bringing “Cytometry from Bench-to-Bedside” has been the focus of her professional activities since 1999 when she started working in translational medicine at Bristol-Myers Squibb.

She co-founded the Flow Cytometry Community within the American Association of Pharmaceutical Scientists (AAPS). This group published the first papers on flow cytometry method and instrument validation. She was a councilor for the International Society for the Advancement of Cytometry (ISAC) and the International Clinical Cytometry Society (ICCS). She serves on the ICCS Advocacy Committee whose mission is to interface with regulatory agencies and has been an invited speaker at FDA/NIST on many occasions.

Virginia is the Chair of the Document Development Committee for the guidance document, CLSI H62- The Validation of Assays Performed by Flow Cytometry. She edited the book, Flow Cytometry in Drug Discovery and Development, as well as journal special issues: Journal of Immunological Methods – Flow Cytometry Biomarkers and Translational Medicine (2011); Cytometry Part B: Clinical Cytometry – Receptor Occupancy (2015) and Cytometry Advancing Next Generation Drug Development (2021).

After obtaining a PhD in virology/immunology from the University of Iowa, Virginia joined Lewis Lanier at DNAX as a postdoctoral fellow where she identified the KIR receptor, KIR3DL1 (CD158E1). She has held leadership roles in several contract research organizations. Currently she is Scientific Affairs Director at Eurofins BioPharma Services.

Message Presenter

Who Should Attend?

Pharmaceutical and biotech professionals working in the development of cell and gene therapies such as:

  • Scientists
  • Managers/Directors of Clinical Development
  • CSOs
  • Program/Project Leads

What You Will Learn

Attendees will:

  • Learn how the application of Design Control during assay design and development will lead to successful outcomes
  • Be introduced to the concept of Measurement Assurance and how it applies to quantitative flow cytometry
  • Learn the impact of instrument set-up, panel design and assay optimization on the validation of the assay specificity and sensitivity
  • Learn to approach the assay specificity and sensitivity validation in alignment with the Clinical Laboratory and Standards Institute (CLSI) H62 Guideline for the Validation of Assays Performed by Flow Cytometry

Xtalks Partner

Eurofins Viracor BioPharma Services

Truly innovative therapeutics are built on advanced science, disruptive technology, and reliable partnerships.  From vaccines to cell and gene therapies, Eurofins Viracor BioPharma is a unique development partner with proven expertise across a broad array of industry-leading analytical technologies, providing integrated testing solutions to help advance your pioneering treatments to patients faster.

We are a team of highly experienced scientists and professionals, providing complex testing and assay development for clinical research. We offer broad experience in molecular infectious disease and immunogenicity testing, along with immune response monitoring, vaccine safety/efficacy assessment, and disease-associated biomarker testing.

Everyday our team works collaboratively with our sponsor partners to solve some of the toughest bioassay challenges, leveraging the unique combination of state-of-the-art technology, and specialized expertise, that drug development companies have come to rely on for their clinical study needs.

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