Antibody Validation for Flow Cytometry

The Presenters

Tomas Kalina, PhD
Childhood Leukemia Investigation
Prague, Czech Republic

Fridtjof Lund-Johansen, PhD
Oslo University Hospital
Norway

Anis Larbi, PhD
Singapore Immunology Network
Singapore

Kelly Lundsten
BioLegend
USA

Ruud Hulspas, PhD
Cellular Technologies Bioconsulting LLC
USA

Pablo Engel, PhD
Department of Cellular Biology and Pathology
University of Barcelona Medical School
Spain

Session Summary

Antibody validation in flow cytometry may be a better-established practice than in other antibody applications, particularly in clinical flow cytometry for diagnostic purposes. Antibody validation for flow cytometry was first addressed in 1982 by the Human Cell Differentiation Molecules, an organization that runs the Human Leucocyte Differentiation Antigens (HLDA) workshops to study leukocyte surface molecules. Through these HLDA workshops, information on novel molecules has accumulated rapidly and led to the characterization and formal designation of more than 400 molecules. This has allowed users to put a lot of trust in (albeit a relatively small number) vendors, who are expected to have validated and verified reactivity, specificity, selectivity, and sensitivity in relevant cell populations and cell types. However, best practice demands that the user finds and validates suitable antibodies for any flow cytometric assay, while there is no consensus on acceptable antibody validation methods. Consequently, the validity of flow cytometry results may be put in question.

This tutorial will present the current status of antibody validation from an academic perspective, as well as from the manufacturer perspective. We will discuss current irreproducibility issues and their relevance to cytometry. We will summarize the needs and challenges specific to flow cytometry, and we will discuss future methods needed for antibody validation.

CMLE Credit: 1.5