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Updated: Jul 2, 2026

Detection and Enrichment of Rare Antigen-specific B Cells for Analysis of Phenotype and Function

Detection and Enrichment of Rare Antigen-specific B Cells for Analysis of Phenotype and Function

Published on: February 16, 2017

Immunophenotypic analysis of PNH cells.

Stephen J Richards¹, Peter Hillmen

¹Haematological Malignancy Diagnostic Service, Leeds, United Kingdom.

Current Protocols in Cytometry

|September 5, 2008

Summary

Paroxysmal nocturnal hemoglobinuria (PNH) diagnosis is improved by flow cytometry, which analyzes glycosylphosphatidylinositol (GPI) anchor expression. This method offers superior sensitivity and specificity compared to traditional lysis-based assays for PNH screening.

Area of Science:

Hematology
Immunology
Molecular Biology

Background:

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired stem cell disorder.
A gene mutation causes an inability to synthesize glycosylphosphatidylinositol (GPI) anchors.
Traditional diagnostic methods for PNH lack specificity and sensitivity.

Purpose of the Study:

To present flow cytometry protocols for PNH diagnosis.
To detail the immunophenotypic analysis of GPI-linked antigens on various blood cells.
To offer a four-color flow cytometry method for simultaneous analysis.

Main Methods:

Flow cytometry for GPI-linked antigen expression analysis.
Immunophenotypic analysis of red blood cells, granulocytes, and monocytes.

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Four-color flow cytometry for simultaneous granulocyte and monocyte analysis.

Main Results:

Flow cytometry provides rapid, sensitive, and specific detection of PNH.
Demonstrates the utility of GPI-linked antigen analysis for PNH diagnosis.
The four-color method allows for efficient, simultaneous analysis of multiple cell types.

Conclusions:

Flow cytometry has largely replaced traditional methods for PNH screening and diagnosis.
This technique offers a significant advancement in identifying PNH.
The described protocols facilitate accurate and efficient PNH diagnostics.