A detailed flow cytometric method for detection of low-level in vivo red blood cell-bound IgG, IgA, and IgM

Wendy Beres, Geralyn M. Meny, Sandra Nance

Research output: Contribution to journalArticlepeer-review

Abstract

Flow cytometric methods are commonly used to analyze white blood cell surface antigen expression. We developed a flow cytometric method to detect red blood cell (RBC)-bound immunoglobulin (Ig)G, IgA, and IgM. RBCs were washed; incubated with fluorescein isothiocyanate (FITC)-conjugated anti-IgG, -IgA, or -IgM; washed; and analyzed on the flow cytometer. The method was optimized by determining the dilution of FITC-conjugated anti-IgG, -IgA, and -IgM providing the greatest amount of fluorescence when tested with Ig-coated RBCs and the least amount of fluorescence when tested with naive RBCs. Tannic acid was used to prepare Ig-coated RBCs. Cross-reactivity of FITC-conjugated anti-IgG, -IgA, and -IgM with Ig-coated RBCs was evaluated, and a reference range was established. Use of this method may assist in clinical evaluation of patients who present with hemolysis and a negative direct antiglobulin test.

Original languageEnglish (US)
Pages (from-to)161-169
Number of pages9
JournalImmunohematology
Volume32
Issue number4
StatePublished - 2016

Keywords

  • Autoimmune hemolytic anemia
  • Direct antiglobulin test
  • Flow cytometry
  • Immunoglobulins
  • Red blood cell
  • Tannic acid

ASJC Scopus subject areas

  • Immunology and Allergy
  • Hematology

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