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Clinical flow cytometric reticulocyte analysis.

B H Davis1, N C Bigelow

  • 1Department of Pathology, Dartmouth Hitchcock Medical Center, Hanover, N.H.

Pathobiology : Journal of Immunopathology, Molecular and Cellular Biology
|January 1, 1990
PubMed
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Flow cytometry (FCM) offers a reliable method for reticulocyte analysis using stains like thiazole orange. This technique provides an early indicator of bone marrow engraftment in transplant patients.

Area of Science:

  • Clinical laboratory diagnostics
  • Hematology
  • Biotechnology

Background:

  • Flow cytometry (FCM) is a versatile technique for analyzing blood cell populations.
  • Accurate reticulocyte counting is crucial for assessing bone marrow function and erythropoiesis.
  • Existing methods for reticulocyte analysis have limitations in terms of speed, accuracy, and standardization.

Purpose of the Study:

  • To evaluate the efficacy of flow cytometric (FCM) analysis for reticulocyte enumeration in a clinical laboratory setting.
  • To compare the performance of different vital stains, specifically acridine orange (AO) and thiazole orange (TO).
  • To establish a reticulocyte maturity index (RMI) using FCM for early detection of bone marrow engraftment.

Main Methods:

  • Utilized flow cytometry (FCM) with various vital stains including acridine orange (AO) and thiazole orange (TO).

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  • Compared FCM reticulocyte counts with traditional microscopic methods, assessing correlation and precision (coefficient of variation < 5%).
  • Developed and validated a reticulocyte maturity index (RMI) based on the mean fluorescent intensity of TO.

    • Main Results:

    • Acridine orange (AO) and thiazole orange (TO) demonstrated excellent correlation with microscopic reticulocyte determinations.
    • Batch analysis of blood samples stored up to 96 hours showed reduced manpower requirements.
    • The reticulocyte maturity index (RMI) using TO proved to be the earliest indicator of marrow engraftment in bone marrow transplant patients, defining early, delayed, and failed engraftment patterns.

    Conclusions:

    • Clinical flow cytometric (FCM) reticulocyte analysis using vital stains like AO and TO is a precise and reliable method.
    • The reticulocyte maturity index (RMI) derived from FCM offers significant clinical value for monitoring bone marrow engraftment.
    • FCM reticulocyte analysis is poised to become the preferred method in clinical laboratories, pending standardization.