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Related Experiment Videos

Light scatter based lymphocyte gate--helpful tool or source of error?

E Krömer1, F Grossmüller

  • 1Ludwig Boltzmann Institute of Leukemia Research and Hematology, Vienna, Austria.

Cytometry
|January 1, 1994
PubMed
Summary

Standard flow cytometry lymphocyte gating may exclude T cells (CD3+). This can lead to inaccurate results, especially when analyzing T cell subsets and their CD4/CD8 ratios in research.

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Area of Science:

  • Immunology
  • Cell Biology
  • Biotechnology

Background:

  • Flow cytometry is a crucial technique for immune cell analysis.
  • Accurate gating is essential for reliable flow cytometry data.
  • Lymphocyte gating typically relies on light scatter properties.

Purpose of the Study:

  • To evaluate the efficacy of standard light scatter-based lymphocyte gating in flow cytometry.
  • To determine if TCR (CD3)-expressing cells are adequately captured by conventional gates.
  • To assess the characteristics of T cell subsets excluded by standard gating strategies.

Main Methods:

  • Analysis of 29 peripheral blood mononuclear cell (PBMC) samples using flow cytometry.
  • Comparison of T cell receptor (TCR) CD3-expressing cells against light scatter-based lymphocyte gates.

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  • Evaluation of CD4 and CD8 T cell subset proportions within and outside the defined gate.

    • Main Results:

    • A significant proportion of T cells (mean 7.9%) were found outside standard lymphocyte gates.
    • The excluded T cell population exhibited an altered CD4/CD8 ratio compared to gated cells.
    • Specific percentages of CD4+ (8.1%) and CD8+ (13.7%) cells were identified outside the gate.

    Conclusions:

    • Standard light scatter-based lymphocyte gating in flow cytometry may lead to the exclusion of T cells.
    • This exclusion can result in erroneous data, particularly impacting studies focused on T cell populations.
    • Researchers should consider alternative or supplementary gating strategies to ensure comprehensive T cell analysis.