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Cytotoxic T Cells-mediated Immune Response01:27

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Quantitative High-throughput Single-cell Cytotoxicity Assay For T Cells
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High-Throughput GLP-Capable Target Cell Visualization Assay for Measuring Cell-Mediated Cytotoxicity.

Anna Welter1,2, Srividya Sundararaman3, Ruliang Li4

  • 1Research & Development Department, Cellular Technology Limited, Shaker Heights, OH 44122, USA. anna.welter@web.de.

Cells
|April 27, 2018
PubMed
Summary

A new Target Cell Visualization Assay (TVA) offers a non-radioactive method to measure cell-mediated cytolysis. This computer image analysis technique matches the sensitivity of traditional assays while providing a traceable data audit trail for regulated environments.

Keywords:
ADCCCD8 cellsCFR-Part 11 complianceNK measurementTVAaudit trailscellular assay validationimmune monitoringmulti-color single cell imaging

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Area of Science:

  • Immunology
  • Cell Biology
  • Assay Development

Background:

  • Immune cells like Natural Killer (NK) and CD8 effector T cells eliminate virus-infected or malignant cells.
  • The chromium release assay, a radioactive method, has been the standard for measuring cell-mediated cytolysis.
  • Non-radioactive, flow cytometry-based methods offer alternatives for single-cell analysis.

Purpose of the Study:

  • To introduce and validate a novel Target Cell Visualization Assay (TVA) for measuring cell-mediated cytolysis.
  • To provide a non-radioactive, computer image analysis-based alternative to existing cytolysis assays.
  • To ensure the assay meets requirements for regulated environments by providing data audit trails.

Main Methods:

  • Development of two versions of the Target Cell Visualization Assay (TVA): one assessing viable (calcein-stained) target cells, and another assessing dead target cells (CFSE/PI staining).
  • Application of single-cell computer image analysis for target cell quantification.
  • Comparison of TVA sensitivity against the standard chromium release assay.

Main Results:

  • The TVA assays demonstrate comparable sensitivity to the traditional chromium release assay.
  • The assay provides a comprehensive data audit trail, including raw, analyzed, and quality-controlled images.
  • Two distinct TVA versions allow for the assessment of either viable or dead target cells.

Conclusions:

  • The Target Cell Visualization Assay (TVA) is a sensitive, non-radioactive method for measuring cell-mediated cytolysis.
  • TVA offers a valuable alternative for cell-mediated cytolysis assessment, particularly in regulated research and clinical settings.
  • The assay's image-based data audit trail enhances transparency and reproducibility.