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Related Experiment Video

Updated: Jul 15, 2025

Enzyme-linked Immunospot Assay ELISPOT: Quantification of Th-1 Cellular Immune Responses Against Microbial Antigens
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Optimization of Peripheral Blood Mononuclear Cell Processing for Improved Clinical ELISpot Assay Performance.

Xinyuan Li1, Shan He2, Jaya Thomas2

  • 1Janssen Research & Development LLC., 1400 McKean Road, Spring House, Pennsylvania, 19477, USA. xli323@its.jnj.com.

The AAPS Journal
|September 28, 2023
PubMed
Summary

Optimizing blood sample processing for cell and gene therapy studies is crucial. This research extends the viable processing window for ELISpot assays, improving monitoring of cellular immune responses and overcoming logistical challenges.

Keywords:
ELISpotPBMCSepMatecell therapycellular immunogenicityclinical trialgene therapyperipheral blood mononuclear cellsvaccine

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Area of Science:

  • Immunology
  • Biotechnology
  • Clinical Research

Background:

  • Cell and gene therapies show promise but face challenges from immune responses.
  • Monitoring cellular immunogenicity is vital for long-lasting clinical efficacy.
  • Enzyme-linked immunospot (ELISpot) assays are key for measuring immune responses but depend on peripheral blood mononuclear cell (PBMC) quality.

Purpose of the Study:

  • To investigate the impact of blood sample processing time on ELISpot assay performance.
  • To optimize sample processing conditions for robust cellular immune response assessment.
  • To develop strategies to mitigate challenges in PBMC preparation for clinical studies.

Main Methods:

  • Examined the effect of delayed blood sample processing on ELISpot assay signals.
  • Optimized various sample processing parameters to recover ELISpot signals.
  • Implemented mitigation strategies for granulocyte contamination in antigen-specific response detection.

Main Results:

  • Overnight delays in blood sample processing led to a loss of ELISpot signals.
  • Optimized processing successfully recovered ELISpot signals for samples processed within 32 hours.
  • Mitigation strategies were identified to address granulocyte contamination issues.

Conclusions:

  • Extended sample processing window for ELISpot assays in cell/gene therapy clinical studies.
  • Provided solutions for logistical challenges in whole blood sample shipment and PBMC preparation.
  • Enhanced the reliability of monitoring cellular immune responses in therapeutic development.