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Related Experiment Videos

Impact of different cell isolation techniques on lymphocyte viability and function.

A B Klein1, S G Witonsky, S Ansar Ahmed

  • 124061-0442,

Journal of Immunoassay & Immunochemistry
|February 3, 2006
PubMed
Summary

Choosing the right lymphocyte isolation method is crucial for accurate immunological assays. Mechanical dissociation techniques impact lymphocyte function, with screen and plunger methods preserving T-cell proliferation better than stomacher or slide methods.

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Area of Science:

  • Immunology
  • Cell Biology
  • Biotechnology

Background:

  • Lymphocyte isolation techniques significantly impact immunological assay outcomes.
  • Comparative assessment of lymphocyte isolation methods, particularly regarding cell recovery, viability, and function, is lacking.
  • Mechanical dissociation methods for lymphoid organs require thorough evaluation.

Purpose of the Study:

  • To determine the effect of different mechanical tissue dissociation methods on lymphocyte viability and function.
  • To compare four distinct mechanical dissociation techniques: metallic screen, sheer force slide, commercial stomacher, and plunger-screen.
  • To evaluate the influence of isolation methods on cell recovery, morphology, apoptosis, necrosis, and mitogen-induced proliferation.

Main Methods:

Related Experiment Videos

  • Lymphoid organs (spleen and thymus) were mechanically dissociated using four methods.
  • Cell viability was assessed using trypan blue exclusion and 7-amino-actinomycin D (7-AAD) staining.
  • Lymphocyte proliferation was measured via Alamar Blue assay, and surface antigen expression via flow cytometry.

    • Main Results:

    • No significant differences were observed in lymphocyte viability, morphology, apoptosis, or necrosis across all tested techniques.
    • Lymphocyte surface antigen expression remained comparable among the different isolation methods.
    • Mitogen-induced splenic T-cell proliferation was significantly higher with metallic screen and plunger-screen methods compared to sheer force slide and commercial stomacher.

    Conclusions:

    • Lymphocyte recovery, morphology, and viability are generally unaffected by the mechanical dissociation techniques evaluated.
    • Lymphocyte function, specifically mitogen-induced proliferation, is negatively impacted by the sheer force slide and commercial stomacher isolation methods.
    • The choice of mechanical dissociation method is critical for preserving lymphocyte function in immunological studies.