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An improved technique for separating rosetted from non-rosetted lymphocytes.

D R Ownby, J McCullough

    Journal of Immunological Methods
    |February 11, 1983
    PubMed
    Summary
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    A new method simplifies lymphocyte separation by eliminating cell resuspension. This technique rapidly isolates rosetted lymphocytes using Percoll density gradients for highly purified cell populations.

    Area of Science:

    • Immunology
    • Cell Biology
    • Biotechnology

    Background:

    • Lymphocyte separation is crucial for immunological studies.
    • Existing methods for separating rosetted lymphocytes can be labor-intensive and require mechanical resuspension.
    • Efficient isolation of specific lymphocyte subsets is essential for research and clinical applications.

    Purpose of the Study:

    • To describe an improved technique for separating rosetted from non-rosetted lymphocytes.
    • To eliminate the need for mechanical resuspension of cell pellets before density gradient separation.
    • To provide a rapid, reproducible method for obtaining highly purified lymphocyte populations.

    Main Methods:

    • A novel density gradient centrifugation technique using Percoll (density 1.078 g/ml).

    Related Experiment Videos

  • Direct overlay of the rosette-containing cell pellet with Percoll, bypassing mechanical resuspension.
  • Separation of human T lymphocytes (rosetted with neuraminidase-treated sheep erythrocytes) from other mononuclear cells.

    • Main Results:

    • Non-rosetted cells floated to the Percoll surface, while rosetted cells remained in the pellet.
    • Resulting T cell populations showed minimal contamination (<0.3% monocytes, <2% B cells).
    • The non-T cell population contained less than 3% T cells.

    Conclusions:

    • The described technique offers a significant improvement over existing methods for lymphocyte separation.
    • This simplified approach is rapid, easily reproducible, and yields highly purified cell populations.
    • The method is effective for isolating specific lymphocyte subsets, such as human T lymphocytes.