Estimation by limiting dilution analysis of human IL 2-secreting T cells: detection of IL 2 produced by single lymphokine-secreting T cells
We present here a culture method for the estimation, in human blood, of the number of lymphocytes that can respond to mitogen by producing interleukin 2 (IL 2). T cells are cultured at limiting dilutions with PHA or Con A in the presence of Epstein Barr virus-transformed human lymphoblastoid cells (EB-LCL), and supernatants are tested 3 days later for IL 2 content by a cell proliferation assay. The distribution of negative wells follows the expected Poisson single-hit relationship, suggesting that the assay is sensitive to single cells of a single limiting cell type. On average, 16.3% of peripheral blood mononuclear cells can produce IL 2 in such clonal cultures (mean of 12 determinations; SD = 5.6%). Surprisingly, irradiation (up to 2000 rad) of the titrated responder cell population diminishes the estimated frequencies by less than 50%. The ability to detect IL 2 levels in cultures containing only a single, nonproliferating T lymphocyte allows us to estimate the amount of IL 2 generated by an individual effector cell during a 3-day culture interval after mitogen stimulation. The average responding, irradiated T cell generates 0.92 pg of IL 2 (median) within 3 days. The method presented provides a straightforward way to provide independent estimates of responding cell number and of lymphokine production per cell in a variety of clinical situations.
- Research Organization:
- Boston Univ. School of Medicine, MA
- OSTI ID:
- 5805384
- Journal Information:
- J. Immunol.; (United States), Vol. 9
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
LYMPHOCYTES
CELL PROLIFERATION
LYMPHOKINES
SECRETION
MITOGENS
BIOLOGICAL EFFECTS
BLOOD
CELL CULTURES
CLONING
DOSE-RESPONSE RELATIONSHIPS
IRRADIATION
STEM CELLS
ANIMAL CELLS
BIOLOGICAL MATERIALS
BLOOD CELLS
BODY FLUIDS
CONNECTIVE TISSUE CELLS
GROWTH FACTORS
LEUKOCYTES
MATERIALS
ORGANIC COMPOUNDS
PROTEINS
SOMATIC CELLS
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