Detection of residual T-cells in T-cell depleted bone marrow: a comparison of immunofluorescence and limiting dilution culture techniques.
The quantitation of T lymphocytes in human bone marrow was assayed by a limiting dilution technique. Phytohaemagglutinin-responsive T cells were maintained in interleukin-2 containing medium with feeder cells for 16 days of culture. Each well was then scored microscopically as positive or negative, or by H3 thymidine incorporation as a measure of DNA synthesis. Minimum Chi square statistics were used to establish the best line of fit to calculate the T lymphocyte frequency in the sample. This method for enumeration of cells was applied to untreated samples of bone marrow, and marrow treated with monoclonal anti-T cell antibodies and rabbit complement. T cells in parallel samples were enumerated by membrane immunofluorescence and flow cytometry. The limiting dilution assay using microscopic analysis produced data consistent with single-hit kinetics, and was the method of choice for detecting T cells in T cell depleted human bone marrow.
Williamson
I K
IK
Dickinson
A M
AM
Proctor
S J
SJ
D000818:Animals; D001772:Blood Cell Count; D001854:Bone Marrow Cells; D005455:Fluorescent Antibody Technique; D006801:Humans; D007201:Indicator Dilution Techniques; D008212:Lymphocyte Depletion; D011817:Rabbits; D013601:T-Lymphocytes