Lymphocyte Blastogenesis Studied by Volume Spectroscopy

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Abstract
The blastogenesis of human blood lymphocytes was studied by measuring the distribution of cellular and nuclear volumes in concanavalin A (Con A)-stimulated leukocyte cultures with a modified Coulter counter. Agglutinated and adherent cells were dispersed by 3 h of incubation with .alpha.-methyl-mannoside and EDTA. Fresh suspensions contained 5-35% monocytes with an average cellular volume 2-2.5 times that of small lymphocytes. The nuclear volume of monocytes was similar to that of small lymphocytes within 10% and remained constant upon stimulation. In cultures given the optimal dose of Con A, 30 .mu.g/ml, measurable cell growth commenced within 8-12 h, whereas nuclear growth began some 6 h later, which is approximately 6-8 h before increased 3H-thymidine incorporation could be detected. The average volume of responding cells grew by a factor of about 4 until mitosis, which started after around 48 h, whereas the average nuclear volume increased by a factor of about 3. The percentage of cells responding by volume growth increased with mitogen concentration even for hyperoptimal concentrations (150 .mu.g/ml) which strongly suppressed 3H-thymidine incorporation. The results exemplify the type of information obtained by volume spectroscopy of leukocyte cultures. This method appears to have several advantages over conventional assays of immune response.