Measurement of single-cell DNA synthesis by pokeweed mitogen-stimulated mononuclear cells with combined light and scanning electron microscopy

Abstract

The response of peripheral blood mononuclear cells to pokeweed mitogen (PWM) stimulation was investigated with combined light (LM) and scanning electron microscopy (SEM). The DNA content of the nucleus of a cell (measured by fluorescence) was compared directly with the diameter and morphological features (by SEM). It was found that upon PWM stimulation lymphocytes transform into blast cells without measurable increase of diploid DNA content (4C) were not seen. The results of DNA fluorescence measurements were comparable with [3H]thymidine uptake profiles, both having peak values on Days 4 and 5 after the start of stimulation. Lymphocyte-enriched populations stimulated by PWM showed a small but distinct population of cells with a diameter of approximately 4-5 micrometers and having a tetraploid DNA content on Days 3, 4 and 5 after the start of stimulation. This reflects the cells' ability to divide in response to mitogen stimulation. SEM showed no characteristic cell-surface morphology for blast cells with increasing DNA content.