Open AccessEffect of electric field pulses on the viability and on the membrane‐bound immunoglobulins of LPS‐activated murine B‐lymphocytes: Correlation with the cell cycle
Author(s) -
Djuzenova Cholpon S.,
Sukhorukov Vladimir L.,
Klöck Gerd,
Arnold W. Michael,
Zimmermann Ulrich
Publication year - 1994
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/cyto.990150107
Subject(s) - antibody , microbiology and biotechnology , immunofluorescence , viability assay , electric field , cell cycle , biophysics , incubation , microsecond , cell , flow cytometry , stimulation , biology , chemistry , immunology , biochemistry , endocrinology , physics , optics , quantum mechanics
The effects, of microsecond electropulses (1–kV/cm) on the viability of murine B lymphocytes and on their binding of antibodies by surface immunoglobulin (Ig) were studied in relation to the cell cycle. Before electropulsing, cultures given 48 h mitogenic stimulation showed at least two cell subpopulations, which were distinguishable by their levels of surface‐Ig expression as assessed with FITC‐labelled antibodies against mouse Ig. The immunofluorescence intensity of cells in S and G2/M phases was higher than that of G0/G1 cells. After exposure of the mitogen‐stimulated lymphocytes to three exponentially decaying (time constantτ=5–40 μs) electric field pulses, dye exclusion assay showed that pulsing at 1 or 2 kV/cm (at 4°C or 20°C) did not cause permeabilization. Field strengths of 3, 4, or 5 kV/cm resulted in 20%, 45%, or 70% of dye‐permeable cells, respectively, if the pulsed cells were transferred to phosphate‐buffered saline on ice for 30 min. Incubation in full medium at 37°C for 30 min (“resealing”) significantly decreased the percentage of permeabilized cells. Electropulsed G0/G1 cells were not only more resistant to direct electric exposure (tolerated higher field strengths) than S+G2/M cells but also responded better to resealing. The surface Ig of lymphocytes pulsed at higher fields and low temperature (4 or 5 kV/cm, τ =5 μs, three pulses, 4°C) was less easily immunostained than in controls or in cells pulsed at 2 kV/cm or less. At 5 kV/cm those cells that were not permeabilized showed a greater reduction in immunostaining, especially if resealed. © 1994 Wiley‐Liss, Inc.