TY - JOUR
T1 - Determination of antibody content in live versus dead hybridoma cells
T2 - Analysis of antibody production in osmotically stressed cultures
AU - Reddy, Sridhar
AU - Bauer, Kenneth D.
AU - Miller, William M
PY - 1992/1/1
Y1 - 1992/1/1
N2 - Hybridomas are known to exhibit increased specific antibody production rated when subjected to environmental stress. Under these conditions, viability is low so that population‐average measurements do not properly reflect the state of viable cells. Even for flow cytometry, which gives a population distribution, special techniques must be used to discriminate between viable and nonviable cells. We describe the use of the vital stain ethidium monoazide (EMA) for independent measurement of intracellular antibody content in live and dead cells via flow cytometry. EMA is shown to be superior to light scattering techniques in identifying dead cells. We apply this technique to show that, in control batch culture, the specific antibody prodution rate and antibody content in live cells are constant during exponential growth, but decrease as cells enter the stationary phase. Antibody is retained in dead cells, but at a lower level than in live cells. We further show that, under hyperosmotic stress, the specific antibody production rate and antibody content in live both remain high during death phase. © 1992 John Wiley & Sons, Inc.
AB - Hybridomas are known to exhibit increased specific antibody production rated when subjected to environmental stress. Under these conditions, viability is low so that population‐average measurements do not properly reflect the state of viable cells. Even for flow cytometry, which gives a population distribution, special techniques must be used to discriminate between viable and nonviable cells. We describe the use of the vital stain ethidium monoazide (EMA) for independent measurement of intracellular antibody content in live and dead cells via flow cytometry. EMA is shown to be superior to light scattering techniques in identifying dead cells. We apply this technique to show that, in control batch culture, the specific antibody prodution rate and antibody content in live cells are constant during exponential growth, but decrease as cells enter the stationary phase. Antibody is retained in dead cells, but at a lower level than in live cells. We further show that, under hyperosmotic stress, the specific antibody production rate and antibody content in live both remain high during death phase. © 1992 John Wiley & Sons, Inc.
KW - antibody production
KW - flow cytometry
KW - hybridoma cells
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U2 - 10.1002/bit.260400811
DO - 10.1002/bit.260400811
M3 - Article
C2 - 18601202
AN - SCOPUS:0026742717
VL - 40
SP - 947
EP - 964
JO - Biotechnology and Bioengineering
JF - Biotechnology and Bioengineering
SN - 0006-3592
IS - 8
ER -