ISSN:
1573-0778
Keywords:
calcium phosphate
;
CHO-K1
;
cytosolic calcium signaling
;
HEK 293
;
laser scanning confocal microscopy
;
transient transfection
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Medicine
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
Abstract For the controlled production of recombinant proteinsin mammalian cells by transient transfection, it maybe desirable not only to manipulate, but also todiagnose the expression success early. Here, weapplied laser scanning confocal microscopy to monitortransfection induced intracellular Ca2+responses. We compared Chinese hamster ovary (CHO K1)versus human embryo kidney (HEK) 293 cell lines, whichdiffer largely in their transfectability. An improvedcalcium phosphate transfection method was used for itssimplicity and its demonstrated upscale potential.Cytosolic Ca2+ signaling appeared to inverselyreflect the cellular transfection fate. Virtually allCHO cells exhibited asynchronous, cytosolicCa2+ oscillations, which peaked 4 h afteraddition of the transfecting solution. Yet, most ofthe HEK cells displayed a slow and continuousCa2+ increase over the time of transfection. CHOcells, when exposed to a transfection-enhancingglycerol shock, strongly downregulated their Ca2+response, including its oscillations. When treatedwith thapsigargin, a Ca2+ store depleting drug,the number of successfully transfected CHO cells was significantly reduced. Our result points tointracellular store release as a critical componentfor the transfection fate of CHO cells, and its early detection before product visualization.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1008150402616
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