The comparison of impedance-based method of cell proliferation monitoring with commonly used metabolic-based techniques.
OBJECTIVE
Determination of cell numbers is a crucial step in studies focused on cytokinetics and cell toxicity. The impedance-based analysis employing electronic sensor array system xCELLigence System allowing label-free dynamic monitoring of relative viable adherent cell amounts was compared with the most utilized methods for relative quantification of viable cell numbers based on a determination of cellular metabolism.


METHODS
Colorimetric assay based on reduction of tetrazolium salt (MTT) by mitochondrial enzymes and chemiluminiscent assay based on intracellular adenosine triphosphate (ATP) determination were compared with the impedance-based system. Cell morphology was compared by microscopic evaluation. Normal human epidermal keratinocytes (NHEK) and normal human dermal fibroblasts (NHDF), together with 3T3 mouse fibroblast and HaCaT keratinocyte cell lines were employed.


RESULTS
The progress of cell growth curves obtained by different methods during 72 hours reflected cell type and cell seeding densities. The impedance-based method was found to be applicable for the determination of the cell proliferation of 3T3 fibroblasts, HaCaT and NHDF, since the comparison of this method with ATP and MTT determinations showed a comparable results. In contrast, the proliferation of NHEK measured by the impedance-based method did not correlate with other methodological approaches. This could be accounted to the specific morphological appearance of these cells.


CONCLUSIONS
The study shows the impedance-based detection of viable adherent cells is a valuable approach for cytokinetics and pharmacological studies. However, the specific morphological characteristics of cell lines have to be considered employing this method for determination of cell proliferation without using other reference methods.
Vistejnova
Lucie
L
CPN s.r.o, Dolní Dobrouc, Czech Republic.
Dvorakova
Jana
J
Hasova
Martina
M
Muthny
Tomas
T
Velebny
Vladimir
V
Soucek
Karel
K
Kubala
Lukas
L
D016475:3T3 Cells; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002452:Cell Count / Q000379:methods*; D002460:Cell Line; D049109:Cell Proliferation; D002478:Cells, Cultured; D003124:Colorimetry; D020405:Dermis / Q000166:cytology; D017097:Electric Impedance; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D015603:Keratinocytes / Q000166:cytology; D008163:Luminescent Measurements; D051379:Mice; D008928:Mitochondria / Q000201:enzymology / Q000378:metabolism; D010084:Oxidation-Reduction; D013778:Tetrazolium Salts / Q000378:metabolism*; D013997:Time Factors
D013778:Tetrazolium Salts; D000255:Adenosine Triphosphate