Image analysis as a tool for quantitative enzyme determination at the cellular level: application for monocytic differentiation of the UM-384 cell line.
Image analysis has been used to determined enzyme activity at the cellular level in individual smeared cells. The counterstains used to visualize smeared cells were chosen to avoid overlap with the chromogene. The amount of the reaction product was quantified by computerised scanning cytophotometry when the conditions of incubation, time and temperature of the reaction, and substrate concentration varied. Under optimal conditions for time, temperature and substrate concentration, a linear relationship was found between enzyme activity determined on smeared cells and in cell lysate. Using these defined conditions, differentiation of UM-384 cells was studied by measuring enzyme activity. After a monocytic differentiation process, induced by sodium butyrate, non-specific esterase cell activity was compared either with differentiation markers (HLA-DR, plasminogen activator inhibitor type 2 and lysozyme) or with markers of proliferation (DNA content) or functional properties (nitroblue tetrazolium reduction and phagocytosis). The results show that, using image analysis, non-specific esterase seems to be a useful means for the assessment of monocytic differentiation whereas myeloperoxidase is not. More generally, quantification of enzyme activity at the cellular level using image analysis can be applied to the study of the differentiation process and may help in the classification of leukemic cells.
Champelovier
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P
Laboratoire de cytologie quantitative, Faculte de Medecine de Grenoble, La Tronche, France.
Seigneurin
D
D
Bosc
D
D
Kolodie
L
L
D000135:Acid Phosphatase / Q000032:analysis; D000943:Antigens, Differentiation / Q000032:analysis; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000032:analysis; D002454:Cell Differentiation / Q000502:physiology*; D002461:Cell Line, Transformed / Q000166:cytology / Q000201:enzymology; D004950:Esterases / Q000032:analysis*; D006684:HLA-DR Antigens / Q000032:analysis; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D007124:Immunoenzyme Techniques; D009000:Monocytes / Q000166:cytology / Q000201:enzymology*; D009113:Muramidase / Q000032:analysis; D009195:Peroxidase / Q000032:analysis*; D017396:Plasminogen Activator Inhibitor 2 / Q000032:analysis; D013481:Superoxides / Q000032:analysis
D000943:Antigens, Differentiation; D006684:HLA-DR Antigens; D017396:Plasminogen Activator Inhibitor 2; D013481:Superoxides; D009195:Peroxidase; D004950:Esterases; D002265:Carboxylic Ester Hydrolases; C019873:chloroacetate esterase; D043182:Carboxylesterase; D000135:Acid Phosphatase; D009113:Muramidase