A simple enzyme-linked immunoassay of human carbonic anhydrase I for the study of developing erythroid cells.
A simple and rapid enzyme-linked immunoassay for human erythrocyte carbonic anhydrase isozyme I was developed. The assay was found to be sensitive enough for the detection of nanogram amounts of the enzyme in incubation mixtures. The first incubation with anti-human carbonic anhydrase I IgG was carried out for 6 hours at room temperature. The second incubation of the enzyme was carried out in the presence of goat plasma C1q coupled with peroxidase for 1 h at room temperature. The enzymatic reaction was performed for 30 min using 2,2'-azino-di(3-ethyl-benzthiazoline sulfonate) as a substrate, and absorbance at 414 nm was recorded. Carbonic anhydrase I was assayed on the range of 1 to 200 ng/ml using this method. The levels of carbonic anhydrase I in K562 cells induced by erythropoietin and in other blood cells were determined. This enzyme-linked immunoassay has application for the study of developing erythroid cells.
Kondo
T
T
First Department of Medicine, School of Medicine, Hokkaido University, Sapporo, Japan.
Murakami
K
K
Isobe
H
H
Taniguchi
N
N
Kawakami
Y
Y
D000911:Antibodies, Monoclonal / Q000276:immunology; D002256:Carbonic Anhydrases / Q000097:blood* / Q000302:isolation & purification; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D004905:Erythrocyte Aging; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans
D000911:Antibodies, Monoclonal; D002256:Carbonic Anhydrases