Enzyme-linked immunosorbent assay for the detection of total cathepsin H in human tissue cytosols and sera.
An enzyme-linked immunosorbent assay (ELISA) was constructed for the determination of total human cathepsin H concentration in clinical samples. Utilising monoclonal and polyclonal antibodies, raised to human liver cathepsin H, the assay is able to detect a mature protein, a precursor molecule and enzyme-inhibitor complexes. The test system permits sensitive and reliable detection of analyte either in tissue cytosols or in sera. The detection limit is 2 ng/ml (n = 10, mean of zero standard +/- 3 SD). The average recovery of cathepsin H, added to the low content samples, was 95.3% +/- 1.8%. The within-run and between-run coefficient of variance (CV) varied from 2.3% to 8.9% and 12.7% to 16.4%, respectively, indicating satisfactory reproducibility of the method. The level of cathepsin H was defined in tissue cytosols of human heart, muscle and kidney and in sera from 30 healthy individuals. Additionally, cathepsin H was measured in sera from 55 patients with primary skin melanoma and from 42 patients with metastatic melanoma. The mean cathepsin H level was significantly higher for both groups of patients compared to normal sera level, being highest for metastatic melanoma patients.
Schweiger
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A
Jozef Stefan Institute, Department of Biochemistry and Molecular Biology, Ljubljana, Slovenia.
Stabuc
B
B
Popovíc
T
T
Turk
V
V
Kos
J
J
D000818:Animals; D000911:Antibodies, Monoclonal; D056655:Cathepsin H; D002403:Cathepsins / Q000032:analysis*; D003546:Cysteine Endopeptidases; D003600:Cytosol / Q000201:enzymology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D007526:Isoelectric Point; D008545:Melanoma / Q000201:enzymology / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D009362:Neoplasm Metastasis
D000911:Antibodies, Monoclonal; D002403:Cathepsins; D003546:Cysteine Endopeptidases; C534126:CTSH protein, human; D056655:Cathepsin H; C534127:Ctsh protein, mouse