Analysis of cathepsin D from breast tissues by capillary electrophoresis.
A rapid and simple method for analyzing cathepsin D in breast tissue based on capillary zone electrophoresis (CZE) is described. After incubating the tissue extracts with hemoglobin as a substrate, a specific peptide is cleaved and separated by CZE in less than 5 min. This peptide is not produced by the action of pepsin or trypsin. It is inhibited by the addition of pepstatin, a specific inhibitor for cathepsin D. Human hemoglobin acted as a better substrate than bovine hemoglobin. The test compared well to a radioimmunoassay. We have shown that peptides can be stacked by the use of acetonitrile. The method demonstrates the advantages of CZE for assay of proteolytic enzymes in general.
Shihabi
Z K
ZK
Department of Pathology, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, NC 27157, USA.
Kute
T E
TE
D001940:Breast / Q000201:enzymology*; D002402:Cathepsin D / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D019075:Electrophoresis, Capillary / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D010436:Pepstatins / Q000494:pharmacology; D012680:Sensitivity and Specificity; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D006454:Hemoglobins; D010436:Pepstatins; C014195:Streptomyces pepsin inhibitor; D002402:Cathepsin D; C031375:pepstatin