Micromethod determination of N-acetyl-alpha-D-glucosaminidase in human leukocytes and study of some of its characteristics.
1. A very easy fluorescent micromethod is described to measure leukocyte N-acetyl-alpha-D-glucosaminidase in human leukocytes. The method presents the following advantages: possibility to measure low activity due to prolonged linearity, very moderate consumption of substrate, use of total homogenate without the need for protein removal by precipitation after the enzyme reaction, and minimal handling after the addition of the stopping reagent. 2. A number of enzyme characteristics as linearity (up to 16 hr), pH optimum (4), thermoresistance at different temperatures (up to 24 hr at room temperature) and at different pH values (no differences between 3-8) and solubilization by digitonin and Triton X-100 were determined. The characteristics (acid pH optimum, binding to Concanavalin A and stability for prolonged time at 0 and 23 degrees C) are in agreement with the known properties of most lysosomal enzymes. 3. The specific activity of the enzyme is very low and an unexplained low latency (about 30%) was observed.
Den Tandt
W R
WR
University of Antwerp (UIA), Faculty of Medicine, Wilrijk, Belgium.
Scharpé
S
S
D000118:Acetylglucosaminidase / Q000097:blood*; D003208:Concanavalin A; D004072:Digitonin / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007962:Leukocytes / Q000737:chemistry / Q000201:enzymology*; D008832:Microchemistry; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D012995:Solubility
D003208:Concanavalin A; D011092:Polyethylene Glycols; D017830:Octoxynol; D000118:Acetylglucosaminidase; D004072:Digitonin