Hexosaminidase A and Total Hexosaminidase, Serum
Method Description Describes how the test is performed and provides a method-specific reference
The hexosaminidases are among the more active of the lysosomal enzymes, which hydrolyze derivatives of beta-D-N-acetylglucosamine and beta-D-N-acetylgalactosamine. Natural substrates are certain sphingolipids (ie, GM2) in which acetylgalactosamine is the terminal monosaccharide. The two hexosaminidase isoenzymes, A and B, differ in their electrophoretic mobility and heat stability. Hexosaminidase A moves toward the anode and is heat labile, while hexosaminidase B moves toward the cathode and is heat stable.
The procedure is performed on a two channel Autoanalyzer equipped with two fluorometers. The substrate used is 4-methylumbelliferyl-N-acetyl-beta-D-glucopyranoside (4-MUF-acetamido-2-deoxy-beta-D-glucopyranoside) from which the fluorescent compound, 4-methylumbelliferone, is liberated by both hexosaminidases.
After sample pickup, a sample splitter divides the patient serum. One-half of the sample travels through a 37 degree C mixing coil with substrate. The other half is first directed through a 53.25 degree C heating coil for 5.5 minutes and is then mixed with substrate and sent through the 37 degree C mixing coil. The hexosaminidase A fraction is destroyed in the heated sample, leaving only hexosaminidase B to react with the substrate. The unheated sample provides the total hexosaminidase (A and B). The reactants are pumped through a fluorometer, and the intensity of the fluorescence is converted to peaks of varying heights on a recorder. Sample peaks are compared to that of a 100 microM beta-methylumbelliferone standard to quantitate both the total and the "B" fraction. The percentage of the "A" fraction that was inactivated by heating is calculated based on these results. The difference in heat inactivation is used to fractionate hexosaminidase activities.(O'Brien JF: Lysosomal storage diseases. In Tietz Textbook of Clinical Chemistry. Edited by CA Burtis, ER Ashwood. Second edition. Philadelphia, PA, WB Saunders Company, 1994. pp 2149-2160)
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Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Tuesday; 10:00 a.m.