DOSAGGIO FLUORIMETRICO NEI LEUCOCITI DI ALCUNI ENZIMI IMPLICATI NELLE MALATTIE DA ACCUMULO

The fluorimetric procedure (substrate: 4 methylumbelliferylglycoside or sulphate) for the microscale assay in leukocytes of a number of enzymes involved in inborn lipid and mucopolysaccharide storage diseases was set up. The following enzymes were studied: β glucosidase; β galactosidase; β N acetylglucosaminidase; β N acetylgalactosaminidase; β glucuronidase; arylsulphatase and α mannosidase. On each, the optimum pH, ionic strength, buffer, the possible requirement for detergents of other factors (albumin) and the saturating amounts of substrate were assessed. All examined enzymes (except for arylsulphatase and β glucosidase) could be easily assayed by the fluorimetric procedure, which proved to be much better than the corresponding colorimetric method for high reproducibility and extreme sensitivity. β Hexosaminidase, β glucuronidase (in the presence of 0.05% albumin) and α mannosidase, could be conveniently assayed using 1 to 5 μg non hemoglobin protein and β galactosidase with 5 to 30 μg. Thus, the leukocyte preparation obtained from 1 ml venous blood is sufficient for carrying out the precise determination of 5 to 10 of these enzymes.