We examined the activity of total N-acetyl-beta-hexosaminidase and of its isoenzyme forms, that represent different stages of the maturation of the lysosomal hydrolase. In both methods the enzyme catalyzes the separation of 4-methylumbelliferone, a fluorescent substance, from 4-methylumelliferyl-2-acetamido-2-deoxy-beta-D-glucopyranoside. We used Leaback's method for the fluorimetric assay of total enzyme, and Ellis's DEAE-cellulose microcolum chromatography for the assay of its components. We obtained a clear separation of each fraction. We will apply these methods in our further studies of children with renal damage, because hexosaminidase seems to be one of the most sensitive markers of tubular damage.