A new diagnostic assay for glycogen storage disease type II in mixed leukocytes

We have established a new method for the enzymatic diagnosis of glycogen storage disease type II (Pompe disease or acid maltase deficiency) using mixed leukocytes. The method employs glycogen and 4-methylumbelliferyl-α-d-glucopyranoside (4MU-αGlc) as substrates for measuring the lysosomal acid α-glucosidase (acid αGlu) activity, and incorporates acarbose to eliminate the interference of unrelated α-glucosidases (predominantly maltase-glucoamylase). It is shown that 3.0 μmol/L acarbose completely inhibits the maltase-glucoamylase activity at pH 4.0, but the lysosomal acid αGlu activity by less than 5%. With this method, we determined the acid αGlu activity in mixed leukocytes from 25 patients with glycogen storage disease type II (2 infantile and 23 late-onset cases), one GAA2/GAA2 homozygote and 30 healthy subjects. In the assay with glycogen as substrate, the addition of acarbose created a clear separation between the patient and the control ranges. In the assay with 4MU-αGlc as substrate, the two ranges were fully separated but remained very close despite the use of acarbose. The separation of the patient and normal ranges was improved considerably by taking the ratio of acarbose-inhibited over uninhibited activity. A GAA2/GAA2 homozygote was correctly diagnosed with 4MU-αGlc but misdiagnosed as patient when glycogen was used as substrate. We conclude that the inclusion of 3.0 μmol/L acarbose in the assays with glycogen and 4MU-αGlc substrates at pH 4.0 allows for the specific measurement of lysosomal acid αGlu activity in mixed leukocytes, thus enabling a reliable diagnosis of glycogen storage disease type II in this specimen.

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