Principle Of Method:
Release of 35SO4 from [N-sulphonate-35S] heparin is measured after low voltage electrophoresis to separate the inorganic sulphate from substrate. An alternative fluorogenic substrate, 4-methylumbelliferyl-á-D-sulphoglucosamide, is available and its use is being evaluated in the laboratory.
Uses And Limitations Of The Method:
The assay is used for determining activity of heparan sulphamidase which is the primary deficiency in Sanfilippo disease type A (MPS IIIA). The natural function of this enzyme is to cleave sulphate groups from the nitrogen atom of glucosamine residues in heparan sulphate. In its absence there is storage of heparan sulphate in the tissues and excess excretion in the urine. The deficiency can be demonstrated in white cells or fibroblasts, although the former may sometimes give equivocal results and the SAS laboratory recommends use of fibroblasts for this assay. MPS IIIA is the most common form of MPS III in the UK. Prenatal diagnosis is possible by assay of chorionic villi or cultured amniotic cells; when amniocentesis is performed investigation of the GAG pattern in the supernatant fluid may assist in the diagnosis. Heterozygote detection is not routinely offered although partial deficiency of the enzyme (in fibroblasts only) can be demonstrated in some carriers.
Specimen Requirements:
For preliminary testing, blood. 5 ml lithium heparin (orange capped tube) unseparated and unfrozen. Send at room temperature to arrive at the laboratory within 24h of venepuncture. For follow-up, fibroblasts cultured from a skin biopsy may be needed. Biopsy material should be collected aseptically into a sterile bottle containing tissue culture medium (available from the laboratory), and sent at room temperature to arrive within 24 hours. Biopsies for tissue culture should not be frozen, Fibroblast cultures established in other laboratories should be sent in plastic 25 cm2 flasks filled with medium.
THE LABORATORY RECOMMENDS USE OF A COURIER SERVICE OR ROYAL MAIL SPECIAL DELIVERY FOR SENDING ALL SPECIMENS TO THE LABORATORY.