Principle Of Method: Two methods are available for this assay, both in use in the laboratory. The older method uses a tritiated disulphated trisaccharide prepared from chondroitin sulphate as substrate. The enzyme acts upon this to produce a monosulphated product that is separated on an anion exchange column and determined by counting. The newer method uses a fluorogenic substrate, 4-methylumbelliferyl-â-D-galactose-6-sulphate. The enzyme acts upon this to release sulphate and 4mu-â-galactoside, which is then acted upon by â-galactosidase to produce galactose and 4-methylumbelliferone. The latter is determined fluorimetrically.
Uses And Limitations Of The Method: The methods are used to diagnose Morquio disease type A (MPS IVA). Currently both methods are run in parallel for suspected patients while the more convenient fluorimetric assay is being evaluated. Since the latter method depends on the presence of â-galactosidase in the tissue being tested it is essential to assay this enzyme first to make sure activity is normal. Morquio A disease is one of the rarer mucopolysaccharidoses. Keratan sulphate is excreted in the urine, in addition to excess chondroitin 6-sulphate, but since this can be missed, and may be normalised in older patients, the enzyme assay is essential for unequivocal diagnosis of the disorder. The enzyme releases sulphate from galactose 6-sulphate residues in keratan sulphate and from N-acetylgalactosamine 6-sulphate residues in chondroitin 6-sulphate; hence the two names. Morquio B disease has clinical similarities to Morquio A but is due to deficiency of â-galactosidase. B.
Prenatal diagnosis of Morquio A disease is possible by assay of the enzyme in chorionic villi or cultured amniotic cells. Heterozygote detection is not offered.
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.
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