Assays / Genetic Enzymes / Glycosaminoglycans

Principle of Method:
Quantitive and qualitative methods are used for urinary glycosaminoglycans (GAGs). In the quantitative method GAGs are determined in urine by reaction with the dye dimethylmethylene blue (DMB) in a reaction that does not require prior precipitation of the GAGs. The colour is measured rapidly after mixing using a Cobas Bio centrifugal analyser. The creatinine content of the urine is measured and results are expressed in terms of this. In the qualitative method
GAGs are precipitated from urine using the cationic detergent cetylpyridinium chloride (CPC). After collection and re-precipitation they are subjected to two-dimensional electrophoresis on cellulose acetate and the separated components visualised with Alcian Blue.

Uses and Limitations of Methods:
The quantitative method is used for screening all the mucopolysaccharides and probably gives rise to fewer false negatives or false positives than other screening procedures. The main limitation is that Morquio disease (MPS IV) may be missed, particularly in older patients. The qualitative method is used to follow up positive results obtained with the quantitative assay. It is also used irrespective of quantitative results in the following situations:

• A positive urinary MPS result has been obtained elsewhere.
• There is a family history of a mucopolysaccharidosis.
• There is a clinical suspicion of Morquio disease (MPS IV).
• There is a particularly strong suspicion of an MPS and the case has been discussed with the SAS laboratory. In practice, most urines sent to the lab are assayed using both methods.

From the pattern of GAGs seen in the qualitative method it is usually possible to make a preliminary diagnosis within the following categories:

• Hurler disease, Hunter disease or Scheie disease
• Sanfilippo disease
• Morquio disease
• Maroteaux-Lamy disease
• Sly disease (MPS VII) is unlikely to be recognised as a specific pattern because of lack of experience in the laboratory with this MPS.
  

Patients must always be followed up by enzymatic assays for unequivocal assignment of type.

Two-dimensional electrophoresis of GAGs is also useful for prenatal diagnosis of MPSs using cell-free amniotic fluid obtained in the second trimester (15 weeks' gestation or later).

Specimen Requirements:
Urine 10-20 ml in a sterile container without preservative. Freeze if not sending immediately but can be transported to the laboratory at room temperature.

THE LABORATORY RECOMMENDS USE OF A COURIER SERVICE OR ROYAL MAIL SPECIAL DELIVERY FOR SENDING ALL SPECIMENS TO THE LABORATORY.

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