Principle Of Method: Skin fibroblasts are treated with a concentration of digitonin which makes the cytoplasmic membrane leaky, but which does not affect the permeability of the peroxisomal membrane. Cells are separated into a supernatant fraction containing cytoplasmic enzymes and a pellet fraction consisting of organelle-associated enzymes for the spectrophotometric determination of catalase activity.
Uses And Limitations Of The Method: This method is used principally for the further characterisation of peroxisomal patients and not as a preliminary screening method. It can be used to categorise patients with biochemical evidence of a peroxisomal disorder into two groups: (1) peroxisomal disorders affecting peroxisome biogenesis or integrity and (2) peroxisomal disorders caused by enzyme defects which do not affect the integrity of the organelle. Disorders which affect peroxisome biogenesis include Zellweger syndrome, neonatal adrenoleucodystrophy and infantile Refsum disease. Patients with these disorders are generally diagnosed by the measurement of very long chain fatty acids levels or plasmalogen biosynthesis activity. Most patients with one of these disorders can be detected using this method. However, it is possible for mature peroxisomes to be present in skin fibroblasts, but absent in a critical organ such as the liver. Thus, the occasional mosaic patient would not necessarily be detected.
Specimen Requirements: Fibroblasts cultured from a skin biopsy are 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 at the laboratory 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.