Iduronate-2-sulfatase, Whole Blood
Method Description Describes how the test is performed and provides a method-specific reference
Whole blood is spotted on filter paper. A 3-mm (one-eighth-inch) disk is punched out of the dried blood spot (DBS) into a microcentrifuge tube and 100 mcL of 0.2%, heat/pH inactivated bovine serum albumin is added as a preincubation extraction which takes place on an orbital shaker for 30 minutes at 37 degrees C. Ten mcL of the extraction liquid is combined with 20 mcL of 1.25 mM 4-methylumbelliferyl-alpha-Ido-A-2S in acetate buffer as the substrate in a black 96-well plate (30 mcL total volume plus DBS). A blank is prepared using only preincubation extraction liquid, substrate, and filter paper punch containing no blood (30 mcL total volume plus blank punches). All patients, controls, and blanks are set up in duplicate (extraction liquid from each of the microcentrifuge tubes). After the incubation period (24 hours at 37 degrees C), 40 mcL of 0.2 M sodium phosphate/0.1 M citric acid, pH 4.5 and 10 mcL of LEBT solution (lysosomal enzymes purified from bovine testis) are added to the same plate. After a second incubation (24 hours at 37 degrees C), 200 mcL of stop buffer (0.5 M NaHCO/0.5 M NaCO plus 0.025% Triton X-100, pH 10.7) is added to all wells (patients, quality control, blanks, calibrators). Calibrators are prepared and analyzed on every plate to calculate enzyme activity results based on fluorescence units in patient wells versus calibrators. The calibration is derived from 4-methylumbelliferone that is serially diluted manually in the plate, with the highest calibrator being equivalent to an enzyme activity of 3.125 nmol/mL blood/hr. The plate is then ready to be read using the spectrofluorometer. Fluorescence readings for duplicate wells are averaged, and the average fluorescence is used to calculate the enzyme activity result.(Civallero G, Michelin K, de Mari J, et al: Twelve different enzyme assays on dried-blood filter paper samples for detection of patients with selected inherited lysosomal storage diseases. Clin Chim Acta 2006;372:98-102)
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Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Analytic Time Defines the amount of time it takes the laboratory to setup and perform the test. This is defined in number of days. The shortest interval of time expressed is "same day/1 day," which means the results may be available the same day that the sample is received in the testing laboratory. One day means results are available 1 day after the sample is received in the laboratory.
Maximum Laboratory Time Defines the maximum time from specimen receipt at Mayo Medical Laboratories until the release of the test result
Specimen Retention Time Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded
Performing Laboratory Location The location of the laboratory that performs the test