Table 1

Assay conditions for each enzyme activity assay.

ENZYME

Punch diameter

Reaction buffer

Fluorogenic substrate

Additional reagents

Incubation time

Stop buffer


alpha-galactosidase A

1.5 mm

30 μL citrate-phosphate 0.3 M pH 5.0

30 + 30* μL

4 MU-alpha-D-galactopyranoside 4.2 mM

10 μL

N-acetyl-galactosamine 0.25 M

20 hours

10%

ethylenediamine 1.32 M pH 11.3


alpha-iduronidase

1.5 mm

10 μL

sodium formate 0.2 M pH 2.8

20 + 20* μL

4MU-alpha-L-idopyranoside

2 mM

10 μL

D-saccharic acid

1,4-lactone monohydrate 3 mM

20 hours

10%

ethylenediamine 1.32 M pH 11.3


beta-glucosidase

3.0 mm

30 μL citrate-phosphate 0.4 M pH 5.2

50 + 50* μL

4MU-beta-D-glycopyranoside 20 mM

40 μL

sodium taurodeoxicolate hydrate 0.75%

20 hours

10%

ethylenediamine 1.32 M pH 11.3


alpha-glucosidase

1.5 mm

20 μL

sodium acetate 0.4 M pH 4.0 and 6.5

20 + 20* μL

4MU-alpha-D-glycopyranoside 2.8 mM

40 μL

acarbose 40 μM

24 hours

10%

ethylenediamine 1.32 M pH 11.3


beta-galactosidase

1.5 mm

20 μL citrate-phosphate 0.1 M pH 4.4

20 + 40* μL

4MU-beta-D-galactopyranoside 0.8 mM

20 μL sodium chloride 0.9%

3 hours

10%

ethylenediamine 1.32 M pH 11.3


* addition in blank wells after incubation

MU: methylumbilliferyl

Müller et al. Diagnostic Pathology 2010 5:65   doi:10.1186/1746-1596-5-65

Open Data