TBS2105 β Hexosaminidase Activity Assay 2022

Beta-Hexosaminidase Activity Colorimetric Assay
Catalog Number Kit Size

TBS2105 100 assays
Description cells in an appropriate volume of cold PBS, approximately one
β-Hexosaminidase are hydrolytic enzyme responsible for the million cells per mL. Centrifuge at 14,000 x g for 10 min at 4°C.
breakdown of carbohydrates. Specifically, β-Hexosaminidase Remove supernatant for assay.
cleaves the terminal β-D-glucuronic acid residue from the
nonreducing terminus of a mucopolysaccharide chain. In humans, Reagent Preparation: Equilibrate all components to 37°C.
these enzymes are found in the lysosome of many tissue types. Briefly vortex or pipette up and down all components to ensure
Loss of β-Hexosaminidase activity results in metabolic disease fresh reconstitution.
and health problems. It is important to detect β-Hexosaminidase Reaction Preparation:
activity in the tested samples for disease examination.
1. Label tubes as #1 through #8 as below diagram.
The Beta-Hexosaminidase Activity Colorimetric Assay provides 2. Add 400 µL of 1x Assay Buffer to Std1, and 200 µL to Std2 to
a simple and sensitive method for monitoring hexosaminidase 8.
activity in biological samples (tissue, cells, serum, urine). This 3. Pipet 100 µL of 10 mM standard stock into Std#1. Then, then
assay uses a synthetic p- nitrophenol derivative (R-pNP) as its make 2x series dilution in Std2 through 7 with addition of 200 µL.
substrate and releases pNP which can be measured at absorbance Std8 is 1x Assay Buffer alone as a standard 0. The standard
(OD 405 nm). The assay can detect as low as 50 µU of NAGase concentration in tube 1 through 7 will be 2000,1000, 500, 250,
activity in a variety of samples. 125, 62.5 and 31.25µM, Tube#8 is Standard 0 as blank.
Applications
100µl 200µl 200µl 200µl 200µl 200µl 200µl
This kit is used for determination of β-Hexosaminidase activity in
biological samples.
Key features
Fast and sensitive: Linear detection range (20 µL sample): 0.05
to 50 U/L for a 30 minutes reaction at 37˚C
10mM
High throughput: Can be readily automated on HTS liquid
handling systems for processing thousands of samples per day.
Kit Contents Std1 Std2 Std3 Std4 Std5 Std6 Std7 Std8
Component 100x RXNS 200x RXNS Assay
Buffer (µL) 400 200 200 200 200 200 200 200
Substrate 10 mL 20 mL
Addition Stock Std1 Std2 Std3 Std4 Std5 Std6
Standard (10mM) 1 mL 2 mL Addition
Hexosaminidase positive 50 µL 100 µL Vol. (µL) 100 200 200 200 200 200 200 0
control Final Conc
Stop Reagent 12ml 24mL (µM) 2000 1000 500 250 125 62.5 31.25 0
Assay Buffer 12ml 24mL 4. Transfer 20 µL of each sample, blank, positive control, and
standards into two separate wells.
STORAGE CONDITIONS 5. Add 80 µL of the substrate solution to all sample, positive
The kit is shipped on ice and should be stored at -20°C for long- control, and blank wells. Add 80 µL of Assay Buffer to each
term storage. Shelf life of 12 months after receipt. standard well (Note: Do not add substrate in the standard). Tap
plate briefly to mix.
PROCEDURES
This assay is based on a kinetic reaction. To ensure identical 3. Incubate at 37°C or desired temperature for 30-60 minutes.
incubation time, addition of Substrate and Stop Reagent to 4. Add 100 µL of Stop Reagent to all wells. Tap plate briefly to
samples should be quick, and mixing should be brief but thorough. mix.
Use of a multi-channel pipettor is recommended. 5. Read OD405nm.
Sample Preparation: Serum and plasma can be assayed CALCULATION

directly. For urine samples containing precipitation, centrifuge at Subtract blank OD (Standard 0, #8) from the standard OD values
10,000 x g, 4°C for 3 minutes and assay the supernatant. and plot the ΔOD against standard concentrations. Determine the
slope, and use the following equation to calculate β-
Cell Lysate: Collect cells by centrifugation at 2,000 x g for 5 min Hexosaminidase activity:
at 4°C. For adherent cells, do not harvest cells using proteolytic
enzymes; rather use a rubber policeman. Homogenize or sonicate
2011 © by Tribioscience 365 San Aleso Ave, Sunnyvale, CA 94085 USA Website: www.tribioscience.com
Tel: 408-498-0197 / 650-917-9269, Fax: 650-618-5498 Email: order@tribioscience.com
Page 1 of 2
Beta-Hexosaminidase Activity Colorimetric Assay
Β-Hexosaminidase Activity (U/L): RELATED PRODUCTS:

DF * (ODSAMPLE – OD BLANK)/ (t * Slope) Tryptase activity colorimetric assay (TBS2101)
where ODSAMPLE is the OD405nm value for each sample and β-Glucuronidase Assay (TBS2110)
ODBLANK is the OD405nm value of the sample blank. Slope is the Caspase-3 Fluorometric Assay kit (TBS3230)
slope of the linear regression fit of the standard points and t is the Cytochrome C Oxidase Activity Assay (TBS2115)
reaction time (30 min). DF is the dilution factor. Fast Glucose Determination Colorimetric/Fluorometric Assay
(TBS2087)
Unit definition: 1 Unit (U) will catalyze the conversion of 1 µmole Glucose Oxidase Activity Colorimetric/Fluorometric Assay
of pNitrophenyl N-acetyl-β-D-glucosaminide to p-Nitrophenol (TBS2088)
and β-Hexosaminidase per min at 37°C Non-esterified Fatty Acid Assy (TBS2203)
Glycerol Colorimetric / Fluorometric Assy (TBS2204)
TYPICAL DATA Protein Assay Kits (TBS2005)
This standard curve is provided for demonstration only as below Cell Nuclear Extract kit (TBS6025)
figure. A standard curve should be generated for each set of
samples assayed.
Research use only.

pNP standard curve
2
R² = 1
1.5
OD at 405nm
0.5
0
0 500 1000 1500 2000
pNP standard concentration(µM)
2011 © by Tribioscience 365 San Aleso Ave, Sunnyvale, CA 94085 USA Website: www.tribioscience.com
Tel: 408-498-0197 / 650-917-9269, Fax: 650-618-5498 Email: order@tribioscience.com
Page 2 of 2

TBS2105 β Hexosaminidase Activity Assay 2022

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TBS2105 β Hexosaminidase Activity Assay 2022

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Beta-Hexosaminidase Activity Colorimetric Assay

Catalog Number Kit Size

Sample Preparation: Serum and plasma can be assayed CALCULATION

Β-Hexosaminidase Activity (U/L): RELATED PRODUCTS:

Research use only.

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