Product Manual

Taurine Assay Kit
Catalog Number
MET-5071

200 assays

FOR RESEARCH USE ONLY
Not for use in diagnostic procedures

Introduction
Taurine, also known as 2-aminoethanesulfonic acid, is a small molecule found in various types ofanimal tissues. Taurine is a large component of bile and represents almost 0.1% of total human bodyweight. In contrast to most biologically occurring acids that contain the weaker acidic carboxyl group,taurine is a sulfonic acid, and is not technically an amino acid due to it lacking the carboxyl group.
Taurine is biochemically synthesized from cysteine through a series of enzymatic reactions in thepancreas: the thiol group of cysteine is oxidized to cysteine sulfinic acid by cysteine dioxygenases,
cysteine sulfinic acid is decarboxylated by sulfinoalanine decarboxylase to form hypotaurine, andhypotaurine is oxidized to form taurine by hypotaurine dehydrogenase.
Taurine has many fundamental functions in mammalian biology. Taurine is conjugated tochenodeoxycholic acid and cholic acid forming the bile salts sodium taurochenodeoxycholate andsodium taurocholate. Taurine's sulfonic acid group has a low pKa which keeps this moiety negatively
charged in the intestinal tract and therefore improves the detergent properties of the bile conjugate.
Taurine is also important for adipose tissue and cardiovascular function as well as maturation andfunction of central nervous system, skeletal muscle, and the retina.
Cell Biolabs’ Taurine Assay Kit is a simple colorimetric assay that measures the total amount of freetaurine present in foods or biological samples in a 96-well microtiter plate format. Taurine found in
conjugated molecules (primary bile acids) is not detected. Each kit provides sufficient reagents toperform up to 200 assays*, including blanks, taurine standards, background controls and unknown
samples. Sample taurine concentrations are determined by comparison with a known taurine standard.
The kit has a detection sensitivity limit of 15.6 µM taurine.
*Note: Each sample replicate requires 2 assays, one treated with Stop Solution and one treated withQuenching Solution. Taurine content is calculated from the difference in OD readings from the 2 wells (SeeCalculation of Results section on page 7).

Assay Principle
Cell Biolabs’ Taurine Assay Kit measures Taurine within food or biological samples. Taurine isoxygenated by taurine dioxygenase in the presence of cofactor into 1-hydroxy-2-aminoethanesulfonicacid which decomposes to aminoacetaldehyde and sulfite. The enzymatic reaction is terminated by theaddition of EDTA (an inhibitor of taurine dioxygenase). The sulfite is then detected with a highly
specific colorimetric thiol probe. Samples and standards are read with a standard 96-well colorimetricplate reader. Samples are compared to a known concentration of taurine standard within the 96-wellmicrotiter plate format (Figure 1).

Figure 1. Taurine Assay Principle2

Related Products
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4.
5.

MET-5053: Total Thiol Assay Kit (Colorimetric)
MET-5054: L-Amino Acid Assay Kit (Colorimetric)
MET-5070: Glycine Assay Kit
STA-631: Total Bile Acid Assay Kit (Colorimetric)
STA-674: Glutamate Assay Kit

Kit Components
Box 1 (shipped at room temperature)
1. Taurine Standard (Part No. 50711C): One 100 µL tube at 100 mM.
2. 10X Assay Buffer (Part No. 50712A): One 10 mL bottle.
3. 10X Reagent A (Part No. 50713B): One 1 mL tube.
4. 10X Reagent B (Part No. 50714A): One 1 mL amber tube.
5. Cofactor (Part No. 50715A): One amber tube containing 250 mg.
6. Stop Solution (Part No. 50717A): One 5 mL bottle.
7. Quenching Solution (Part No. 50718A): One 5 mL bottle.
Box 2 (shipped on blue ice packs)
1. 10X Taurine Dioxygenase (Part No. 50716D): One 1 mL tube containing recombinant Taurine
Dioxygenase from Escherichia coli.
2. Developing Solution (Part No. 50719C): One 10 mL bottle.

Materials Not Supplied
1. Distilled or deionized water
2. 1X PBS
3. Standard 96-well microtiter plate and/or cell culture microplate

StorageUpon receipt, store the Taurine Standard, Taurine Dioxygenase, and Developing Solution at -20ºC.
Store the remaining components at 4ºC.

Preparation of Reagents••

1X Assay Buffer: Dilute the 10X Assay Buffer to 1X with deionized water. Stir to homogeneity.
Store at room temperature.
100X Cofactor: Weigh out at least 6 milligrams of solid Cofactor. Resuspend by vortexing
thoroughly in 1X Assay Buffer at 6 mg/mL. The 100X Cofactor will change color to bright orange
over the first several minutes of resuspension, but is stable for 8 hours at room temperature. Mix
100X Cofactor thoroughly just before use in reaction mix below. Discard unused 100X Cofactor
after 8 hours at room temperature.
3•Reaction Mix: Prepare a Reaction Mix by diluting the 10X Reagent A 1:10, 10X Reagent B 1:10,100X Cofactor 1:100, and Taurine Dioxygenase 1:10 in 1X Assay Buffer. For example, add 100
μL 10X Reagent A, 100 μL 10X Reagent B, 10 µL 100X Cofactor, and 100 µL of Taurine
Dioxygenase to 690 μL of 1X Assay Buffer for a total of 1 mL. This Reaction Mix volume isenough for 20 assays. The Reaction Mix is stable for 1 day at 4ºC.

Note: Prepare only enough for immediate use by scaling the above example proportionally.

Preparation of Samples••••

Tissue lysates: Sonicate or homogenize tissue sample in cold PBS or 1X Assay Buffer andcentrifuge at 10000 x g for 10 minutes at 4°C. Perform dilutions in 1X Assay Buffer.
Cell lysates: Resuspend cells at 1-2 x 106 cells/mL in PBS or 1X Assay Buffer. Homogenize orsonicate the cells on ice. Centrifuge to remove debris. Cell lysates may be assayed undiluted ordiluted as necessary in 1X Assay Buffer.
Urine: To remove insoluble particles, centrifuge at 10,000 rpm for 5 min. The supernatant may be
assayed undiluted or diluted as necessary in 1X Assay Buffer.
Serum or Plasma: Deproteinate the sample by running it through a centrifugal filter unit (Forexample use an Amicon Ultra 0.5 mL 10K Cat. No. UFC501024) and collecting the flow through.
To remove insoluble particles, centrifuge at 10,000 rpm for 5 min. The supernatant may be assayed
undiluted or diluted as necessary in 1X Assay Buffer.

Notes:All samples should be assayed immediately or stored at -80°C for up to 1-2 months. Run proper controls
as necessary. Optimal experimental conditions for samples must be determined by the investigator.
Always run a standard curve with samples.

Preparation of Standard Curve
Prepare fresh Taurine Standards before use by diluting in 1X Assay Buffer according to Table 2.
Standard Tubes
12345678100 mM Taurine Solution (µL)
5250 of Tube #1250 of Tube #2250 of Tube #3
250 of Tube #4
250 of Tube #5
250 of Tube #60

1X Assay Buffer (µL)
495
250
250
250250250250
250Taurine (µM)
1000500
250
125
62.5
31.3
15.60Table 2. Preparation of Taurine Standards.

Assay Protocol
1. Prepare and mix all reagents thoroughly before use. Each sample, including unknowns andstandards, should be assayed in duplicate or triplicate.
Note: Each sample replicate requires two paired wells, one to be treated with Stop Solution and one withQuenching Solution. While the Stop Solution contains EDTA (which inhibits Taurine Dioxygenase), theQuenching Solution contains both EDTA and a quenching reagent that specifically destroys any sulfite produced
by Taurine Dioxygenase, therefore serving as a background control.

42. Add 50 µL of each Taurine Standard or unknown sample into wells of a 96-well microtiter plate.
3. Add 50 µL of Reaction Mix to each well. Mix the well contents thoroughly and incubate for 30
minutes at room temperature.
4. Add 50 µL of Stop Solution to the Taurine Standards and one half of the paired sample wells.
5. Add 50 µL of Quenching Solution to the other half of the paired sample wells.
6. Add 50 µL of Developing Solution to all of the wells and mix thoroughly.
7. Incubate the plate on an orbital shaker for 3 minutes at room temperature.
8. Read the plate at 405-415 nm using a microplate spectrophotometer.

Example of Results
The following figures demonstrate typical Taurine Assay Kit results. One should use the data below forreference only. This data should not be used to interpret or calculate actual sample results.

Figure 2: Taurine Standard Curve.

Figure 3: Taurine Detection in Human Urine or Serum. Human samples were tested undiluted according to
the Assay Protocol (note: human serum was deproteinated according to the Preparation of Samples section).

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Calculation of Results
1.
2.
3.
4.

Determine the average optical density (OD) values for each sample, control, and standard.
Subtract the average zero standard value from itself and all standard values.
Graph the standard curve (see Figure 2).
Subtract the sample well values containing Quenching Solution from the sample well values containing
Stop Solution to obtain the difference. The OD difference is due to the enzyme Taurine Dioxygenase
activity (see Figure 3):

Net OD = ODStop – ODQuenching
5. Compare the Net OD of each sample to the standard curve to determine and extrapolate the quantity oftaurine present in the sample. Only use values within the range of the standard curve.

References
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Olive, MF (2002). Amino Acids. 23: 345–57.
Dominy, J Jr; Thinschmidt, JS; Peris, J; Dawson, R Jr; Papke, RL (2004). J. of Neurochem. 89:1195–205.
7. Foos, TM; Wu, JY (2002). Neurochem. Res. 27: 21–6.
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10. El Idrissi A, Messing J, Scalia J, Trenkner E; Messing; Scalia; Trenkner (2003). Adv. Exp. Med.
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Recent Product Citations
1. Justice, C.N. et al. (2023). Intra-ischemic hypothermia cardioprotection involves modulation ofPTEN/Akt/ERK signaling and fatty acid oxidation. Physiol Rep. 11(4):e15611. doi:
10.14814/phy2.15611.
2. Ying, Z. et al. (2022). Impact of Different Diets on Adult Tri-Spine Horseshoe Crab, Tachypleus
tridentatus. J Ocean Univ China. 21(3):541-548. doi: 10.1007/s11802-022-5199-4.
3. Yang, Z. et al. (2019). Serum Metabolomic Profiling Identifies Key Metabolic Signatures
Associated with Pathogenesis of Alcoholic Liver Disease in Humans. Hepatol Commun. 3(4):542557. doi: 10.1002/hep4.1322.

Warranty
These products are warranted to perform as described in their labeling and in Cell Biolabs literature when used inaccordance with their instructions. THERE ARE NO WARRANTIES THAT EXTEND BEYOND THIS EXPRESSEDWARRANTY AND CELL BIOLABS DISCLAIMS ANY IMPLIED WARRANTY OF MERCHANTABILITY ORWARRANTY OF FITNESS FOR PARTICULAR PURPOSE. CELL BIOLABS’s sole obligation and purchaser’s
exclusive remedy for breach of this warranty shall be, at the option of CELL BIOLABS, to repair or replace the products. Inno event shall CELL BIOLABS be liable for any proximate, incidental or consequential damages in connection with theproducts.

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