H-D-Phe-Homopro-Arg-pNA . 2 acetate (Synonyms: S-2238 acetate) |
Catalog No.GA22292
One-Click Copy Product Info
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H-D-Phe-Homopro-Arg-pNA . 2 acetate is a substrate of thrombin. After being cleaved by thrombin, this substrate releases p-nitroaniline (pNA), and its concentration can be determined by the absorbance at 405nm.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 115388-96-0
Sample solution is provided at 25 µL, 10mM.
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H-D-Phe-Homopro-Arg-pNA . 2 acetate is a substrate of thrombin. After being cleaved by thrombin, this substrate releases p-nitroaniline (pNA), and its concentration can be determined by the absorbance at 405nm.
H-D-Phe-Homopro-Arg-pNA is commonly used in thrombin activity assays, antithrombin/heparin cofactor activity measurements, plasma heparin content determination, and thrombin inhibitor screening [1-4].
References:
[1]. Lottenberg R, Hall JA, Fenton JW 2nd, Jackson CM. The action of thrombin on peptide p-nitroanilide substrates: hydrolysis of Tos-Gly-Pro-Arg-pNA and D-Phe-Pip-Arg-pNA by human alpha and gamma and bovine alpha and beta-thrombins. Thromb Res. 1982;28(3):313-332. doi:10.1016/0049-3848(82)90114-1.
[2]. Abildgaard U, Lie M. Antithrombin (heparin cofactor) assay with “new” chromogenic substrates (S-2238 and Chromozym TH). Thromb Res. 1977;11(4):549-553. doi:10.1016/0049-3848(77)90067-8.
[3]. Larsen ML, Abildgaard U, Teien AN, Gjesdal K. Assay of plasma heparin by amidolytic determination of thrombin with the chromogenic substrate H-D-Phe-Homopro-Arg-pNA (S-2238). Thromb Res. 1978;13(2):285-288. doi:10.1016/0049-3848(78)90071-1.
[4]. Wettstein L, Knaff PM, Kersten C, et al. Peptidomimetic inhibitors of TMPRSS2 block SARS-CoV-2 infection in cell culture. Commun Biol. 2022;5(1):681. doi:10.1038/s42003-022-03613-4.
The following protocol is recommended as a general guideline only and should be modified according to your specific requirements.
1.1 Preparation of Stock Solution
Dissolve one vial containing 10mg of H-D-Phe-Homopro-Arg-pNA . 2 acetate in 14.865mL sterile distilled water to prepare a 1.0mM stock solution.
Note: It is recommended to aliquot the H-D-Phe-Homopro-Arg-pNA stock solution and store it at 4°C protected from light.
1.2 Preparation of Working Solution
1) Transfer 7.5mL of H-D-Phe-Homopro-Arg-pNA stock solution.
2) Add 1.0mL of 3mg/mL Polybrene (GC19206) solution.
3) Add 1.5mL sterile distilled water.
4) Mix thoroughly to prepare a 0.75mM H-D-Phe-Homopro-Arg-pNA working solution.
Store at 4°C. The working solution is stable for 1 week.
1.3 Procedure
1) Add 50μL plasma sample to be tested into 3.50mL heparin-containing buffer for dilution.
2) Transfer 400μL of the diluted plasma and pre-incubate at 37°C for 2-6min.
3) Add 100μL thrombin solution (10 NIH U/mL) and mix thoroughly（if needed）.
4) After exactly 30s, add 300μL prewarmed H-D-Phe-Homopro-Arg-pNA working solution and mix vigorously.
5) After another exactly 30s, add 300μL 50% acetic acid immediately to terminate the reaction.
6) Keep the reaction mixture protected from light until absorbance measurement.
Measure absorbance at 405nm using a spectrophotometer.
7) Generate a standard curve and calculate the concentration of antithrombin-heparin cofactor (AT-III) in the plasma sample.
1.4 Blank Controls
1) Normal plasma blank:
400μL 1.00U/mL AT-III standard solution + 300μL 50% acetic acid + 400μL distilled water, mixed thoroughly.
2) Sample plasma blank:
400μL diluted plasma sample + 300μL 50% acetic acid + 400μL distilled water, mixed thoroughly.
References:
[1]. Friberger P, Knös M, Gustavsson S, Aurell L, Claeson G. Measurement of antithrombin III in normal and pathologic states using chromogenic substrate S-2238. Comparison with immunoelectrophoretic and factor Xa inhibition assays. Thromb Res. 1977;11(6):755-763. doi:10.1016/0049-3848(77)90086-1.
| Cas No. | 115388-96-0 | SDF | |
| Synonyms | S-2238 acetate | ||
| Formula | C27H36N8O5 · 2 C2H4O2 | M.Wt | 672.74 |
| Solubility | H2O : 125 mg/mL (204.02 mM; Need ultrasonic); DMSO : 100 mg/mL (163.22 mM; Need ultrasonic) | Storage | Store at -20°C |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 1.4865 mL | 7.4323 mL | 14.8646 mL |
| 5 mM | 297.3 μL | 1.4865 mL | 2.9729 mL |
| 10 mM | 148.6 μL | 743.2 μL | 1.4865 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
g
μL
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >98.00% Appearance: A solid
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 3 reference(s) in Google Scholar.)