Suc-Ala-Phe-Lys-AMC |
Catalog No.GA23548
One-Click Copy Product Info
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Suc-Ala-Phe-Lys-AMC is a highly sensitive fluorogenic substrate.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 73207-91-7
Sample solution is provided at 25 µL, 10mM.
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Suc-Ala-Phe-Lys-AMC is a highly sensitive fluorogenic substrate. Suc-Ala-Phe-Lys-AMC can be specifically recognized and cleaved by plasmin, releasing the AMC fluorophore. Enzyme activity is quantified by measuring the change in fluorescence intensity. Suc-Ala-Phe-Lys-AMC can be used for the detection and related research of plasmin and gingipain K[1-2].
References:
[1] Götz MG, James KE, Hansell E, et al. A new class of potent and selective inhibitors of asparaginyl endopeptidases (legumains) from evolutionarily diverse pathogens. J Med Chem. 2008 May 8;51(9):2816-32.
Determination of Plasmin Activity Using Suc-Ala-Phe-Lys-AMC[1]
This protocol is adapted from research data and provided for reference only. Adjustments may be necessary based on specific experimental requirements.
1. Reagent Preparation: Dissolve Suc-Ala-Phe-Lys-AMC in a buffer to prepare a working concentration substrate solution (e.g., 0.045mM Ala-Phe-Lys-AMC or 0.8mM Suc-Ala-Phe-Lys-AMC). Use 0.05M Tes/NaOH buffer (pH=7.5). Prepare a 20% (v/v) dimethyl sulfoxide solution as an auxiliary solvent; this concentration has no effect on enzyme activity. Prepare a series of plasmin standard solutions with known activity (CTA units).
2. Sample Pretreatment: For crude extract samples, to avoid non-specific hydrolysis of the substrate by aminopeptidases, a protected substrate such as Suc-Ala-Phe-Lys-AMC must be used. Cell culture supernatant samples can be used directly after removing cell debris by centrifugation. When testing samples that may contain plasminogen activators (e.g., urokinase) or thrombin, the inhibitor Trasylol should be prepared.
3. Plasmin Activity Assay: Under conditions of 24°C, add the substrate solution, Tes buffer, and 20% DMSO to a final reaction volume of 0.5mL. Then, initiate the reaction by adding the enzyme solution (e.g., plasmin, urokinase, or thrombin) or the test sample. The reaction time is typically 5 minutes or can be adjusted based on enzyme activity.
4. Fluorescence Measurement and Quantification: Plasmin hydrolyzes Suc-Ala-Phe-Lys-AMC, releasing the fluorescent group 7-amino-4-methylcoumarin. After the reaction, measure the fluorescence intensity under standardized conditions (calibrate the fluorometer daily with 10nM 7-amino-4-methylcoumarin) using a spectrofluorometer. Calculate the enzyme activity by comparing the rate of fluorescence intensity change against a standard curve.
Precautions:
(1) This method is extremely sensitive, capable of detecting as little as 10⁻⁵CTA units of plasmin. The reaction rate is linear over at least a 100-fold range of enzyme concentrations. By utilizing the inhibitor Trasylol and differences in the kinetic constants of the substrates, plasmin activity can be specifically assayed even in the presence of enzymes like thrombin or urokinase.
(2) For your safety and health, please wear a lab coat and disposable gloves during operation.
References:
[1] Pierzchala PA, Dorn CP, Zimmerman M. A new fluorogenic substrate for plasmin. Biochem J. 1979 Dec 1;183(3):555-9. doi: 10.1042/bj1830555.
| Cas No. | 73207-91-7 | SDF | |
| Formula | C32H39N5O8 | M.Wt | 621.69 |
| Solubility | Soluble in DMSO | 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 | 1.6085 mL | 8.0426 mL | 16.0852 mL |
| 5 mM | 321.7 μL | 1.6085 mL | 3.217 mL |
| 10 mM | 160.9 μL | 804.3 μL | 1.6085 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 34 reference(s) in Google Scholar.)