Recombinant Human Trypsin 2/PRSS2 Protein, CF

Catalog # Availability Size / Price Qty
3586-SE-010
Product Details
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Recombinant Human Trypsin 2/PRSS2 Protein, CF Summary

Product Specifications

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave the fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002). The specific activity is >8,000 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Trypsin 2/PRSS2 protein
Ala16-Ser247, with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Ala16
Structure / Form
Pro form
Predicted Molecular Mass
26 kDa
SDS-PAGE
34 kDa, reducing conditions

Product Datasheets

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

3586-SE

Formulation Supplied as a 0.2 μm filtered solution in HCl and NaCl.
Shipping The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Activation Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
  • Assay Buffer: 0.1 M Tris, 0.15 M NaCl, 10 mM CaCl2, 0.05% Brij-35 (w/v), pH 8.0
  • Recombinant Human Enteropeptidase/Enterokinase (rhEnterokinase) (Catalog # 1585-SE)
  • Recombinant Human Trypsin 2/PRSS2 (rhTrypsin 2) (Catalog # 3586-SE)
  • Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
  • 1,10 Phenanthroline (Sigma, Catalog # 320056), 0.6 M stock in DMSO
  • Substrate II: MCA-Arg-Pro-Lys-Pro-Val-Glu-Nva-Trp-Arg-Lys(DNP)-NH2 (Catalog # ES002), 2 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
Activate rhEnterokinase with Thermolysin.
  1. Dilute rhEnterokinase to 100 µg/mL in Activation Buffer.
  2. Dilute Thermolysin to 3.16 µg/mL in Activation Buffer.
  3. Mix 50 µL of 100 µg/mL rhEnterokinase with 50 µL of 3.16 µg/mL Thermolysin.
  4. Incubate reaction at 37 °C for 30 minutes.
  5. Stop reaction by adding 100 µL of 20 mM 1, 10 Phenanthroline to the reaction.
Activate rhTrypsin 2 with activated rhEnterokinase.
  1. Dilute activated rhEnterokinase to 0.4 µg/mL in Assay Buffer.
  2. Dilute rhTrypsin 2 to 200 µg/mL in Assay Buffer.
  3. Mix 25 µL of 0.4 µg/mL rhEnterokinase with 25 µL of 200 µg/mL rhTrypsin 2.
  4. Incubate reaction at room temperature for 30 minutes.
Perform reaction.
  1. Dilute activated rhTrypsin 2 to 0.05 µg/mL in Assay Buffer.
  2. Dilute substrate to 20 µM in Assay Buffer.
  3. In a plate load 50 µL of 0.05 μg/mL of rhTrypsin 2 and include a Substrate Blank of 50 µL Assay Buffer.
  4. Start the reaction by adding 50 µL of 20 µM Substrate to wells.
  5. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
  6. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU)
amount of enzyme (µg)

     *Adjusted for Substrate Blank

     **Derived using calibration standard MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).

Per Well:
  • rhTrypsin 2: 0.0025
  • Substrate: 10 μM
Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: Trypsin 2/PRSS2

Human Trypsin 2, encoded by the PRSS2 gene, is also known as anionic Trypsin (1). Accounting for approximately 1/3 of the total trypsin content in the pancreatic juice, it is one of the trypsins produced in the pancreas. It consists of a signal peptide (residues 1‑15), a pro region (residues 16‑23), and a mature chain (residues 24‑247). Trypsin 2 is synthesized as a zymogen and secreted into the duodenal lumen, where it is activated by enteropeptidase (2). It plays a central role in digestion and activating other pro-enzymes. In human pancreatic diseases and chronic alcoholism, it is up-regulated (3).

References
  1. Emi, M. et al. (1986) Gene. 41:305.
  2. Halfon, S. et al. (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, et al.) pp. 1483-1488, Academic Press, San Diego.
  3. Rinderknecht, H. et al. (1979) Gut. 20:886.
Entrez Gene IDs
5645 (Human); 22072 (Mouse); 25052 (Rat)
Alternate Names
Anionic trypsinogen; EC 3.4.21; EC 3.4.21.4; MGC111183; MGC120174; MGC120175; protease serine 2 preproprotein; protease, serine, 2 (trypsin 2); protease, serine, 2, preproprotein; PRSS2; Serine protease 2; Tesp4; TRY2TRY8; TRY8; TRYP2; Trypsin 2; Trypsin II; trypsin-2; trypsinogen 2; Trypsinogen 20

FAQs

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Recombinant Human Trypsin 2/PRSS2 Protein, CF
By Anonymous on 12/05/2018
Application: Immunoassay Standard
Reason for Rating: We have used this IRT-2 to prepare dried blood spot Standards and Controls. These DBS material is used in our multiplex assay (Luminex xMAP Platform) to screen newborn babies for Cystic Fibrosis.

We started with only IRT-2 and then added a combination of IRT-1 + IRT-2. Both with excellent results.