Recombinant Human Serpin E2 (carrier-free)

Product Details

Source

Human Serpin E2, amino acids Ser20-Pro398 (Accession # NM_001136528) with a C-terminal TG-8H-GGQ tag was expressed in 293E cells.

Molecular Mass

The 392 amino acid recombinant protein has a predicted molecular mass of approximately 43.4 kD. The non-reduced and DTT-reduced proteins migrate at 50 - 60 kD by SDS-PAGE.

Purity

> 95 % as determined by Coomasie stained SDS-PAGE

Formulation

0.22 µm filtered protein solution is in PBS, pH 7.2.

Endotoxin Level

Less than 1.0 EU per µg of protein as determined by the LAL method.

Concentration

10 and 25 µg sizes are bottled at 200 µg/mL. 100 µg size and larger sizes are lot-specific and bottled at the concentration indicated on the vial. To obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.

Storage & Handling

Unopened vial can be stored between 2°C and 8°C for up to 2 weeks, at -20°C for up to six months, or at -70°C or colder until the expiration date. For maximum results, quick spin vial prior to opening. The protein can be aliquoted and stored at -20°C or colder. Stock solutions can also be prepared at 50 - 100 µg/mL in appropriate sterile buffer, carrier protein such as 0.2 - 1% BSA or HSA can be added when preparing the stock solution. Aliquots can be stored between 2°C and 8°C for up to one week and stored at -20°C or colder for up to 3 months. Avoid repeated freeze/thaw cycles.

Activity

Recombinant human Serpin E2 inhibits the activity of trypsin from bovine pancreas (at 10 ng/ml). The IC₅₀ value is less than 1.84 nM (80 ng/mL).

Application

Bioassay

Application Notes

Human  Serpin E2 Activity Assay

Human Serpin E2 activity is measured by its ability to inhibit cleavage of a fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH₂,  by Trypsin from Bovine pancreas. The potency of hSerpin E2 is described as an IC₅₀ value.

Materials

1. Recombinant human Serpin E2 (Predicted MW: 43.4 kDa)
2. Assay Buffer: pH 7.5, 50 mM Tris, 10 mM CaCl₂ 0.15 M NaCl, 0.05% Brij-35.
3. Bovine Trypsin
4. Trypsin substrate: Mca-RPKPVE-Nval-WRK(Dnp)-NH₂

 

Activity assay procedures

1. Dilute hSerpin E2 in 16.0 µg/mL in Assay Buffer.
2. Make serial dilution of hSerpin E2 in Assay Buffer from 16.0 µg/mL to 0.0156 µg/mL, and include 0 ng/mL as control.
3. Dilute Trypsin in 0.2 ug/mL in Assay Buffer.
4. Add into a tube 20 µL of each serially-diluted hSerpin E2 including the control without hSerpin E2, and then add 20 µL of 0.2 µg/mL trypsin solution. Briefly vortex, and then centrifuge the tubes to mix the solutions.
5. Incubate the mixture at RT for 30 min.
6. Add 160 µL of Assay Buffer into each tube prepared in step 4 after incubation to make diluted assay mixtures.
7. Dilute the substrate in Assay Buffer at 20 µM.
8. Load into a well plate 50 µL of each assay mixture and start the reaction by adding 50 µL of 20 µM Substrate. Include a substrate blank containing 50 µL Assay Buffer and 50 µL of 20 µM Substrate.
9. Read at excitation and emission wavelengths of 320 nm and 405 nm, respectively, in kinetic mode for 5 minutes.
10. Analyze the data with non-linear regression to estimate IC₅₀ value of hSerpin E2 towards bovine trypsin.

 

Final assay conditions per well

• Trypsin: 1 ng (10 ng/mL)
• hSerpin E2: 800, 400, 200, 100, 50, 25, 12.5, 6.25, 3.13, 1.56, 0.781, 0 ng/mL
• Substrate: 10 µM

BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue-ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are verified in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at tech@biolegend.com.

Antigen Details

Structure

Monomer

Distribution

Tissue: Brain, Kidney, Ovary

Cells: Monocyte, macrophage, fibroblast, myoblasts, vascular smooth muscle cells

Function

Blood coagulation, cell migration, neurite outgrowth.

Interaction

Thrombin, uPA, tPA, Plasmin

Ligand/Receptor

Glycosaimnoglycan, Heparan sulfate

Bioactivity

Inhibit thrombin, uPA, tPA, and plasmin.

Biology Area

Neuroscience

Molecular Family

Enzymes and Regulators

Antigen References

1. Bergeron S, et al. 2010. Mol. Cancer 9:271.
2. Fayard B et al. 2009. Cancer Res. 69:5690.
3. Law RH, et al. 2006. Genome Biology 7:216.
4. Crisp RJ, et al. 2000. J. Biol. Chem. 275:19628.
5. Baker JB, et al. 1980. Cell. Aug;21(1):37-45.

Gene ID

5270 View all products for this Gene ID

UniProt

View information about Serpin E2 on UniProt.org

Documentation

• Technical Data Sheet (pdf)
• Certificate of Analysis
• Material Safety Data Sheet

Reviews

Related Pages & Pathways

Related FAQs

Why choose BioLegend recombinant proteins?

     • Each lot of product is quality-tested for bioactivity as indicated on the data sheet.
     • Greater than 95% Purity or higher, tested on every lot of product.
     • 100% Satisfaction Guarantee for quality performance, stability, and consistency.
     • Ready-to-use liquid format saves time and reduces challenges associated with reconstitution.
     • Bulk and customization available. Contact us.
     • Learn more about our Recombinant Proteins.

How does the activity of your recombinant proteins compare to competitors?

We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!

What is the specific activity or ED50 of my recombinant protein?

The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.

Have your recombinants been tested for stability?

Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.

Does specific activity of a recombinant protein vary between lots?

Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.

How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?

Use formula Specific activity (Units/mg) = 10^6/ ED50 (ng/mL)

Certificate of Analysis