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Human Serpin G1/C1 Inhibitor (Plasma) Protein, CF

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Summary
Reactivity HuSpecies Glossary
Applications Inhibition Activity
Format
Carrier-Free

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Human Serpin G1/C1 Inhibitor (Plasma) Protein, CF Summary

Details of Functionality
Measured by its ability to inhibit Recombinant Human Complement Component C1s (Catalog # 2060-SE) cleavage of a colorimetric peptide substrate, N-carbobenzyloxy-Lys-ThioBenzyl ester (Z-K-SBzl). The IC50 is <2.6 nM, as measured under the described conditions.
Source
Human plasma-derived Serpin G1/C1 Inhibitor protein
The human plasma used for the isolation of this product were certified by the supplier to be HIV-1 and HBsAg negative at the time of shipment. Human blood products should always be treated in accordance with universal handling precautions.
N-terminal Sequence
NPNATSSS
Protein/Peptide Type
Natural Enzymes
Gene
SERPING1
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Dilutions
  • Inhibition Activity
SDS-PAGE
93 kDa, reducing conditions
Publications
Read Publications using
2488-PI in the following applications:

Packaging, Storage & Formulations

Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after reconstitution.
Buffer
Lyophilized from a 0.2 μm filtered solution in Sodium Acetate and NaCl.
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Reconstitution Instructions
Reconstitute at 0.5 mg/mL in sterile, deionized water.
Assay Procedure
  • Assay Buffer: 50 mM Tris, 10 mM CaCl2, 0.15 M NaCl, and 0.05% Brij-35 (w/v), pH 7.5 (TCNB)
  • Human Serpin G1/C1 Inhibitor (hSerpin G1) (Catalog # 2488-PI)
  • Recombinant Human Complement Component C1s (rhC1s) (Catalog # 2060-SE)
  • Substrate Z-Lys-SBzl (Bachem, Catalog # M-1300), 10 mM in DMSO
  • 5,5’-Dithobis(2-Nitrobenzoic acid) (DTNB) (Sigma, Catalog # D-8130), 10 mM in DMSO
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Fluorescent Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhC1s to 2 µg/mL in Assay Buffer.
  2. Prepare a curve of hSerpin G1 (MW: 93,000 Da) in Assay Buffer. Make the following serial dilutions: 2000, 1000, 500, 100, 50, 25, 12.5, 4, 0.8 and 0.2 nM.
  3. Combine 25 µL of diluted rhC1s and 25 µL of hSerpin G1 at each concentration of the curve. Include two rhC1s blanks containing Assay Buffer in place of hSerpin G1.
  4. Incubate mixtures at room temperature for 30 minutes.
  5. Dilute the mixtures 5-fold by combining 50 µL of reaction mixture with 200 µL Assay Buffer.
  6. Dilute substrate to 200 μM with Assay Buffer with 200 μM DTNB.
  7. Load into plate 50 µL of the diluted incubated mixtures, and start the reaction by adding 50 µL of 200 µM substrate/DTNB mixture.
  8. Read at an absorbance of 405 nm in kinetic mode for 5 minutes.
  9. Derive the 50% inhibition concentration (IC50) for hSerpin G1 by plotting OD/min (or specific activity) vs. concentration with 4-PL fitting.
  10. The specific activity for rhC1s at each point may be determined using the following formula (if needed):

     Specific Activity (pmol/min/µg) =

 Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/M
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank
     **Using the extinction coefficient 13260 M-1cm-1
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rhC1s: 0.010 µg (1.33 nM)
  • hSerpin G1 curve: 100, 50, 25, 5, 2.5, 1.25, 0.625, 0.2, 0.04 and 0.01 nM
  • Substrate: 100 µM
  • DTNB: 100 µM
      		•

      Notes

      This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

      Alternate Names for Human Serpin G1/C1 Inhibitor (Plasma) Protein, CF

      • C1 Inhibitor
      • C1IN
      • C1INH
      • C1NH
      • HAE1
      • HAE2
      • Serpin G1
      • SERPING1 serpin peptidase inhibitor, clade G (C1 inhibitor), member 1

      Background

      As a member of the serpin superfamily of serine protease inhibitors, Serpin G1/C1 inhibitor is the physiological inhibitor of activated C1r and C1s, two serine proteases involved in the classical complement pathway. In addition, it inhibits plasma kallikrein and coagulation factor XIIa, two serine proteases involved in the processing of kininogen to release bradykinin. Therefore, it plays an important role in regulating activation of both the complement and contact systems (1). Serpin G1 deficiency results in hereditary angioedema, which is characterized by recurrent episodes of localized angioedema of the skin, gastrointestinal mucosa or upper respiratory mucosa (2). The deduced amino acid sequence of human Serpin G1 precursor consists of 500 residues with a signal peptide. The mature protein of 478 amino acid residues is heavily glycosylated (1).

      1. Davis, A.E. III et al. (1993) Methods Enzymol. 223:97.
      2. Davis, A.E. III (2004) Drug News Perspect. 17:439.

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      Publications for Serpin G1/C1 Inhibitor (2488-PI)(4)

      We have publications tested in 1 confirmed species: Human.

      We have publications tested in 4 applications: Bioassay, ELISA Developmet, ELISA Standard, Enzyme Assay.


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      Bioinformatics

      Gene Symbol SERPING1