murine MAb against Gla (matrix bound) residues
Description
REF ADG3570Mx is a monoclonal antibody (REF 3570) directed against gamma-carboxyglutamyl (Gla) residues found in various proteins of human and other origin, and snake venoms, bound to SEPHAROSE™ CL-4B. It is intended for use in the affinity purification of Gla-containing proteins such as prothrombin, factor VII/VIIa, factor IX/IXa, factor X/Xa, Protein C, Protein S and growth arrest-specific protein-6 (Gas6).
Properties
The antibody is bound to the sepharose at a concentration of 5 mg of antibody/mL of settled gel.
Purification Protocol
Using the following buffers, a suggested purification protocol is:
Loading/Washing Buffer: 20 mM Tris-HCl, pH 7.4
150 mM NaCl, 10 mM EDTA,
Elution Buffer: 20 mM Tris-HCl, pH 7.4
150 mM NaCl, 50 mM CaCl2,
1. Pack the gel in a suitable column, approx.
16 mm in diameter
2. Equilibrate the column with Loading Buffer at a
flow rate of 2 mL/min.
3. Load the sample to be purified using a flow rate
of 0.5 mL/min.
4. Wash the column with Washing Buffer using a
flow rate of 2 mL/min.
5. Once the A280 of the effluent has decreased to
the baseline, elute the bound protein with
Elution Buffer using a flow rate of 2 mL/min.
6. Dialyze the elution fraction against a suitable
buffer (e.g. TBS, Tris Buffered Saline, pH 7.4)
and store at –20°C.
Note: As Ca2+ ions in the sample may inhibit binding, the presence of a chelator such as EDTA (2-10 mM) is recommended during the binding step. In some cases, Ca2+ concentrations >50 mM may enhance the elution step.
Example
Factor X (5 µg) was purified using ADG3570Mx sepharose (500 µl, 1:1 suspension) in a spin column. The fractions were analysed by Western blot analysis using (A) the monoclonal anti-Gla antibody M3B or (B) a polyclonal anti-Factor X antibody. (1= flow through, 2-4 Wash, 5-8 Eluate, M= Protein marker)
![938107_ADG3570Mx](/uploads/artikeldetails/938107_adg3570mx.jpg)
Presentation
Approximately 1 mL of settled gel immobilized with 5 mg of purified antibody with approximately 1.0 mL of PBS, pH 7.4 with 0.02% NaN3 added as a preservative.
Storage
Store the immobilized gel at 2° - 8°C. The gel should be washed thoroughly with PBS, pH 7.4 pH prior to use to remove traces of the sodium azide preservative.
Category: Research use only
Type: Antibody
Product Availability: Worldwide
Manufacturer: ImmBioMed GmbH & Co KG, Germany
For more information please click .pdf icon below.
References
- Brown, M. A., et al. Journal of Biological Chemistry 2000, 275: 19795-19802.
- Stenberg, L. M. et al. Biochemical and Biophysical Research Communications 2001, 280: 1036–1041.
- Stenberg, L. M. et al. Biochemical and Biophysical Research Communications 2001, 283: 454–459.