MagVigen™ – SH surface nanoparticles provide you with the flexibility of coupling to various molecules through simple bioconjugation reactions. The resulting MagVigen™ bioconjugates could be exploited to achieve highly specific binding for cell isolation, protein, DNA/RNA purification or immunoprecipitation assays. Examples of biomolecules that could be covalently bound to MagVigen™ surfaces include primary antibody, protein/peptide, DNA/RNA or other ligands.
Advantages of MagVigen™ magnetic nanoparticles
- Magnetically responsive to a magnet, easy for bio-conjugation and purification
- Smaller nanoparticle size, higher binding capacity, longer settling time, compatible to automation and high throughput workflow
- Optimal surface chemistry, low non-specific binding
- Consistent, high quality results
Product and Related Product Contents
- MagVigen™ – SH surface nanoparticles (Cat# 21002) are provided in phosphate buffered saline (PBS), pH 7.4. Each vial contains 1 ml of solution with a particle concentration of 4 mg/ml.
- Washing Buffer (10X), Cat# A20001.
- Magnet, Cat# A20003.
All materials except the magnet should be stored at 4°C for up to 1 year.
Antibody Conjugation to MagVigen™ -SH Surface
- Determine needed surface coverage of antibody per nanoparticle.
- Desalt Antibody-SMCC using NAP column.
- Mix purified Antibody-SMCC with MagVigen™-SH nanoparticles. Incubate overnight under continuous rotation at room temperature.
- Separate out MagVigen™-Antibody by magnetic purification.
- Wash 1-3 times with PBS or other buffer solution. Remove non-magnetically captured solution.
Note 1: The general range is about 0.3 -1.5 mg antibody per mg of MagVigen™.
Mix Antibody in water with SMCC (Succinimidyl trans- 4(maleimidylmethyl) cyclohexane-1-carboxylate) based crosslinker in PBS solution. Incubate for 40-60 min.
Note 2: SMCC is used to crosslink the amine groups on antibody with the –SH groups from MagVigen™. SMCC based crosslinker is suggested because of its superior chemical stability when used with our nanoparticles and its ease of use.
Note 3: The ratio of SMCC to antibody is 1:10.
Note: This step removes the free SMCC from the Antibody- SMCC mixture.
Note: One wash could be sufficient for most applications.
Resuspend washed MagVigen™-Antibody conjugates into preferred buffer, ready to use.
This protocol was optimized for enrichment of 1-10 μg rabbit or mouse antibody in a volume of 100 μl. For a smaller size of sample, it is recommended to add extra Washing Buffer to reach a 100 μl reaction volume. For larger scale of purification, adjust the amount of reagents accordingly.
- Dilute 10X Washing Buffer with PBS to 1X.
- Vortex MagVigen™ nanoparticles for 10-20 seconds.
- Take 5-50 μl nanoparticle solution (for 1-10 μg antibody), add it to 100 μl 1X Washing Buffer, and vortex to mix.
- Separate the nanoparticles from the solution by placing the magnet on the side of the tube for 2-5 min and remove the supernatant carefully (with magnet still on the side).
- Remove magnet and wash the nanoparticles with 100 μl 1X Washing Buffer. Repeat step 4, and remove supernatant.
- Add 100 μl sample solution containing desired antibodies to the nanoparticle pellet, mix well, and incubate with gentle rotation for 2 hours at room temperature or 4°C overnight.
- After incubation, use the magnet to separate nanoparticle- antibody complex from the solution and remove the supernatant.
- Wash nanoparticle-antibody complex with 100 μl 1X Washing Buffer twice and remove supernatant.
- Elute captured antibody from the nanoparticles by adding 90 μl Elution Buffer, mix well, and incubate for 1 min at room temperature.
- Separate the nanoparticles from the eluted antibody with magnet. Transfer supernatant to a clean tube and immediately neutralize the eluate by adding 10 μl Tris (1M, pH=8.0). The enriched antibody is ready to use for subsequent evaluation.
Note: A clear precipitate containing dark brown colored nanoparticles should become visible on the side of the micro-centrifuge tube.
MagVigen™ for Antibody Purification and Concentrating:
General Optimal Proportion: 1 mg of magnetic beads for 50 μg of antibody. The proportion may be optimized based on specific antibody or protein property.
Note for Large Volume (>50 ml) Protein Purification:
- Increase incubation time to ensure yield
- Increase magnetic pull down time to ensure majority of beads forming the pellet, this could be confirmed by total clearness of the supernatant.