Anti-His Magnetic Beads 

MCE Anti-His Magnetic Beads are used for immunoprecipitation (IP) of specific His-tagged proteins expressed in bacterial and mammalian cells and in vitro expression systems. Anti-His magnetic beads are based on carboxyl magnetic beads, with 2 μm particle size, covalently coupling with high quality mouse IgG monoclonal antibody that recognizes the His tag. Magnetic beads are removed from the solution manually by using a magnetic stand or automatically by using an instrument. With high loading of His-tagged protein and high specificity, Anti-His Magnetic Beads are also suitable for Co-immunoprecipitation and purification of His-tagged protein.

4°C, 2 years.

Do not centrifuge, dry or freeze the magnetic beads.

1. Preparation of Magnetic Beads

1.1 Resuspend the Magnetic Beads in the vial (tilt and rotate for 2 minutes or gently pipette for 10 times, do not vortex). Transfer 25 μL of Anti-c-Myc Magnetic Beads suspension into a new tube.

1.2 Add 500 μL of wash buffer to the beads and gently pipette to mix. Place the tube into a magnetic stand (MCE Cat. No.: HY-K0200) to collect the beads against the side of the tube. Remove and discard the supernatant. Repeat this step for 2 times

2. Protein Binding

2.1 Add 500 μL of cell lysate (the sample containing c-Myc-tagged protein) to the washed beads. For Ag binding, incubate for 2 hours at room temperature or overnight at 4°C while gently rotating the tube.

2.2 Place the tube into a magnetic stand to collect the beads against the side of the tube. Remove and discard the supernatant.

NOTE: Occasional aggregation of magnetic beads during the binding process doesn’t affect experimental results.

3. Washing

Add 500 μL of wash buffer to the Magbeads-Ag complex and mix gently. Place the tube into a magnetic stand to collect the beads against the side of the tube. Remove and discard the supernatant. Repeat this step for 3 times.

4. Elution & Detection

Three elution methods are recommended according to protein characteristics or further usage:

1) Elution with sample buffer for gel electrophoresis and immuoblotting. Add 100 μL of 1× SDS-PAGE loading buffer to each tube and boil for 5 minutes. Cool and place the tube into a magnetic stand to collect the beads and transfer the supernatant to a new tube. Keep the supernatant containing the target antigen for SDS-PAGE analysis

2) Elution with Elution Buffer A under acidic condition. Add 100 μL of Elution Buffer A to each tube. Incubate with gentle shaking or on a rotator for 10 minutes at room temperature. Place the tube into a magnetic stand to collect the beads and transfer the supernatant to a new tube. Adding 10-20 μL of Neutralization Buffer for each 100 μL of eluate to neutralize the low pH, which may help preserve bioactivity of target protein.

3) Elution with Elution Buffer B under native condition. Add 3-5 (v/v) volume of Elution Buffer B to each tube. Incubate with gentle shaking or on a rotator for 1 hour at room temperature or 2 hours at 4°C. Place the tube into a magnetic stand to collect the beads and transfer the supernatant to a new tube. For immediate use, store the eluates at 4°C, or store at -20°C for long term storage.