The Thermo Scientific Pierce Magnetic IP/Co-IP Kit provides the magnetic beads, positive control and reagents to perform immunoprecipitation (IP) assays of HA-tagged proteins or co-IP reactions with HA-tagged bait proteins.
The complete Magnetic IP/Co-IP Kit can be used to perform 40 IP or Co-IP assays. It includes magnetic beads coated with immobilized anti-HA antibody, lysis/wash buffer, low pH elution buffer, neutralization buffer, HA-tag positive control lysate, and non-reducing sample buffer. Protocols are provided for both manual and automated workflows.
Highlights:
- Specific – highly specific anti-HA monoclonal antibody enables high yields of immunoprecipitation products
- Fast – Optimized protocol is completed in approximately 1 hour
- Low non-specific binding – stable, pre-blocked beads provide highly purified HA-tagged proteins
- Versatile – beads are compatible with manual and automated workflows (e.g., Thermo Scientific KingFisher Instruments)
- Convenient and easy – complete kit includes all necessary reagents and easy-to-follow instructions
Includes:
The kit contains magnetic beads immobilized with anti-HA antibody, lysis/wash buffer, low pH elution buffer, neutralization buffer, HA-tag positive control lysate, and non-reducing sample buffer.
Applications:
- High-throughput immunoprecipitation of recombinant HA-tagged proteins and co-IP of interacting proteins
Product Details:
The hemagglutinin (HA) peptide (YPYDVPDYA), derived from the human influenza virus HA protein, is one of several fusion protein tags used for recombinant protein expression. The Pierce Magnetic HA-Tag IP/Co-IP Kit uses a specific, high-affinity immobilized antibody for rapid purification of HA-tagged fusion proteins from bacterial and mammalian cell lysates, as well as from lysates prepared with the Pierce Human in vitro Translation Kits. The beads are incubated with a cell lysate containing HA-tagged protein, the fusion protein is captured, and the beads are subsequently washed and then eluted using low pH elution buffer or non-reducing sample buffer. The protocol and buffers have been optimized for both IP and co-IP reactions, enriching for specific protein interaction complexes in the eluted samples. Anti-HA antibody can be used to detect HA-tagged protein by Western blot analysis.
| Characteristics of Thermo Scientific Pierce Anti-HA Magnetic Beads. |
| Composition: |
High-affinity mouse IgG1 monoclonal antibody covalently coupled to the surface of blocked magnetic beads |
| Magnetization: |
Superparamagnetic (no magnetic memory) |
| Mean Diameter: |
1μm (nominal) |
| Density: |
2.0g/cm3 |
| Bead Concentration: |
10mg/mL |
| Binding Capacity: |
≥10μg of GST-ERK-HA (70kDa fusion protein)/mg of beads |
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Immunoprecipitation of HA-tagged protein with the Thermo Scientific Pierce Magnetic HA-Tag IP/Co-IP Kit. HA-tagged BAD was expressed using a Pierce in vitro Expression System. Using the KingFisher Flex Instrument, 25μL each of Pierce Anti-HA Magnetic Beads or Anti-HA-tag Magnetic Beads (MBL International Corp.) were added to a 96 deep-well plate. The beads were incubated for 1 hour at room temperature with the HA-tagged BAD expressed in HeLa lysate, washed twice in IP Lysis/Wash Buffer, washed once in water and then eluted with low-pH Elution Buffer, pH 2.0. Samples were resolved by SDS-PAGE and analyzed by Western blot using the Thermo Scientific Anti-HA Monoclonal Antibody (above) and by silver stain for non-specific binding (below). The Pierce beads produced a higher yield of HA-BAD than MBL beads. Non-specific binding was minimal for both bead samples tested. |
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Better co-immunoprecipitation (co-IP) with Thermo Scientific Pierce Anti-HA Magnetic Beads. HA-tagged BAD was expressed in a Pierce in vitro Expression System which includeas a HeLa lysate containing 14-3-3 proteins, a family of multifunctional regulatory proteins that bind BAD in vivo. Using a magnetic stand, 50μL each of Pierce Anti-HA Magnetic Beads and Anti-HA-tag Magnetic Beads (MBL International Corp.) were added to microcentrifuge tubes. The beads were incubated for 1 hour at room temperature with lysate containing HA-tagged BAD, washed twice with PBS containing 0.05% Tween*-20, washed once with water and then eluted in 30% acetonitrile/0.5% formic acid. Samples were dried in a Speedvac concentrator and reconstituted in reducing SDS-PAGE sample buffer. One half of each eluate was resolved by SDS-PAGE and analyzed for 14-3-3 by Western blot (above) and by silver stain (below). The Pierce Anti-HA Magnetic Beads detected co-immunoprecipitated 14-3-3 whereas the MBL beads did not. Non-specific binding was minimal for both beads tested. |
Related Products:
Pierce Anti-HA Magnetic Beads
Other Pierce Magnetic Beads
Pierce HA-Tag IP/Co-IP Kit (agarose)
Pierce Anti-HA Agarose and Peptide
Anti-HA Antibody
KingFisher Magnetic Particle Processors
Cell Lysis Reagents
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