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Pierce HA-Tag Magnetic IP/Co-IP Kit


Magnetic beads for efficient isolation of HA-tagged proteins and interactors.

Pierce HA-tag Magnetic IP/Co-IP Kit

The Thermo Scientific Pierce Magnetic HA-Tag IP/Co-IP Kit contains specific immunoaffinity magnetic beads and reagents to perform immunoprecipitation assays of HA fusion proteins or co-IP experiments using HA-tagged bait proteins.

Unlike traditional IP procedures based on capture with Protein A/G beads, this kit uses magnetic beads containing pre-immobilized anti-HA antibody. These Pierce Anti-HA Magnetic Beads ensure specific binding of HA-tagged protein complexes from biological samples. Because the antibody is covalently attached to the beads, IP-targets are easily eluted and recovered without antibody contamination. The complete IP kit includes the magnetic beads, lysis/wash buffer, low-pH elution buffer, neutralization buffer, HA-tag positive control lysate, and non-reducing sample buffer for SDS-PAGE. Protocols are provided for both manual and automated magnetic separation workflows. Sufficient components are provided to perform 40 IP or co-IP assays.

Highlights:

  • Specific magnetic beads – covalently immobilized high-quality anti-HA monoclonal antibody enables high yields of immunoprecipitation products
  • Low non-specific binding – stable, pre-blocked beads and specific antibody minimize off-target binding
  • Trouble-free elution – low-pH elution buffer ensures recovery of HA-tagged protein interaction complexes without antibody leaching contamination
  • Convenient and fast – complete kit and easy-to-follow instructions provide optimized protocols to perform IP or Co-IP experiment in approximately 1 hour
  • Versatile – magnetic beads are compatible with manual and automated magnetic separation workflows (e.g., Thermo Scientific KingFisher Instruments)

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 (clone 2-2.2.14) 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.

For information about the binding capacity and other properties of the magnetic beads used in this kit, see Pierce Anti-HA Magnetic Beads (Part No. 88836).

Immunoprecipitation of HA-tagged protein with magnetic beads 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.

 

Co-IP with HA-tagged proteins using magnetic beads 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 Resources:

Epitope Tag Product Selection Guide – links to antibodies, resins, magnetic beads and IP kits

Related Products:

Anti-HA Magnetic Beads
Anti-HA Agarose
HA-Tag IP/Co-IP Kit (agarose)
Anti-HA Antibody and other Anti-Tag Antibodies
Fusion Protein Purification Resins and Kits
KingFisher Magnetic Particle Processors
Cell Lysis and Protein Extraction Reagents


Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
88838 HA-Tag Magnetic IP/Co-IP Kit
Formulation: Six-component kit
Sufficient For: 40 IP reactions using 25µL of anti-HA magnetic beads

Kit Contents:
HA-tagged Positive Control (Part No. 26180X), 500μL

Application Set (Part No. 88838X):
Pierce Anti-HA Magnetic Beads, 1mL
Pierce IP Lysis/Wash Buffer, 2 × 50mL
Non-Reducing Sample Buffer (5X), 5mL
Elution Buffer (pH 2.0), 5mL
Neutralization Buffer (pH 8.5), 1mL
40-rxn kit Product Instructions for product #88838 HA-Tag Magnetic IP/Co-IP Kit MSDS for product #88838 HA-Tag Magnetic IP/Co-IP Kit Certificate of Analysis for product #88838 HA-Tag Magnetic IP/Co-IP Kit $413.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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