The Thermo Scientific Pierce Magnetic c-Myc-Tag IP/Co-IP Kit contains specific immunoaffinity magnetic beads and reagents to perform immunoprecipitation assays of c-Myc fusion proteins or co-IP experiments using c-Myc-tagged bait proteins.
Unlike traditional IP procedures based on capture with Protein A/G beads, this kit uses magnetic beads containing pre-immobilized anti-c-Myc antibody. These Pierce Anti-c-Myc Magnetic Beads ensure specific binding of c-Myc 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, c-Myc-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.
- Specific magnetic beads – covalently immobilized high-quality anti-c-Myc 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 c-Myc-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)
The c-Myc peptide (EQKLISEEDL) derived from the C-terminus region of human c-Myc protein is one of several fusion protein tags used for recombinant protein expression. The Pierce c-Myc Magnetic IP/Co-IP Kit uses a specific, high-affinity immobilized antibody (clone 9E10) for rapid immunoprecipitation of c-Myc 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 c-Myc 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-c-Myc antibody can be used to detect c-Myc 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-c-Myc Magnetic Beads (Part No. 88842).
Better immunoprecipitation results with Thermo Scientific Pierce Anti-c-Myc Magnetic Beads. Green Renilla luciferase c-Myc fusion protein was expressed in 293T cells. For IP, identical aliquots of the cell lysate were incubated in duplicate for one hour at room temperature with anti-c-Myc magnetic beads from each manufacturer. For all conditions, IP products were eluted identically using low-pH buffer. Eluted fractions (25µL each) were separated by 12% SDS-PAGE, transferred to PVDF membrane, and detected via anti-c-Myc antibody (Part No. MA1-980), goat anti-mouse secondary antibody, and chemiluminescent substrate (Part No. 34080). Note: Manufacturers supply magnetic beads slurries at different concentrations and recommend different amounts of beads per microgram of lysate for IP. For this reason, only 10µL of CST beads were used in this experiment. Nevertheless, Pierce beads provide equal or greater binding capacity, even in experiments involving carefully matched numbers of beads (data not shown).
Co-IP of BCL2-Luc with c-Myc-tagged BAD in 293T cells. This experiment tests the specificity of co-immunoprecipitation by comparing results from 293T cells cultured alone (293T) or with vectors expressing BCL2-Luc and/or BAD-c-Myc fusion proteins (labeled BCL2 and BAD, respectively). The Western blot reveals the presence of BCL2 detected through its fused luciferase domain (i.e., via an anti-Luc antibody). Although BCL2 is expressed in two cultures (as indicated by its detection in two of the Control Lysates), it is purified by anti-c-Myc magnetic beads (IP or Co-IP lanes) only when co-expressed and captured through its interaction with c-Myc-tagged BAD (BAD+BCL2 lane). For the IP experiments, lysates were incubated with anti-c-Myc magnetic beads (50µL) for two hours at 4°C, then processed and eluted using the kit components and procedure.
Epitope Tag Product Selection Guide – links to antibodies, resins, magnetic beads and IP kits
Other Anti-Tag Antibodies
Anti-c-Myc Magnetic Beads
Fusion Protein Purification Resins and Kits
KingFisher Magnetic Particle Processors
Cell Lysis and Protein Extraction Reagents