Thermo Scientific Pierce Anti-HA Agarose is a high-affinity immunoprecipitation resin ideal for purification of recombinant HA-tagged proteins expressed in human in vitro expression systems and bacterial and mammalian cell lysates.
The anti-HA antibody used to manufacture Pierce Anti-HA Agarose is a highly specific mouse IgG1 monoclonal antibody (clone 2-2.2.14) that recognizes the HA epitope tag (YPYDVPDYA) derived from the human influenza hemagglutinin (HA) protein. The support is crosslinked 4% beaded agarose. This anti-HA affinity resin can be packed into gravity purification columns, spin purification columns or cartridges for FPLC instruments to purify HA-fusion proteins expressed in bacterial or mammalian cells.
- Specific – highly specific anti-HA monoclonal antibody (clone 2-2.2.14) enables high yield and high purity for immunoprecipitation
- Scalable – available in 2 and 10mL resin package sizes to allow for larger scale purifications or immunoprecipitations
- Versatile – can be used in spin or gravity columns as well as in FPLC cartridges
- Convenient – reagents to elute and detect HA-tagged fusion proteins are available separately
Pierce Anti-HA Agarose consists of a highly specific monoclonal anti-HA antibody that is covalently immobilized on a crosslinked 4% beaded agarose support. The product is supplied as a 50% slurry. Upon incubation with a sample, HA-tagged fusion protein is captured on the agarose beads. After simple washing steps, the tagged protein is easily eluted from the resin using 0.1M glycine (pH 2.0-2.8), 3M NaSCN or 50mM NaOH depending on the downstream application of the purified protein. For elution of highly functional proteins, Pierce HA-peptide can also be used to elute the HA-tagged protein. Anti-HA antibody may be used to detect the presence of the tagged protein by Western blot.
In experiments with a HA-tagged fusion protein (37kDa), the resin provided a binding capacities of 60 to 150nmol protein per mL of settled resin. The elution capacity was at least 6 to 15nmol per mL of settled resin using 3M NaSCN. Binding and elution capacity will vary depending on the HA-fusion protein and the method of elution.
||Immunoprecipitation of HA-tagged protein (GST-PI3K-SH2-HA) from E. coli lysate with the Thermo Scientific Pierce HA-Tag IP/Co-IP Kit. Coomassie Stained gel. Lane 1: MW marker, Lane 2: 15µL of 0.25mg/mL of positive control lysate, Lane 3: 15µL of Flow-through, Lane 4: 15µL of Glycine, pH 2.8 elution fraction.
||Purification of HA-tagged protein from the Pierce human in vitro expression lysate. HA-tagged GFP (from Pontellina plumata) was expressed with the Pierce Human In Vitro Protein Expression System and purified by incubation with Pierce Immobilized Anti-HA Agarose. Anti-HA agarose slurry (50µL) was added to a Pierce Spin column (Part No. 69705) along with 60µL of the in vitro translation reaction diluted to a final volume of 200µL in TBS. The resin and sample were mixed for 1 hour at 4°C with end-over-end mixing. The resin was pelleted by centrifugation and was washed 3X with 10 column volumes of TBS-T. HA-tagged GFP was eluted from the resin by adding 3 x 1 column volume of 1mg/mL Pierce Influenza Hemagglutinin (HA) Peptide (Part No. 26184). The elution fractions were combined and 20% of the elution fraction was electrophoresed in SDS-PAGE and stained with Gel Code Blue (Part No. 24590). Lane 1: immunoprecipitation eluate from human in vitro translation reaction containing no DNA (negative control), Lane 2: immunoprecipitation eluate from human in vitro translation reaction containing GFP-HA DNA, Lane 3: MW Marker
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Epitope Tag Product Selection Guide – links to antibodies, resins, magnetic beads and IP kits
IgG Elution Buffer
HA-Tag IP/Co-IP Kit (agarose)
HA-Tag Magnetic IP/Co-IP Kit (magnetic)
Anti-HA Magnetic Beads
Anti-HA Antibody and other Anti-Tag Antibodies
Fusion Protein Purification Resins and Kits
Cell Lysis and Protein Extraction Reagents