Description

PA006838: Biotinylated Recombinant Anti-Myc-Tag Monoclonal Antibody (Clone 9E10), Human IgG1 Fc Silent Kappa (Agarose conjugated)

Agarose conjugated Myc-Tag human IgG1 monoclonal antibody is made by covalently linking a monoclonal anti-Myc antibody to the agarose which enables immunoprecipitation (IP) of Myc tagged proteins or co-immunoprecipitation (Co-IP) of their interacting partners.
Source: This monoclonal antibody is produced by immunizing animals with a synthetic peptide containing epitope Myc (KLH-coupled).
Clone: 9E10.
Isotype: Human IgG1 Fc Silent Kappa.
Specificity: Anti-Myc-Tag Mouse mAb detects transfected proteins containing the Myc epitope tag.
Storage: The product is supplied as a 50% slurry in storage buffer (1xPBS, pH 7.4, containing 0.1% NaN3). Store the product at 4°C and do not freeze.
RECOMMENDED ELUTION BUFFER: 0.2 M Glycine, pH 2.5

PA006839: Biotinylated Recombinant Anti-Myc-Tag Monoclonal Antibody (Clone 9E10), Human IgG1 Fc Silent Kappa (Magnetic Microbead Conjugated)

Magnetic Microbead conjugated Myc-Tag human IgG1 monoclonal antibody is made by covalently linking a monoclonal anti-Myc antibody to the agarose which enables immunoprecipitation (IP) of Myc tagged proteins or co-immunoprecipitation (Co-IP) of their interacting partners.
Source: This monoclonal antibody is produced by immunizing animals with a synthetic peptide containing epitope Myc (KLH-coupled).
Clone: 9E10.
Isotype: Human IgG1 Fc Silent Kappa.
Specificity: Anti-Myc-Tag Mouse mAb detects transfected proteins containing the Myc epitope tag.
Storage: The product is supplied as a 50% slurry in storage buffer (1xPBS, pH 7.4, containing 0.1% NaN3). Store the product at 4°C and do not freeze.
RECOMMENDED ELUTION BUFFER: 0.2 M Glycine, pH 2.5

Immunoprecipitation procedure
The work can be performed in 1.5 ml micro-centrifuge tubes or in spin columns.

1. Thoroughly resuspend the Anti-Myc Agarose by inverting the tube or vial several times.

2. Add 20-50 ul 50% slurry of Anti-Myc Agarose into cell lysate using a wide-bore pipette tip. Note: the lysate should be fresh, and for a well expressed tagged protein, 200 ul lysate (200-500 ug total protein) usually yields a good IP result.

3. Incubate with gentle mixing for 2 h to overnight at 4°C.

4. Wash the beads with 1 ml TBS buffer or lysis buffer, such as RIPA (50 mM Tris HCl, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40, 0.5% sodium deoxycholate), centrifuge for 3 min at 2,000xg, and discard the supernatant. Wash 3 times, avoid losing beads during washes.

5. Elution of the Myc tagged protein.
Option 1. Elution with elution buffer.
Add 30-50 ul elution buffer to the beads, gently tap the tube to mix well, immediately centrifuge for 3 min, transfer the supernatant very carefully to a fresh tube (Avoid transferring any beads).
Note: Neutralize the eluant immediately by add 1 ul of 1.5 M Tris, pH 9.0 per 20 ul Elution buffer.

Option 2. Elution with Myc peptide
Add 30-50 ul Myc peptide solution (100 ug/ml Myc peptide in TBS buffer), gently tap the tube to mix well, incubate for 10 min, centrifuge for 3 min, and transfer the supernant to a fresh tube. TBS buffer: 50 mM Tris HCl, 150 mM NaCl, pH 7.4.

Option 3. Elution with SDS loading buffer
Add 30 ul 2x SDS loading buffer, gently tap the tube to mix well, boil at 100°C for 5 min, centrifuge for 3 min, transfer the supernatant to a fresh tube.
Note: in this case, the supernatant contains not only the binding proteins, but also IgG (heavy and light chains).

6. Prepare SDS-PAGE gel for western blotting or proceed to other assays.