Western blot analysis of extracts from COS cells transfected with Myc-Chorionic Somatomammotropin Hormone-Like 1 (CSL) protein (lane 1). Immunoprecipitation of COS/CSL lysates with Mouse IgG (Sepharose® Bead Conjugate) #3420 (lane 2) or Myc-Tag (9B11) (Sepharose® Bead Conjugate) Mouse mAb (lane 3). Blots were probed with Myc Tag (71D10) Rabbit mAb #2278.
This Cell Signaling Technology antibody is immobilized via covalent binding of primary amino groups to N-hydroxysuccinimide (NHS)-activated Sepharose® beads. Myc-Tag (9B111) Mouse mAb (Sepharose® Bead Conjugate) is useful for the immunoprecipitation of Myc-tagged recombinant proteins.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol. Store at –20°C. Do not aliquot the antibodies.
This protocol is intended for immunoprecipitation of native proteins for analysis by western immunoblot or kinase activity.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
10X Cell Lysis Buffer: (#9803) 20 mM Tris (pH 7.5), 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton X-100, 2.5 mM Sodium pyrophosphate, 1 mM β-glycerophosphate, 1 mM Na3VO4, 1 μg/ml Leupeptin
NOTE: CST recommends adding 1 mM PMSF (#8553) before use*.
Proceed to one of the following specific set of steps.
NOTE: To minimize masking caused by denatured IgG heavy chains (~50 kDa), we recommend using Mouse Anti-Rabbit IgG (Light-Chain Specific) (L57A3) mAb (#3677) or Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127). To minimize masking caused by denatured IgG light chains (~25 kDa), we recommend using Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127).
posted December 2007
Protocol Id: 27
Myc-Tag (9B11) Mouse mAb detects transfected proteins containing the Myc epitope tag. The antibody recognizes the Myc-tag fused to either the amino- or carboxy-termini of targeted proteins in transfected cells.
All Species Expected
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues 410-419 of human c-Myc (EQKLISEEDL).
Epitope tags are useful for the labeling and detection of proteins using immunoblotting, immunoprecipitation, and immunostaining techniques. Because of their small size, they are unlikely to affect the tagged protein’s biochemical properties.
Epitope tags are useful for the labeling and detection of proteins using immunoblotting, immunoprecipitation and immunostaining techniques. Due to their small size, they are unlikely to affect the tagged protein's biochemical properties.
The Myc epitope tag is widely used to detect expression of recombinant proteins in bacteria, yeast, insect and mammalian cell systems (1).