Confocal immunofluorescent analysis of COS-7 cells, transiently transfected with a DYKDDDDK-tagged fusion protein, using DYKDDDDK Tag Antibody (Binds to same epitope as Sigma's Anti-FLAG® M2 Antibody) (Alexa Fluor® 594 Conjugate) (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).Learn more about how we get our images
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% formaldehyde diluted in 1X PBS.
NOTE: Formaldehyde is toxic, use only in a fume hood.
NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.
posted November 2006
revised November 2013
Protocol Id: 182
Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.
DYKDDDDK Tag Antibody (Binds to same epitope as Sigma's Anti-FLAG® M2 Antibody) (Alexa Fluor® 594 Conjugate) detects exogenously expressed DYKDDDDK-tagged proteins in cells. The antibody recognizes the DYKDDDDK peptide fused to either the amino- or carboxy-terminus of targeted proteins. The binding specificity of this antibody is NOT dependent on the presence of divalent metal cations.
All Species Expected
Polyclonal antibodies are produced by immunizing animals with a synthetic DYKDDDDK peptide.
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.
The DYKDDDDK peptide has been used extensively as a general epitope tag in expression vectors. This peptide can be expressed and detected with the protein of interest as an amino-terminal or carboxy-terminal fusion (1).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. Anti-FLAG is a registered trademark of Sigma-Aldrich Biotechnology. DRAQ5 is a registered trademark of Biostatus Limited. The Alexa Fluor dye antibody conjugates in this product are sold under license from Life Technologies Corporation for research use only, except for use in combination with DNA microarrays. The Alexa Fluor® dyes (except for Alexa Fluor® 430 dye) are covered by pending and issued patents. Alexa Fluor® is a registered trademark of Molecular Probes, Inc.