DYKDDDDK Tag (FLAG) Antibody Biotin Conjugated

SKU: orb345881

Description

DYKDDDDK antibody (Biotin)

Images & Validation

• Tested Applications: ELISA, IHC, WB
• Dilution Range: ELISA: 1:10,000 - 1:50,000, IHC: 1:1,000 - 1:5,000, WB: 1:2,000 - 1:10,000
• Application Notes:
  This antibody is optimally suited for monitoring the expression of FLAG tagged fusion proteins. As such, this antibody can be used to identify fusion proteins containing the FLAG epitope. The antibody recognizes the epitope tag fused to the carboxy or amino- terminus of targeted proteins. This antibody has been tested by ELISA and western blotting against both the immunizing peptide and FLAG containing recombinant proteins. Although not tested, this antibody is likely functional for immunoprecipitation and immunocytochemistry, and other immunodetection techniques. The epitope tag peptide sequence was first derived from the 11-amino-acid leader peptide of the gene-10 product from bacteriophage T7. Now the most commonly used hydrophilic octapeptide is DYKDDDDK. Biorbyt's polyclonal antibody to detect FLAG conjugated proteins binds FLAG containing fusion proteins with greater affinity than the widely used monoclonal M1, M2 and M5 clones, and shows greater sensitivity in most assays. Affinity purification of the polyclonal antibody results in very low background levels in assays and low cross-reactivity with other cellular proteins.

Key Properties

• Antibody Type: Primary Antibody
• Host: Rabbit
• Clonality: Polyclonal
• Isotype: IgG
• Immunogen: This antibody was purified from whole rabbit serum prepared by repeated immunizations with the Enterokinase Cleavage Site (ECS) peptide DYKDDDDK (Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys) conjugated to KLH using maleimide. This antibody reacts with FLAG conjugated proteins.
• Purity: This affinity purified antibody is directed against the FLAG motif and is useful in determining its presence in various assays. This polyclonal anti-FLAG tag antibody detects over-expressed proteins containing the FLAG epitope tag. To date this antibody has reacted with all amino- terminal FLAG tagged proteins so far tested. In western blotting of bacterial extracts the antibody does not cross-react with endogenous proteins.
• Conjugation: Biotin

Storage & Handling

• Storage: Store vial at 4° C prior to restoration. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use.
• Form/Appearance: Lyophilized
• Buffer/Preservatives: Preservative: 0.01% (w/v) Sodium Azide. Stabilizer: 10 mg/mL Bovine Serum Albumin (rAlbumin) - Immunoglobulin and Protease free; Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
• Concentration: 1.0 mg/mL
• Expiration Date: 12 months from date of receipt.
• Disclaimer: For research use only

Alternative Names

rabbit antibody for the detection of FLAG conjugated proteins biotin conjugated, rabbit anti DYKDDDDK Biotin

Affinity Purified Antibody to detect FLAG™ conjugated proteins detects both C terminal linked and N terminal linked FLAG™ tagged recombinant proteins by western blot. This antibody was used at a dilution of 1:2500 to detect 1.0 µg of recombinant protein containing either the FLAG™ epitope tag linked at the carboxy (C) or the amino (N) terminus of the recombinant protein. A 4-20% gradient gel was used to resolve the protein by SDS-PAGE. The protein was transferred to nitrocellulose using standard methods. After blocking, the membrane was probed with the primary antibody for 1 h at room temperature followed by washes and reaction with a 1:10000 dilution of IRDye® 800 conjugated Gt-a-Rabbit IgG (H&L) MX10 for 30 min at room temperature.

Biorbyt's antibody to detect FLAG™ conjugated proteins is shown to detect as little as 3 ng of amino-terminal FLAG™ tagged recombinant protein by western blot. This antibody was used at a 1:1000 dilution to detect 3-fold serial dilutions of amino-terminal FLAG™-Bacterial Alkaline Phosphatase (BAP) fusion protein (Sigma P-7582) starting at 1.0 µg of protein as shown in lanes 1-6 respectively. A 4-20% gradient gel was used to separate the protein by SDS-PAGE. The protein was transferred to nitrocellulose using standard methods. After blocking, the membrane was probed with the primary antibody for 1 h at room temperature followed by washes and reaction with a 1:10000 dilution of IRDye® 800 conjugated Gt-a-Rabbit IgG (H&L) for 30 min at room temperature.