DYKDDDDK tag polyclonal antibody (Biotin)
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More Files
- More Functions
Western Blot
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 1:2,500 to detect 1.0 ug 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 1:10,000 dilution of IRDye® 800 conjugated Gt-a-Rabbit IgG (H&L) MX10 (code 611-132-122) for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.
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Specification
Product Description
Rabbit polyclonal antibody raised against synthetic peptides of FLAG epitope tag. The antibody is conjugated with Biotin.
Immunogen
A synthetic peptide (conjugated with KLH) corresponding to DYKDDDDK tag.
Sequence
DYKDDDDK
Host
Rabbit
Specificity
Carboxy and amino terminal linked FLAG tagged recombinant proteins. 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 affinity purified antibody is directed against the FLAG motif and is useful in determining its presence in various assays. This polyclonal 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.
Form
Lyophilized
Conjugation
Biotin
Preparation Method
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. Residues of glycine and cysteine were added to the carboxy terminal end to facilitate coupling. This antibody reacts with FLAG conjugated proteins.
Purification
Affinity purification
Isotype
IgG
Conjugation Note
FP Value: 10-20 moles Biotin per mole of IgG
Recommend Usage
ELISA (1:10000-1:50000)
Immunohistochemistry (1:1000-1:5000)
Western Blot (1:2000-1:10000)
The optimal working dilution should be determined by the end user.Storage Buffer
Lyophilized from 0.02 M potassium phosphate, 0.15 M sodium chloride, pH 7.2 (10 mg/mL BSA (immunoglobulin and protease free), 0.01% (w/v) sodium azide).
Storage Instruction
Store vial at 4°C prior to restoration.
After reconstitution with 100 uL deionized water (or equivalent), aliquot contents and freeze at -20°C or below for extended storage.
Aliquot to avoid repeated 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.Note
This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
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Applications
Western Blot
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 1:2,500 to detect 1.0 ug 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 1:10,000 dilution of IRDye® 800 conjugated Gt-a-Rabbit IgG (H&L) MX10 (code 611-132-122) for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.Immunohistochemistry
Enzyme-linked Immunoabsorbent Assay
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