WB result of FLAG Tag Recombinant Rabbit mAb
Primary antibody: FLAG Tag Recombinant Rabbit mAb at 1/10000 dilution
Lane 1: 293T transfected with empty vector whole cell lysate 20 µg
Lane 2: 293T expressing fusion protein with FLAG tag in the middle whole cell lysate 20 µg
Lane 3: 293F expressing C-Flag fusion protein whole cell lysate 20 µg
Lane 4: 293F expressing N-Flag fusion protein whole cell lysate 20 µg
Secondary antibody: Goat Anti- rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution
Predicted MW: 15 kDa
Observed MW: 22 kDa
FLAG Tag Recombinant Rabbit mAb (S-SC044)
FLAG Tag Recombinant Rabbit mAb (S-SC044)
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Product Details
Product Details
Product Specification
| Host | Rabbit |
| Clone Number | S-SC044 |
| Antibody Type | Recombinant mAb |
| Isotype | IgG |
| Application | WB, ICC, IP |
| Reactivity | Species Independent |
| Purification | Protein A |
| Concentration | 0.5 mg/ml |
| Conjugation | Unconjugated |
| Physical Appearance | Liquid |
| Storage Buffer | PBS, 40% Glycerol, 0.05% BSA, 0.03% Proclin 300 |
| Stability & Storage | 12 months from date of receipt / reconstitution, -20 °C as supplied |
Dilution
| application | dilution | species |
| WB | 1:5000-1:50000 | Species Independent |
| IP | 1:50 | Species Independent |
| ICC | 1:500 | Species Independent |
Background
FLAG-tag, or FLAG octapeptide, or FLAG epitope, is a peptide protein tag that can be added to a protein using recombinant DNA technology, having the sequence DYKDDDDK (where D=aspartic acid, Y=tyrosine, and K=lysine). It is one of the most specific tags and it is an artificial antigen to which specific, high affinity monoclonal antibodies have been developed and hence can be used for protein purification by affinity chromatography and also can be used for locating proteins within living cells. FLAG-tag has been used to separate recombinant, overexpressed protein from wild-type protein expressed by the host organism. FLAG-tag can also be used in the isolation of protein complexes with multiple subunits.
Picture
Picture
Western Blot
IP
FLAG Tag Rabbit mAb at 1/50 dilution (1 µg) immunoprecipitating FLAG Tag in 0.4 mg 293F expressing C-Flag fusion protein whole cell lysate.
Western blot was performed on the immunoprecipitate using FLAG Tag Rabbit mAb at 1/10000 dilution.
Secondary antibody (HRP) for IP was used at 1/1000 dilution.
Lane 1: 293F expressing C-Flag fusion protein whole cell lysate 20 µg (Input)
Lane 2: FLAG Tag Rabbit mAb IP in 293F expressing C-Flag fusion protein whole cell lysate
Lane 3: Rabbit monoclonal IgG IP in 293F expressing C-Flag fusion protein whole cell lysate
Predicted MW: 15 kDa
Observed MW: 22 kDa
Immunocytochemistry
ICC shows positive staining in FLAG tag fusion protein transfected 293F cells (top panel) and negative staining in vector-transfected 293F cells (below panel). Anti- FLAG Tag antibody was used at 1/500 dilution (Green) and incubated overnight at 4°C. Goat polyclonal Antibody to Rabbit IgG - H&L (Alexa Fluor® 488) was used as secondary antibody at 1/1000 dilution. The cells were fixed with 100% ice-cold methanol and permeabilized with 0.1% PBS-Triton X-100. Nuclei were counterstained with DAPI (Blue).
