Rabbit Polyclonal ChIP Grade Anti-GFP Antibody from Abcam

Abcam’s ChiP-grade anti-GFP antibody is designed to detect 5 ng of recombinant GFP, within a minute of exposure to film.

  • Product Name - Anti-GFP antibody - ChIP Grade
  • Description - Rabbit polyclonal to GFP - ChIP Grade
  • Host Species - Rabbit
  • Tested Applications - It is suitable for ChIP, ICC/IF, ELISA, Flow Cyt, IHC-Wholemount, ChIP/Chip, IHC-FrFl, WB, IHC-Fr, IHC-P, ICC, IHC-FoFr, and electron microscopy.
  • Species Reactivity - It is species-independent.
  • Immunogen - It is a recombinant full-length protein that corresponds to GFP.

Positive Control

Abcam’s antibody detects 5 ng of recombinant GFP (utilizing ECL Plus or ECL) within a minute of exposure to film. This also includes rabbit reticulocytes and Human HEK293 cells.

General Notes

The overall concentration of IgG has been established to be 5 mg/mL; however, the specific IgG concentration is not known. During short-term storage, the product should be kept refrigerated all the time. Risk of bacterial contamination can be reduced with sterilized equipment.

The ab290 is an extremely versatile antibody that provides a more intense signal when compared to other commercially available anti-GFP antibodies. On Western blot (WB), the antibody identifies the GFP fraction from cell extracts that express recombinant GFP fusion proteins. In addition, the antibody has been demonstrated to be useful on mouse sections attached with formalin.

In immunocytochemistry, the ab290 antibody offers an excellent signal on recombinant YES-GFP chimeras that are expressed in COS cells (McCabe et al. 1999). It is often utilized in IP-Western and Immunoprecipitation (IP) protocols and has also been effectively utilized in HRP immunohistochemistry at a ratio of 1:200 on whole-mount mouse embryos. Moreover, ab6556 is a purified version of this antibody.

The anti-GFP antibody also detects the improved form of GFP. It even detects yellow fluorescent protein (YFP) — a genetic mutant of green fluorescent protein (GFP) — as well as venus YFP.

Applications

Abcam’s Abpromise guarantee covers the usage of ab290 in the tested applications listed in the table below. Recommended starting dilutions have been mentioned; the end-user should determine the ideal concentrations/dilutions.

Application Abreviews Notes
Flow Cyt ☆☆☆☆☆ 1/50–1/300.
The ab171870-Rabbit polyclonal IgG can also be used as an isotype control with this antibody.
ELISA   Should be used at an assay dependent concentration.
ICC/IF ☆☆☆☆☆ 1/500–1/1000.
Abcam recommends Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150081) secondary antibody.
ChIP ☆☆☆☆☆ Should be used at an assay dependent concentration.
IHC-FrFl ☆☆☆☆☆ Should be used at an assay dependent concentration.
ChIP/Chip ☆☆☆☆☆ 5 µg should be used for 1 µg of chromatin. PubMed: 17289569.
IHC-Wholemount ☆☆☆☆☆ Should be used at an assay dependent concentration. PubMed: 27287807.
Electron Microscopy   1/1000–1/4000
IHC-FoFr ☆☆☆☆☆ 1/200–1/500
ICC ☆☆☆☆☆ 1/200–1/1000.
Abcam recommends Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150081) secondary antibody.
IHC-P ☆☆☆☆ 1/500–1/1000. Heat-mediated antigen retrieval should be performed through the microwave technique before starting with IHC staining protocol.
IHC-Fr ☆☆☆☆☆ Should be used at an assay dependent concentration. It has been reported to work at dilutions of up to 1/3000. Secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) should be used.
IP ☆☆☆☆☆ Should be used at an assay dependent concentration. Should be used at 1 µL per 10 cm tissue culture dish (10-µL protein A agarose CL4B should also be used to precipitate the immune complex).
WB ☆☆☆☆☆ 1/1000–1/2500.
It is best to utilize 12.5% SDS-PAGE and to transfer to PVDF membrane. In addition, 1x Blotto (or 3% BSA in PBS) should be used for diluting and blocking purposes. PBS should be used in 3 x 5 minutes washing steps all through the immunolabeling process.
It is recommended to probe with ab290 at 1:1000–1:5000 dilution and utilize Goat Anti-Rabbit IgG H&L (HRP) (ab205718) at 1:5000 dilution with ECL detection technique.
The ab290 antibody has been shown to work at 1:50,000 and dilutions falling in this range should be tested upon seeing high background. Both incubation steps should be ensured at 22 °C for one hour.

 

Target

  • Subunit Structure - Monomer
  • Tissue Specificity - Photocytes
  • Sequence Similarities - It belongs to the GFP family.
  • Biophysicochemical Properties - Absorption: Abs (maximum) = 395 nm. While the ab290 antibody displays a smaller absorbance peak at 470 nm, the fluorescence emission spectrum peaks at 509 nm with a shoulder at 540 nm.

Relevance

The ab290 antibody functions as an energy-transfer acceptor. It ensures that the blue chemiluminescence of the protein aequorin is transduced into green fluorescent light through energy transfer. The antibody fluoresces in vivo on receiving energy from the Ca2+-activated photoprotein aequorin.