Overview

  • Product nameAnti-GFP antibody [9F9.F9]
    See all GFP primary antibodies
  • Description
    Mouse monoclonal [9F9.F9] to GFP
  • SpecificityAssayed by ELISA for direct binding of antigen recognizes wild type and recombinant GFP. Well suited to titrate GFP in solution using either form of the antibody as the capture or detection antibodies. Shows no reactivity against red fluorescence protein (RFP). This antibody is known to cross react with the wild type (wt), recombinant (rGFP) and enhanced (eGFP) forms.
  • Tested applicationsSuitable for: WB, IHC-Fr, Sandwich ELISA, ICC/IF, IPmore details
  • Immunogen

    Fusion protein corresponding to GFP aa 1-246. Full Length Fusion Protein. Derived from the jellyfish Aequorea victoria.

Properties

  • FormLiquid
  • Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage bufferpH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 0.87% Sodium chloride, 0.42% Potassium phosphate
  • Concentration information loading...
  • PurityProtein A purified
  • Purification notesGFP Monoclonal Antibody was prepared from tissue culture supernatant by Protein A affinity chromatography.
  • ClonalityMonoclonal
  • Clone number9F9.F9
  • IsotypeIgG1
  • Light chain typekappa
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab1218 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/3000 - 1/30000.

(See Abreviews). Recognises the immunogen under native conditions and reduced/denatured conditions.

IHC-Fr 1/1000 - 1/5000.

Works also with eYFP.

Sandwich ELISA 1/80000 - 1/500000. Can be paired for Sandwich ELISA with Goat polyclonal to GFP (Biotin) (ab6658).

ab1218 can be used to detect GFP by ELISA (sandwich or capture) for the direct binding of antigen. Biotin conjugated monoclonal anti-GFP is well suited to titrate GFP in a sandwich ELISA in combination with polyclonal anti-GFP as a capture antibody.

ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration. PubMed: 16135826

Target

  • RelevanceFunction: Energy-transfer acceptor. Its role is to transduce the blue chemiluminescence of the protein aequorin into green fluorescent light by energy transfer. Fluoresces in vivo upon receiving energy from the Ca2+ -activated photoprotein aequorin.

    Subunit structure: Monomer.

    Tissue specificity: Photocytes.

    Post-translational modification: Contains a chromophore consisting of modified amino acid residues. The chromophore is formed by autocatalytic backbone condensation between Ser-65 and Gly-67, and oxidation of Tyr-66 to didehydrotyrosine. Maturation of the chromophore requires nothing other than molecular oxygen.

    Biotechnological use: Green fluorescent protein has been engineered to produce a vast number of variously colored mutants, fusion proteins, and biosensors. Fluorescent proteins and its mutated allelic forms, blue, cyan and yellow have become a useful and ubiquitous tool for making chimeric proteins, where they function as a fluorescent protein tag. Typically they tolerate N- and C-terminal fusion to a broad variety of proteins. They have been expressed in most known cell types and are used as a noninvasive fluorescent marker in living cells and organisms. They enable a wide range of applications where they have functioned as a cell lineage tracer, reporter of gene expression, or as a measure of protein-protein interactions. Can also be used as a molecular thermometer, allowing accurate temperature measurements in fluids. The measurement process relies on the detection of the blinking of GFP using fluorescence correlation spectroscopy.

    Sequence similarities: Belongs to the GFP family.

    Biophysicochemical properties: Absorption: Abs(max)=395 nm
    Exhibits a smaller absorbance peak at 470 nm. The fluorescence emission spectrum peaks at 509 nm with a shoulder at 540 nm.
  • Alternative names
    • GFP antibody
    • Green fluorescent protein antibody
    • yfp antibody

Anti-GFP antibody [9F9.F9] images

  • All lanes : Anti-GFP antibody [9F9.F9] (ab1218) at 1 mg/ml (1 hour at room temperature)

    Lane 1 : HeLa lysate 50ng with BLOTTO, overnight at 4°C
    Lane 2 : HeLa lysate 100ng with BLOTTO, overnight at 4°C
    Lane 3 : HeLa lysate 500ng with BLOTTO, overnight at 4°C

    Blocking peptides at 5 % per lane.

    Secondary
    IRDye® 800 conjugated Goat anti-mouse IgG (H+L), 45 minutes at room temperature at 1/2500 dilution

    Additional bands at : 27 kDa. We are unsure as to the identity of these extra bands.
  • ab1218 staining GFP in Monkey Kidney cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with 1% Triton X-100 and blocked with 3% BSA for 1 hour at 25°C. Samples were incubated with primary antibody (1/500) for 12 hour at 4°C. An undiluted Alexa Fluor® 488-conjugated Goat polyclonal was used as the secondary antibody

    See Abreview

  • ab1218 staining GFP in human prostate epithelial primary cells by Immunocytochemistry/ Immunofluorescence. Cells (ARPE-19 cells transfected with mouse Rab27A-GFP) were fixed with paraformaldehyde, permeabilized with 0.5% Triton x100 and blocking with 5% serum was performed for 20 minutes at 25°C. Samples were incubated with primary antibody (1/500: in 1% goat serum, 0.1%TX100, 1XPBS) for 16 hours at 4°C. An Alexa Fluor®546-conjugated goat polyclonal to mouse IgG was used as secondary antibody at 1/500 dilution. DAPI was used to stain the cell nuclei (blue) and mRab27A-GFP with ab1218 (red).

    See Abreview

  • All lanes : Anti-GFP antibody [9F9.F9] (ab1218) at 1/1000 dilution (2 hours at 25°C)

    Lane 1 : J774 with Milk, 2 hours at 25°C
    Lane 2 : J774+pEGFP-N1 with Milk, 2 hours at 25°C

    Lysates/proteins at 20 µg per lane.
    Blocking peptides at 5 % per lane.

    Secondary
    Goat anti-mouse HRP at 1/10000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Additional bands at : 26 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 1 minute

    Image courtesy of an anonymous Abreview

    See Abreview

  • Immunoprecipitation using ab1218 at 0.01µl/µl HEK (human embryonic kidney) 293 whole cell lysate incubated with protein G matrix for 1 hour at 4°C.

    Total protein input was 3,000,000 cells.

    Lane 1: GFP-RhoA wt

    Lane 2: GFP-Rac1 wt

    Lane 3: GFP-empty

    The western blot antibody was ab290 (undiluted).

    See Abreview

References for Anti-GFP antibody [9F9.F9] (ab1218)

This product has been referenced in:
  • So EC  et al. The Rab-binding Profiles of Bacterial Virulence Factors during Infection. J Biol Chem 291:5832-43 (2016). IP . Read more (PubMed: 26755725) »
  • Ilie A  et al. A Christianson syndrome-linked deletion mutation (?(287)ES(288)) in SLC9A6 disrupts recycling endosomal function and elicits neurodegeneration and cell death. Mol Neurodegener 11:63 (2016). Read more (PubMed: 27590723) »

See all 98 Publications for this product

Product Wall

Application Western blot
Sample Hamster Cell lysate - whole cell (CHO cells overexpressing a GFP fusion protein)
Gel Running Conditions Reduced Denaturing (12%)
Loading amount 40 µg
Specification CHO cells overexpressing a GFP fusion protein
Blocking step Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Mar 25 2016

Application Western blot
Loading amount 100000 cells
Gel Running Conditions Reduced Denaturing
Sample Human Cell lysate - whole cell (HEK293T)
Specification HEK293T
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Dr. Brian Emmer

Verified customer

Submitted Mar 20 2015

We do not have data for reactivity of ab1218 with acGFP but a customer sent a positive review of ab290 for IP and WB of acGFP and acGFP-tagged p27. Here is the URL of the review http://www.abcam.com/GFP-antibody-ChIP-Grade-ab290/reviews/33378 .

Application Western blot
Loading amount 30 µg
Gel Running Conditions Reduced Denaturing
Sample Chinese Hamster Cell lysate - whole cell (CHO cell)
Specification CHO cell
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Username

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Submitted Jan 22 2015

Application Immunocytochemistry/ Immunofluorescence
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Sample Monkey Cell (kidney)
Specification kidney
Permeabilization Yes - triton x-100
Fixative Formaldehyde
Username

Abcam user community

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Submitted Jun 11 2014

Application Western blot
Loading amount 60 µg
Gel Running Conditions Reduced Denaturing
Sample Human Cell lysate - whole cell (HEK293T)
Specification HEK293T
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

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Verified customer

Submitted Mar 17 2014

Abcam has not validated the combination of species/application used in this Abreview.
Application Immunohistochemistry (Frozen sections)
Sample Mouse Tissue sections (E10.5 mouse embryo)
Permeabilization Yes - PBS / 0.5% v/v Triton X-100 (Sigma)
Specification E10.5 mouse embryo
Blocking step NOVOCASTRA protein block as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 25°C
Fixative Paraformaldehyde
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Submitted Feb 25 2014

Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HELA)
Permeabilization Yes - 0.2% Triton100, permeablize after primary incubation
Specification HELA
Fixative Formaldehyde
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Abcam user community

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Submitted Oct 18 2013

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 20 µg
Gel Running Conditions Non-reduced Denaturing (10)
Sample Human Cell lysate - whole cell (HeLa cells)
Specification HeLa cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

Submitted May 28 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Rat Cell (fibroblasts)
Specification fibroblasts
Fixative Formaldehyde
Permeabilization No
Blocking step BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Dec 20 2012

1-10 of 64 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"