• Product name
    Anti-GFP antibody (Biotin)
    See all GFP primary antibodies
  • Description
    Goat polyclonal to GFP (Biotin)
  • Host species
  • Conjugation
  • Specificity
    Antibody recognizes wild type, recombinant and enhanced forms of GFP (EGFP). No reaction was observed against Human, Mouse and Rat Serum Proteins.
  • Tested applications
    Suitable for: WB, IP, ICC/IF, Sandwich ELISA, IHC-P, In situ hybridization, IHC-Frmore details
  • Species reactivity
    Reacts with: Species independent
  • Immunogen

    Recombinant full length protein corresponding to GFP aa 1-246.


    Database link: P42212

  • General notes

    Designed to detect GFP and its variants in ELISA (sandwich or capture), immunoblotting and immunoprecipitation.

    Biotinamidocaproate N-Hydroxysuccinimide Ester (BAC) Biotin/Protein Ratio: 10-20 BAC molecules per goat IgG molecule.


  • Form
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 0.44% Potassium phosphate, 0.88% Sodium chloride

    10 mg/mL BSA, immunoglobulin and protease free
  • Concentration information loading...
  • Purity
    Affinity purified
  • Purification notes
    This product was prepared from monospecific antiserum by immunoaffinity chromatography using Green Fluorescent Protein (Aequorea victoria) coupled to agarose beads followed by solid phase adsorption(s) to remove any unwanted reactivities.
  • Primary antibody notes
    Designed to detect GFP and its variants in ELISA (sandwich or capture), immunoblotting and immunoprecipitation.
  • Clonality
  • Isotype
  • Research areas


Our Abpromise guarantee covers the use of ab6658 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 1/2000 - 1/10000.
IP Use at an assay dependent concentration.
ICC/IF 1/1000 - 1/5000.
Sandwich ELISA 1/10000 - 1/50000. Can be paired for Sandwich ELISA with Mouse monoclonal [9F9.F9] to GFP (ab1218).

May be used as capture or detection antibody in a Sandwich ELISA.

IHC-P 1/250.
In situ hybridization Use at an assay dependent concentration. PubMed: 24262147
IHC-Fr 1/5000.


  • Relevance
    Function: 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


  • Anti-GFP antibody (Biotin) (ab6658)
    Additional bands at: 28 kDa. We are unsure as to the identity of these extra bands.

  • ab6658 staining GFP in human melanoma cells recovered from nude mice by Immunocytochemistry/ immunoflurescence. Cells were fixed with formaldehyde, permeabilized with 0.25% Triton X-100 RT for 10min and blocking with commercially available blocking buffer was performed for 30 minutes at RT. Samples were incubated with primary antibody (1:50) for 18 hours at 4°C. An Alexa Fluor®488-conjugated donkey polyclonal to goat IgG was used as secondary antibody at 1/100 dilution. Green color indicates GFP/Fibrolast cells, while red color indicates Ki67 positive cells, most of them are tumor cells (Abcam`s ab15580 was used for the detection).

    See Abreview

  • Immunofluorescence Microscopy using ab6658. Tissue: Drosophila melanogaster late stage embryonic central nervous system. Fixation: 0.5% PFA. Antigen retrieval: not required. Primary antibody: Anti-GFP antibody at a 1/1,000 for 1 h at RT. Secondary antibody: AlexaFluor 488™ conjugated anti-Goat antibody at 1/300 for 45 min at RT. Panel A: shows a lateral view (ventral left). Panels B and C: shows ventral views of whole mount embryos at 63x magnification (plus 2x digital zoom). In all panels, anterior is up. Staining: tau-GFP cell bodies (large arrowhead) and axons of motorneurons (arrow) and interneurons (small arrowhead) as green fluorescent signal.

  • Immunofluorescence Microscopy using ab6658. Tissue: Sf-1:Cre mice crossed to the Z/EG reporter line. Mouse brain (coronal view, 20X magnification). Fixation: 4%PFA/PBS with o/n fixation, and subsequently transferred to a 30% sucrose solution. Antigen retrieval: frozen in OCT freezing medium (Sakura) and cryostat sectioned at 40 microns. Primary antibody: Goat anti- GFP was used at 1/500 dilution in free floating imunnohistochemistry to detect GFP. Secondary antibody: Fluorchrome conjugated Anti-goat IgG secondary antibody was used for detection at 1:10,000 for 45 min at RT.Localization: Sf-1+ neurons and their processes of the ventromedial nucleus of the hypothalamus. Staining: eGFP as green fluorescent signal and sections were counterstained with DAPI.

  • ab6658 staining GFP in rat brain cells infected with viruses containing GFP under a CMV promoter by Immunocytochemistry/ Immunofluorescence.Cells were fixed with formaldehyde, permeabilized using 0.2% Triton, blocked with 20% serum and then incubated with ab6658 at a 1:50 dilution for 20 hours at 25°C. The secondary used was an Alexa-Fluor 488 conjugated rabbit polyclonal, used at a 1/200 dilution.

    See Abreview


This product has been referenced in:
  • Muralidharan B  et al. An Efficient System for Gene Perturbation in Embryonic Hippocampal Progenitors Using Ex Vivo Electroporation Followed by In Vitro Dissociated Cell Culture. J Exp Neurosci 12:1179069518767404 (2018). Read more (PubMed: 29760561) »
  • He S  et al. Adult zebrafish Langerhans cells arise from hematopoietic stem/progenitor cells. Elife 7:N/A (2018). Read more (PubMed: 29905527) »
See all 52 Publications for this product

Customer reviews and Q&As

1-10 of 18 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Mouse Cell lysate - whole cell (E. coli)
Total protein in input
4000 µg
Immuno-precipitation step
Other - Dynabeads
E. coli

Abcam user community

Verified customer

Submitted Feb 26 2018

Abcam guarantees this product to work in the species/application used in this Abreview.
Western blot
Mouse Cell lysate - whole cell (Fibroblast)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
30 µg
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Hongwei Shao

Verified customer

Submitted Sep 06 2017


I am sorry to confirm that the binding constant is not often determined for research grade antibodies and there is no binding constant information available for this particular product.

I am sorry we have no data to share to help answer your question on this occasion.

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I performed the following molecular weight calculations for this antibody:

Assuming 15 biotin molecules per IgG molecule:

MW IgG = 150 kDa
MW biotin = 244.31 g/mol = 244.31 Da = 0.24431 kDa
MW ab6658 = 150 + (15*0.24431) ˜ 154 kDa

So the molecular weight of the biotinylated antibody should be ˜ 154 kDa.

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We do not determine the percentage of molecules conjugated to biotin, but it is expected to be close to 100%. The conjugation ratio between our biotin and IgG is usually 5-6 molecules of biotin per molecule of IgG.

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The unconjugated version of ab6658 is ab6673.


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Thank you very much for contacting us with your question.

In general, using a secondary antibody will help amplify the signal from the primary antibody. With a pre-conjugated primary antibody, the signal may be weaker than using a conjugated secondary antibody. The avidin-biotin complex will also help amplify the signal, and there are other benefits to using a pre-conjugated primary (no cross-reaction between the secondary and samples, faster assay, etc), so all of these points should be weighed when considering whether to use a pre-conjugated primary or a secondary antibody step.

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>pecies specificity is not applicable for this product. You can try this antibody for detection of GFP in any type of cells if they express GFP protein.

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Abcam guarantees this product to work in the species/application used in this Abreview.
Immunocytochemistry/ Immunofluorescence
Rat Cell (Brain tissue slices)
Yes - Triton 0.2%
Brain tissue slices
Blocking step
Serum as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 20% · Temperature: 25°C

Dr. Efrat Shema

Verified customer

Submitted Aug 04 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Rat Tissue sections (Brain)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Sodium citrate pH 6.0
Yes - Triton 0.2%
Blocking step
Serum as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 20%

Dr. Efrat Shema

Verified customer

Submitted Aug 02 2011

1-10 of 18 Abreviews or Q&A


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