Overview

  • Product name
    Anti-GFP antibody (FITC)
    See all GFP primary antibodies
  • Description
    Goat polyclonal to GFP (FITC)
  • Host species
    Goat
  • Conjugation
    FITC. Ex: 493nm, Em: 528nm
  • Tested applications
    Suitable for: IHC-FoFr, IHC-Fr, WB, ICC/IFmore details
  • Immunogen

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

    MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTT GKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFF KDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNV YIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHY LSTQSALSKDPNEKRDHMVLLEFVTAAGITHGMDELYK


    Database link: P42212

  • Positive control
    • WB: Recombinant A. victoria GFP protein.
  • General notes

    Designed to detect GFP and its variants in ELISA (sandwich or capture), immunoblotting and immunoprecipitation. Fluorescein conjugated anti-GFP was assayed by immunofluorescence microscopy on prokaryotic (E.coli) and eukaryotic (CHO cells) expression systems and was shown to detect GFP containing inserts. Significant amplification of signal was detected using fluorochrome conjugated anti-GFP relative to the fluorescence of GFP alone.
    In case of unexpected background, use pre-adsorbed secondary antibodies.

    Fluorescein isothiocyanate (FITC) (MW 390 daltons) Absorption Wavelength: 495 nm Emission Wavelength: 528 nm Fluorochrome/Protein Ratio: 3.5 moles FITC per mole of Goat IgG

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 0.42% Potassium phosphate, 0.87% Sodium chloride, 1% BSA

    BSA Immunoglobulin and Protease free
  • Concentration information loading...
  • Purity
    Protein G purified
  • Purification notes
    GFP Fluorescein Conjugated Antibody 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. Fluorescein conjugated anti-GFP was assayed by immunofluorescence microscopy on prokaryotic (E.coli) and eukaryotic (CHO cells) expression systems and was shown to detect GFP containing inserts. Significant amplification of signal was detected using fluorochrome conjugated anti-GFP relative to the fluorescence of GFP alone.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab6662 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
IHC-FoFr 1/250.
IHC-Fr Use at an assay dependent concentration.
WB 1/10000 - 1/100000.
ICC/IF 1/200 - 1/400.
IF 1/500 - 1/2500.

Target

  • 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

Images

  • Anti-GFP antibody (FITC) (ab6662) + Recombinant A. victoria GFP protein (ab84191)

    Secondary
    Fluorescein goat secondary antibody for 60 minutes at RT at 1/1000 dilution


    Block: Blocking buffer for 30 minutes at RT.

  • Oscillation of Cyclin E and E2F target gene expression is deregulated in de2f1b mutant salivary glands.

    Salivary glands of control and de2f1b mutant early (80–85 hr AEL) third instar larvae expressing PCNA-GFP (green, ab6662) are stained with anti-dE2F1 (red). The region where high PCNA-GFP is observed with low dE2F1 is marked by an asterisk.

    For Immunostaining, third instar imaginal discs and salivary glands were dissected in PBS and immediately fixed in 4% formaldehyde in PBS for 20 minutes at room temperature with the exception of tissues subjected to anti-dE2F1 staining that were fixed for 30 minutes on ice. Fixed tissues were then washed with 0.3% PBST (0.3% TritonX-100 in 1XPBS) and 0.1% PBST (0.1% TritonX-100 in 1XPBS). Samples were incubated with appropriate amount of primary antibody in 0.1% PBST and 1%BSA overnight. Samples were then washed with 0.1% PBST, incubated in secondary antibody in 0.1% PBST and 1% BSA for 2 hours, followed by several washes in 0.1% PBST prior to mounting.

  • Immunofluorescence Microscopy using ab6662.

    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 minutes 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.

  • ab6662 staining mouse brain tissue sections (inducible GFP reporter) by IHC-Fr.

    The tissue was paraformaldehyde fixed and blocked with serum and then incubated with the antibody at a 1/1000 dilution for 1 hour.

    Staining is shown in the left hand panel. The middle panel shows staining with a rabbit anti-GFP antibody and the right hand panel shows the merged images (plus DAPI).

    ab6662 gives no noticable background and it is found that when viewing on an epifluorescent the exposure time is significantly reduced.

    See Abreview

  • Immunofluorescence Microscopy using ab6662.

    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.

    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 1:500 at 1/10,000 for 45 minutes 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.

  • These pictures show confocal immunofluorescence using GFP-expressing glial cells (green) transplanted into the lesioned rat spinal cord.

    Detected using ab6662 and a standard FITC filter set. Axons are labeled red by an antibody to neurofilament-200 and a rhodamine secondary antibody. The upper panel shows the centre of the transplant site at low power. Numerous GFP-positive cells can be seen mingling with axons. The lower panel shows, at high power in a single optical section, how ab6662 reveals the morphology of the transplanted cells to such an extent that their close interactions with axons are obvious - the cell depicted can be seen wrapping around a neurofilament-200 positive axon.

    These images were kindly supplied as part of the review submitted by Andrew Toft.

References

This product has been referenced in:
  • Li Y  et al. Genetic targeting of Purkinje fibres by Sema3a-CreERT2. Sci Rep 8:2382 (2018). IHC-FoFr . Read more (PubMed: 29403069) »
  • Kim M  et al. An alternatively spliced form affecting the Marked Box domain of Drosophila E2F1 is required for proper cell cycle regulation. PLoS Genet 14:e1007204 (2018). Read more (PubMed: 29420631) »
See all 83 Publications for this product

Customer reviews and Q&As

1-10 of 15 Abreviews or Q&A

Application
Immunohistochemistry free floating
Sample
Mouse Tissue sections (brain)
Specification
brain

James Bogenpohl

Verified customer

Submitted Jul 16 2018

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Intestine)
Permeabilization
No
Specification
Intestine
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Dec 18 2012

Answer

Ab6662 n’a effectivement pas encore été testé en cytométrie en flux et nous n’offrons pas d’échantillon gratuit à des fins de tests préliminaires. Cependant, si vous souhaitez tester cet anticorps en cytométrie en flux, il me sera possible de vous offrir une remise. Après soumission d’une Abreview avec vos résultats, un avoir de la valeur de l'anticorps testé vous sera offert.

Étapes à suivre :

1. Nous informer que vous souhaitez tester l’anticorps en cytométrie en flux et nous vous enverrons un code promotionnel (valable 4 mois) à utiliser après obtention et envoi de vos résultats,

2. Acheter l'anticorps,

3. Nous envoyer vos résultats (positifs ou négatifs) obtenus sous forme d'Abreview (www.abcam.com/abreviews).

4. L'envoi de vos résultats activera le code et vous donnera un avoir de la valeur de l'anticorps testé.

J'espère que ces informations vous aideront. N'hésitez pas à me contacter si vous avez besoin de plus de renseignements.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Intestine)
Specification
Intestine
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Apr 17 2012

Answer

ab6662 is directly conjugated to the FITC molecule, thus there is no secondary antibody present. The FITC molecule is attached directly to the Fc portion of the antibody via covalent bonds to amine groups. The FITC is not irreversibly attached to the antibody, however, because the bonds formed are covalent strong chemicals would be required which could also damage the antibody. Lastly, there is no unconjugated FITC in the vial; the only FITC present will be conjugated to the antibody.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
African Green Monkey Cell (COS-7)
Specification
COS-7
Fixative
Paraformaldehyde
Permeabilization
Yes - Methanol, 30 min at -20°C
Blocking step
FCS as blocking agent for 30 minute(s) · Concentration: 20% · Temperature: RT°C

Dr. Ioannis Gavvovidis

Verified customer

Submitted Dec 09 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Mouse Tissue sections (brain)
Specification
brain
Fixative
Paraformaldehyde
Antigen retrieval step
None
Permeabilization
No
Blocking step
Commercially available blocking buffer as blocking agent for 30 minute(s) · Concentration: 0.5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jan 07 2008

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Brain and spinal cord)
Specification
Brain and spinal cord
Fixative
Methanol
Permeabilization
Yes - 0.1% Triton X100
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2.5% · Temperature: 25°C

Dr. Joshua Weiner

Verified customer

Submitted Jul 06 2007

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Inducible GFP reporter (Brain))
Specification
Inducible GFP reporter (Brain)
Fixative
Paraformaldehyde
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 30 ul/ml

Dr. Joshua Breunig

Verified customer

Submitted Feb 09 2007

Answer

This GFP antibody will cross react with YFP, CFP, eGFP, but not RFP.

Read More

1-10 of 15 Abreviews or Q&A

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