GFP Antibody [Biotin]

Lane 1: HeLa cells. Lane 2: mock transfected HeLa cell lysate. Load: 35 ug per lane. Primary antibody: GFP antibody at 1 ug/ml for 1 h at room temperature. Secondary antibody: IRDye 800 conjugated Donkey-a-Goat IgG [H&L] MX7 secondary antibody at 1:2,500 for 45 min at RT. Block: 5% BLOTTO overnight at 4C. Predicted/Observed size: 27 kDa, 33 kDa for GFP. Other band(s): none.

GFP-Biotin was tested in GFP-Hela cells against a Streptavidin conjugated 550 secondary. Hela-GFP are illuminated in Green and nuclei in Blue (DAPI).

Lane 1: GFP. Lane 2: None. Load: 50 ng per lane. Primary antibody: GFP antibody Biotin conjugated at 1:1,000 for 60 min at RT. Secondary antibody: Peroxidase streptavidin secondary antibody at 1:40,000 for 30 min at RT. Blocking: incubated with blocking buffer for 30 min at RT. Predicted/Observed size: 28 kDa, 28 kDa for GFP. Other band(s): None.

IF analysis of GFP in mouse aorta, liver, brain, and lung cells. Image courtesy of product review by Laura Shankman.

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 40um. Primary antibody: Goat anti-GFP was used at 1:500 dilution in free floating IHC 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 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.

Staining of transgenic mouse pancreas, expressing GFP in beta cells. Image courtesy of product review by Dr. Yves Heremans of Vrije Universiteit Brussel.

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.

ELISA Results of GFP antibody [Biotin] tested against purified GFP protein. Each well was coated in duplicate with 10ug/mL of GFP protein. The starting dilution of antibody was 5ug/mL and the X-axis represents the Log10 of a 3-fold dilution. This titration is a 4-parameter curve fit where the IC50 is defined as the titer of the antibody. Assay performed using 3% Fish Gel/PBS Blocking buffer and SA-HRP conjugated.

• Reactivity: All-NA
• Applications: WB, ELISA, ICC/IF, IHC, KD, KO
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Biotin

GFP Antibody [Biotin] Summary

• Description: For extended storage aliquot contents and freeze at -20C or below. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room
  
  This 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. Assay by immunoelectrophoresis resulted in a single precipitin arc against anti-Goat Serum, anti-biotin and purified and partially purified Green Fluorescent Protein (Aequorea victoria) Serum
• Immunogen: The immunogen is a Green Fluorescent Protein (GFP) fusion protein corresponding to the full length amino acid sequence (246aa) derived from the jellyfish Aequorea victoria. (Uniprot: P42212)
• Specificity: No reaction was observed against Human, Mouse and Rat Serum Proteins.
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Purity: Immunogen affinity purified

Applications/Dilutions

• Dilutions:
  • ELISA 1:50000-1:80000
  • Immunocytochemistry/ Immunofluorescence 1:100-1:1000
  • Immunohistochemistry 1:1000-1:5000
  • Immunohistochemistry-Frozen 1:10-1:500
  • Immunohistochemistry-Paraffin 1:10-1:500
  • Knockdown Validated
  • Knockout Validated
  • Western Blot 1:2000-1:10000
• Application Notes: GFP Antibody is suitable for immunoblotting, ELISA, immunohistochemistry, immunomicroscopy as well as other antibody based assays using streptavidin or avidin conjugates requiring lot-to-lot consistency.
• Readout System:
  • Streptavidin Native Protein NBP2-29367
  • Streptavidin Native Protein NBP2-29370
  • Streptavidin Native Protein NBP2-29371

Reactivity Notes

No reaction was observed against Human, or Rat serum proteins. Known Cross Reactivity: rGFP. YFP differs from GFP due to a mutation at Thr203Tyr; antibodies raised against full-length GFP should also detect YFP and other variants. Reactivity in transgenic mice with GFP. Reactivity in human cell lines transfected will a GFP construct.

Packaging, Storage & Formulations

• Storage: Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.
• Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2, 10 mg/mL Bovine Serum Albumin (BSA) - Immunoglobulin and Protease free
• Preservative: 0.01% Sodium Azide
• Purity: Immunogen affinity purified

Alternate Names for GFP Antibody [Biotin]

• eGFP
• Enhanced Green Fluorescent Protein
• GFP
• GFPuv
• green fluorescent protein (gfp)
• Green Fluorescent Protein

Background

Green fluorescent protein (GFP) is a 27 kDa protein derived from the bioluminescent jellyfish Aquorea Victoria. GFP can be excited by light in the blue to ultraviolet spectrum and has an excitation peak at 395 nm with an emission peak at 509 nm in the green spectrum. GFP autocatalytically forms a fluorescent pigment from natural amino acids present in the nascent protein. GFP is widely used as a reporter (tag) for gene expression, enabling researchers to visualize and localize GFP-tagged proteins within living cells without the need for chemical staining. GFP fluorescence is stable under fixation conditions and antibodies are suitable for a variety of applications including ELISA, ICC/IF, IHC, IP and WB (1-3).

References
1. Shi, C., Pan, F. C., Kim, J. N., Washington, M. K., Padmanabhan, C., Meyer, C. T., . . . Means, A. L. (2019). Differential Cell Susceptibilities to Kras(G12D) in the Setting of Obstructive Chronic Pancreatitis. Cell Mol Gastroenterol Hepatol. doi:10.1016/j.jcmgh.2019.07.001

2. Zhao, S., Fortier, T. M., & Baehrecke, E. H. (2018). Autophagy Promotes Tumor-like Stem Cell Niche Occupancy. Curr Biol, 28(19), 3056-3064.e3053. doi:10.1016/j.cub.2018.07.075

3. Zusso, M., Lunardi, V., Franceschini, D., Pagetta, A., Lo, R., Stifani, S., . . . Moro, S. (2019). Ciprofloxacin and levofloxacin attenuate microglia inflammatory response via TLR4/NF-kB pathway. J Neuroinflammation, 16(1), 148. doi:10.1186/s12974-019-1538-9

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for GFP Antibody (NB100-1678)(11)

Publications using NB100-1678

• Kim K, Wann J, Kim HG et al. Uncoupling protein 1-driven Cre ( Ucp1-Cre ) is expressed in the epithelial cells of mammary glands and various non-adipose tissues bioRxiv : the preprint server for biology 2023-10-22 [PMID: 37905088] (Immunohistochemistry)
• Abe I, Oguri Y, Verkerke ARP et al. Lipolysis-derived linoleic acid drives beige fat progenitor cell proliferation Developmental cell 2022-12-05 [PMID: 36473459] (IHC-P)
  
  Details:
  Dilution used in IHC-P 1:200
• Li Y, Imai N, Nicholls HT et al. Thioesterase superfamily member 1 undergoes stimulus-coupled conformational reorganization to regulate metabolism in mice Nat Commun 2021-06-10 [PMID: 34108467] (ICC/IF)
  
  Details:
  This citation used the Biotin format of this antibody.
• Li J, Li E, Czepielewski RS et al. Neurotensin is an anti-thermogenic peptide produced by lymphatic endothelial cells Cell Metab 2021-05-26 [PMID: 34038712]
  
  Details:
  Citation using the Biotin format of this antibody.
• Sethi NS, Kikuchi O, Duronio GN et al. Early TP53 alterations engage environmental exposures to promote gastric premalignancy in an integrative mouse model Nat Genet 2020-02-07 [PMID: 32025000] (IHC-P)
• Sonzogni O, Haynes J, Seifried LA et al. Reporters to mark and eliminate basal or luminal epithelial cells in culture and in vivo PLoS Biol 2018-06-21 [PMID: 29924804] (ICC/IF)
• Roh HC, Tsai LTY, Shao M et al. Warming Induces Significant Reprogramming of Beige, but Not Brown, Adipocyte Cellular Identity Cell Metab 2018-04-17 [PMID: 29657031]
• Relation T, Yi T, Guess AJ et al. Intratumoral Delivery of Interferongamma-Secreting Mesenchymal Stromal Cells Repolarizes Tumor-Associated Macrophages and Suppresses Neuroblastoma Proliferation In Vivo Stem Cells 2018-02-13 [PMID: 29430789] (ICC/IF)
• Relation T, Yi T, Guess AJ et al. Mule Regulates the Intestinal Stem Cell Niche via the Wnt Pathway and Targets EphB3 for Proteasomal and Lysosomal Degradation Cell Stem Cell 2016-05-18 [PMID: 27184401] (IF/IHC)
• Edelblute CM, Heller LC, Malik MA, Heller R. Activated air produced by shielded sliding discharge plasma mediates plasmid DNA delivery to mammalian cells Biotechnol Bioeng 2015-06-03 [PMID: 26041378]
  
  Details:
  This citation used the Biotin version of this antibody.
• Long JZ, Svensson KJ, Tsai L et al. A Smooth Muscle-Like Origin for Beige Adipocytes Cell Metab 2014-04-02 [PMID: 24709624] (IHC-P)
  
  Details:
  This citation used the Biotin version of this antibody.

Reviews for GFP Antibody (NB100-1678) (3)

Average Rating: 4.7

(Based on 3 reviews)

We have 3 reviews tested in 1 species: Mouse.

reviewed by: Verified Customer IHC-P Mouse 06/27/2013

Summary

• Application: Immunohistochemistry-Paraffin
• Sample Tested: Transgenic mouse pancreas, expressing GFP in beta cells
• Species: Mouse

reviewed by: Verified Customer ICC Mouse 11/08/2011

Summary

• Application: Immunocytochemistry
• Sample Tested: Mouse Aorta, Liver, Brain, Lung
• Species: Mouse
• Lot: 23463

reviewed by: Verified Customer IHC-Fr Mouse 02/17/2010

Summary

• Application: Immunohistochemistry-Frozen
• Sample Tested: Mouse Tissue-Brain
• Species: Mouse
• Lot: 23000

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