Green Fluorescent Protein (GFP) antibody

Details for Product No. ABIN398304
Independently validated
Antigen
Reactivity
Aequorea victoria
(337), (31), (11), (8), (4), (2), (2), (1), (1), (1)
Host
Rat
(152), (149), (54), (26), (3), (2)
Clonality (Clone)
Monoclonal ()
Conjugate
Un-conjugated
(25), (19), (14), (9), (9), (3), (3), (3), (3), (3), (3), (3), (3), (2), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1)
Application
Western Blotting (WB), ELISA
(300), (155), (105), (67), (66), (38), (37), (31), (30), (29), (13), (11), (10), (10), (8), (4), (3), (2), (1)
Pubmed 10 references available
Quantity 100 μL
Options
Shipping to United States (Change)
Availability Will be delivered in 4 to 5 Business Days
'Independent Validation' Badge
Lot Number 100316
Method validated Western Blotting
Positive Control 293FT cells transduced with eGFP virus
Negative Control untransduced 293FT cells
Notes A major band was observed at the correct molecular weight in the two positive (eGFP-transduced) cell lysates. No major bands were observed in the negative control (untransduced cells). Some additional faint bands of lower molecular weight were also observed in the positive controls, this may be because the antibody was used at 1:500 instead of the recommended 1:1000 dilution. Microscope images of transduced cells are provided to demonstrate successful eGFP-transduction of cells.
Catalog No. ABIN398304
244.20 $
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Purpose For biochemical analysis of GFP-tagged fusion proteins.
Clone 3H9
Isotype IgG2a
Specificity 3H9 efficiently recognizes green fluorescent proteins from Aequorea victoria such as eGFP, wtGFP, YFP, or CFP, but no red fluorescent proteins.
Characteristics Fluorescent proteins (FP) are powerful tools to study protein localization and dynamics in living cells in particular the prominent green fluorescent protein (GFP) from Aequorea victoria.
Here we offer a rat monoclonal antibody (MAb) against various GFPs, designated as 3H9. The 3H9 MAb is characterized by high affinity and specificity. It is suitable for immunoblotting and ELISA.
With this versatile antibody GFP and fusions thereof become attractive tools for biochemical applications like identification of interacting proteins or Western Blotting.
Purification Hybridoma supernatant. The Hybridoma was cultured in protein free medium. Tissue culture supernatant was concentrated by centrifugation using a 30 kDa exclusion membrane to an antibody concentration of 1 mg/ml.
Background The first green fluorescent protein (GFP) was isolated from Aequorea victoria and has a molecular weight of 27 kDa. Genetic mutations of GFP resulted in fluorescent proteins with different excitation and emission spectra e.g. eGFP, YFP, CFP. Fluorescent proteins became attractive tools for a variety of applications.
Research Area Tags/Labels
Application Notes Western Blot: Used at a dilution of 1/1000. Detects a band of approximately 31 kDa (the predicted molecular weight of GFP).
Comment

IF: not working

Restrictions For Research Use only
Format Liquid
Concentration 1 mg/mL
Preservative Sodium azide
Precaution of Use This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Handling Advice Do not freeze.
Storage 4 °C
Expiry Date 12 months
Supplier Images
anti-Green Fluorescent Protein (GFP) antibody Western Blot analysis using anti-GFP antibody 3H9. Primary antibody: 0.2 ug of GFP antibody 3H9 incubation for 1 h at RT. Secondary antibody: anti-rat-HRP (Jackson ImmunoResearch Europe). Detection with ECL Western Blot Detection Kit
Product cited in: Sieber, Lange, Kollmorgen et al.: "Sharpin contributes to TNFα dependent NFκB activation and anti-apoptotic signalling in hepatocytes." in: PLoS ONE, Vol. 7, Issue 1, pp. e29993, 2012 (PubMed).

Ingmundson, Nahar, Brinkmann et al.: "The exported Plasmodium berghei protein IBIS1 delineates membranous structures in infected red blood cells." in: Molecular microbiology, Vol. 83, Issue 6, pp. 1229-43, 2012 (PubMed).

Garcia-Santisteban, Zorroza, Rodriguez: "Two nuclear localization signals in USP1 mediate nuclear import of the USP1/UAF1 complex." in: PLoS ONE, Vol. 7, Issue 6, pp. e38570, 2012 (PubMed).

Bouasker, Simard: "The slicing activity of miRNA-specific Argonautes is essential for the miRNA pathway in C. elegans." in: Nucleic acids research, Vol. 40, Issue 20, pp. 10452-62, 2012 (PubMed).

Lee, Fischer: "Drosophila Tel2 is expressed as a translational fusion with EpsinR and is a regulator of wingless signaling." in: PLoS ONE, Vol. 7, Issue 9, pp. e46357, 2012 (PubMed).

Becker, Allmann, Hofstätter et al.: "Direct homo- and hetero-interactions of MeCP2 and MBD2." in: PLoS ONE, Vol. 8, Issue 1, pp. e53730, 2013 (PubMed).

Lestini, Laptenok, Kühn et al.: "Intracellular dynamics of archaeal FANCM homologue Hef in response to halted DNA replication." in: Nucleic acids research, Vol. 41, Issue 22, pp. 10358-70, 2013 (PubMed).

Kríz, Pospíchalová, Masek et al.: "β-arrestin promotes Wnt-induced low density lipoprotein receptor-related protein 6 (Lrp6) phosphorylation via increased membrane recruitment of Amer1 protein." in: The Journal of biological chemistry, Vol. 289, Issue 2, pp. 1128-41, 2014 (PubMed).

Anton, Bauer, Keck et al.: "A p38 substrate-specific MK2-EGFP translocation assay for identification and validation of new p38 inhibitors in living cells: a comprising alternative for acquisition of cellular p38 inhibition data." in: PLoS ONE, Vol. 9, Issue 4, pp. e95641, 2014 (PubMed).

Galili, Dylla, Lüdke et al.: "Converging circuits mediate temperature and shock aversive olfactory conditioning in Drosophila." in: Current biology : CB, Vol. 24, Issue 15, pp. 1712-22, 2014 (PubMed).

Primary Antibody
  • Antigen: Green Fluorescent Protein (GFP)
  • Catalog number: ABIN398304
  • Supplier: Chromotek
  • Supplier catalog number: 3H9
  • Lot number: 100316
Secondary Antibody
  • Antibody: Alexa Fluor 680 Goat Anti-Rat IgG (H+L)
  • Supplier: Life Technologies
  • Catalog number: A-21096
  • Lot number: N/A
Controls
  • Positive control: 293FT cells transduced with eGFP virus
  • Negative control: untransduced 293FT cells
Protocol
  • 20 µg of total protein from prepared cell lysates was run on a 4-12% SDS-PAGE gel.
  • The gel was run at 180 V for 40 minutes and transferred to a PVDF membrane using a wet-transfer apparatus.
  • The membrane was rinsed in TBST for 10 min in TBST.
  • The membrane was blocked in 3% BSA for 30 min.
  • The membrane was incubated in 1:500 primary antibody diluted in 3% BSA at 4°C overnight.
  • The membrane was washed 3 times for 10 min in TBST.
  • The membrane was incubated in secondary antibody at 1:10000 for 1 hr at RT.
  • The membrane was washed 3 times for 10 min in TBST.
  • The membrane was imaged using a fluorescent imaging system at 679nm⁄702nm.
Experimental Notes Fluorescent microscopy images are included to confirm eGFP transduction of 293FT cells.
Validation Images
anti-Green Fluorescent Protein (GFP) antibody Figure 1: A. Western blot of lysates from untransduced 293FT cells (Lane 2) and 293FT cells transduced with eGFP virus (Lane 3 and 4) probed with anti-GFP antibody (upper panel) or for alpha tubulin (lower panel). B. Confocal image of 293FT cells used to generate Lysate1 and Lysate2.
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