Dendra 2 antibody

Catalog No. ABIN361314

Product Details

Key Features:

• Dendra2 Antibody (ABIN361314) independently validated for Immunocytochemistry by Neurology Lab (INSERM).
• Publications cite use for Immunoblotting as well as Immunostaining.
• Dendra2 belongs to a new class of photoactivatable fluorescent proteins (PAFPs). PAFPs allow to photolable and track fusion proteins in vivo. Dendra2 Antibody (ABIN361314) can be used for control purposes in this setting.

Target: Dendra 2

Reactivity: Coral

Host: Rabbit

Clonality: Polyclonal

Conjugate: Un-conjugated

Application: Western Blotting (WB), Flow Cytometry (FACS), Immunocytochemistry (ICC)

Purpose: Anti-Dendra2 purified

Specificity: The rabbit polyclonal antibody PAb (836) recognizes Dendra2 tag, a green-to red photoswitchable protein derived from an octocoral (Dendronephthya).

Purification: Purified by protein-A affinity chromatography.

Purity: > 95 % (by SDS-PAGE)

Immunogen: Dendra2 tag

Isotype: IgG

Application Details

Application Notes: Western blotting: Recommended dilution: 1-2 μg/mL.
Flow cytometry: Recommended dilution: 1-4 μg/mL. Extracellular and intracellular staining.

Restrictions: For Research Use only

Independent Validation (ICC)

Validation #101498 (Immunocytochemistry)

by: Plasticity in cortical networks and epilepsy, Institut du Fer à Moulin, UMR839 INSERM/UPMC

No.: #101498

Date: 11/03/2017

Antigen: Dendra2

Lot Number: R-053-AI-03

Method validated: Immunocytochemistry

Positive Control: Rat hippocampal cells transfected with a Dendra2-tagged protein (Kv2.1_Dendra)

Negative Control: Non transfected cells in the same preparation

Notes:

ABIN361314 specifically labels Dendra+ cells in hippocampal cell cultures transfected with a construct bearing Dendra2.

Validation Images

Hippocampal primary culture co-transfected with Kv2.1_Dendra and HomerDsRed (A, red fluorescence of Homer), stained with anti-Dendra2 ABIN361314 (B, revealed with A647-coupled secondary antibody) and anti-MAP2 antibody (C, revealed with A488-coupled secondary antibody). Merged images in (D), showing the labeling of anti-Dendra2 antibody (blue) that reveals the transfected cell identified by the fluorescence of HomerDsRed (red) but not the non-transfected cells (visible by the immunolabeling of MAP2, green). Scale bar: 10µm, 63x magnification.

Full Methods

Primary Antibody: ABIN361314

Secondary Antibody: donkey anti-rabbit A647 conjugated antibody (Jackson Immunoresearch, 711-605-152, lot 123104)

Full Protocol:

• Grow hippocampal cells in Neurobasal medium 1x (Gibco, ref. 21103-049, lot 1880280) supplemented with B27 1x (Gibco, ref. 17-504-044, lot 1733317), 2mM L-glutamine (Gibco, ref. 25030-024, lot 1881896) and antibiotics (200U/ml penicillin, 200µg/ml streptomycin, Gibco, ref. 15140-122), at 37°C and 5% CO₂ at a density of 120×10³ cells/ml on 18-mm diameter glass coverslips (Assistent) pre-coated with 50µg/ml poly-D,L-ornithine (Sigma-Aldrich, P3655, lot SLBL9424V).
• Transfect cells with 0.5µg of Kv2.1_Dendra and 0.5µg of Homer_DsRed constructs (in-house made) using Transfectin Lipid transfection reagent (BioRad, ref 170-3351, lot 4206491) following the manufacturer´s instructions.
• Fix hippocampal cell cultures on coverslips with 4% buffered formalin for 15min at RT.
• Wash slides 2x in PBS (prepared from DPBS 10x, Gibco, 14200-067, lot 1754645).
• Permeabilize cells with 0.25% Triton X-100 (Roth, 3051.3) in PBS for 4min at RT.
• Wash slides 2x in PBS.
• Block non-specific binding with PBS-Goat serum 10% (Goat serum: Gibco, 16210-064, lot 1158993) for 30min at RT.
• Incubate slides with primary rabbit anti-Dendra2 antibody (antibodies-online, ABIN361314, lot R-053-AI-03) diluted 1:100 and mouse anti-MAP2 antibody (Chemicon, MAB 378, lot LV1414361) diluted 1:200 in PBS-Goat serum 10% for 1h at RT.
• Wash slides 2x 5min in PBS-Goat serum 5%.
• Incubate with secondary donkey anti-rabbit A647 conjugated antibody (Jackson Immunoresearch, 711-605-152, lot 123104) and donkey anti-mouse A488 conjugated antibody (Jackson Immunoresearch, 715-545-150, lot 123589) diluted 1:250 in PBS-Goat serum 10% for 45min at RT.
• Wash slides 1x 5min in PBS.
• Mount coverslips with 10-20µl of Mowiol antifade reagent (Calbiochem, 475904, lot D00136768).
• Image acquisition on a Nikon Eclipse Ti microscope, 63x magnification.

Experimental Notes:

Staining pattern of Kv2.1_Dendra observed with ABIN361314 reflects the staining of the native protein Kv2.1 (obtained with its specific antibody).

Handling

Concentration: 1 mg/mL

Buffer: Phosphate buffered saline (PBS), pH 7.4, 15 mM sodium azide

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

Storage Comment: Store at 2-8°C. Do not freeze.

References

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Morgner, Bornes, Hahn, López-Iglesias, Kroese, Pritchard, Vennin, Peters, Huijbers, van Rheenen: "A Lamb1Dendra2 mouse model identifies basement-membrane-producing origins and dynamics in PyMT breast tumors." in: Developmental cell, Vol. 58, Issue 7, pp. 535-549.e5, (2023) (PubMed).

Zhang, Zhou, Li, Mo, Ma, Ni, Yang, He, Luo: "Tel2 regulates redifferentiation of bipotential progenitor cells via Hhex during zebrafish liver regeneration." in: Cell reports, Vol. 39, Issue 1, pp. 110596, (2022) (PubMed).

Wang, Fan, Khawaja, Liu, Zhan, Kodama, Chin, Li, Le, Zhou, Condello, Grinberg, Seeley, Miller, Mok, Gestwicki, Cuervo, Luo, Gan: "Microglial NF-κB drives tau spreading and toxicity in a mouse model of tauopathy." in: Nature communications, Vol. 13, Issue 1, pp. 1969, (2022) (PubMed).

Singh, Ye, Kim: "Dual leucine zipper kinase regulates Dscam expression through a non-canonical function of the cytoplasmic poly(A)-binding protein." in: The Journal of neuroscience : the official journal of the Society for Neuroscience, (2022) (PubMed).

Chen, He, Luo: "Brain vascular damage-induced lymphatic ingrowth is directed by Cxcl12b/Cxcr4a." in: Development (Cambridge, England), Vol. 149, Issue 13, (2022) (PubMed).

Singh, Kim: "Generation and validation of pX-UASTattB for dose-dependent misexpression studies in Drosophila." in: microPublication biology, Vol. 2022, (2022) (PubMed).

Teh, Singh, Ren, Huang, Ariyarathne, Salamon, Wang, Kotake, Fujita: "Surface-localized glycoproteins act through class C ARFs to fine-tune gametophore initiation in Physcomitrium patens." in: Development (Cambridge, England), Vol. 149, Issue 24, (2022) (PubMed).

Bellec, Pinot, Gicquel, Le Borgne: "The Clathrin adaptor AP-1 and Stratum act in parallel pathways to control Notch activation in Drosophila sensory organ precursors cells." in: Development (Cambridge, England), Vol. 148, Issue 1, (2021) (PubMed).

Dong, Wang, Kao, Diaz, Tasset, Kaushik, Thiruthuvanathan, Zintiridou, Nieves, Dzieciatkowska, Reisz, Gavathiotis, DAlessandro, Will, Cuervo: "Chaperone-mediated autophagy sustains haematopoietic stem-cell function." in: Nature, Vol. 591, Issue 7848, pp. 117-123, (2021) (PubMed).

Chen, Li, Ni, Chen, Yang, He, Luo: "Acute brain vascular regeneration occurs via lymphatic transdifferentiation." in: Developmental cell, Vol. 56, Issue 22, pp. 3115-3127.e6, (2021) (PubMed).

Dong, Aguirre-Hernandez, Scrivo, Eliscovich, Arias, Bravo-Cordero, Cuervo: "Monitoring spatiotemporal changes in chaperone-mediated autophagy in vivo." in: Nature communications, Vol. 11, Issue 1, pp. 645, (2020) (PubMed).

Chen, He, Ni, Yang, Zhang, Luo: "Cerebrovascular Injuries Induce Lymphatic Invasion into Brain Parenchyma to Guide Vascular Regeneration in Zebrafish." in: Developmental cell, Vol. 49, Issue 5, pp. 697-710.e5, (2019) (PubMed).

Patrizio, Renner, Pizzarelli, Triller, Specht: "Alpha subunit-dependent glycine receptor clustering and regulation of synaptic receptor numbers." in: Scientific reports, Vol. 7, Issue 1, pp. 10899, (2018) (PubMed).

Bergé, Campagne, Mignolet, Holden, Théraulaz, Manley, Allain, Viollier: "Modularity and determinants of a (bi-)polarization control system from free-living and obligate intracellular bacteria." in: eLife, Vol. 5, (2017) (PubMed).

McEwen, Maher, Mo, Gottardi: "E-cadherin phosphorylation occurs during its biosynthesis to promote its cell surface stability and adhesion." in: Molecular biology of the cell, Vol. 25, Issue 16, pp. 2365-74, (2015) (PubMed).

Target Details

Target: Dendra 2

Alternative Name: Dendra2