GFP | Green Fluorescence Protein
AS20 4443 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Green Fluorescence Protein (GFP)

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Product Information
Rabbit
Reactivity
Application examples

194 µg/well of total protein extracted freshly from tobacco with write exact buffer components (50mM Tris pH 7.5, 150 mM NaCl, 1 mM EDTA, 10% Glicerol, 1 mM DTT, 1X Pefablock) and denatured with SDS 2% and 0.1% Bromophenol blue (Laemmli buffer) at 95°C for 5 min.... were separated on 12 % SDS-PAGE and blotted 2hs to PVDF/nitrocellulose (pore size of um), using wet transfer. Blot was blocked with 5% milk for 1h/RT with agitation. Blot was incubated in the primary antibody at a dilution of 1: 10 000 in TBS-T ON/4°C (with or without milk) with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed 3 times for 10 min in TBS-T at RT with agitation. Blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, AS09 602) diluted to 1:20 000 in for 1h/RT with agitation. The blot was washed as above and developed for 2 min with Agrisera ECLSuperBright with name and supplier. Exposure time was 60 seconds.
Courtesy of Delfina Gagliardi Instituto de Agrobiotecnología del Litoral (IAL), Argentina
Super-resolution image of immunofluorescent localization of GFP-fused protein in Nicotiana tabacum (tobacco) leaf tissue using Agrisera anti-GFP antibodies (AS20 4443) and donkey anti-rabbit IgG DyLight 650 conjugated secondary antibodies (Green). Calcofluor White (Blue) and Tissue Autofluorescence (Yellow),
Method:
Fixation: 2% paraformaldehyde + 0.5% glutaraldehyde, dehydration in ethanol, embedment and UV polymerization in HM20 resin
Cell wall digestion: no
Membrane permeabilization: DMSO-IGEPA
Air drying; No
Antigen retrieval: No
Blocking buffer: 4% w/v non-fat milk powder in 1X Phosphate Buffered Saline buffer, prepared freshly, filtered with 0.2 µm filer
Washing buffer: 1X Phosphate Buffered Saline buffer Primary antibody dilution and incubation time: 1:100; 1 hour at room temperature
Secondary antibody dilution and incubation time and supplier: 1:200 Donkey anti-rabbit IgG (H&L) DyLight®650 (Agrisera, AS12 2254); 1 hour at room temperature
Co-staining of the nucleus (DAPI): No
Cell wall and nucleus staining: Calcofluor White
All material was freshly prepared. All incubation volumes were kept the same: 200 μL per 22x22 mm cover glass/sample
Courtesy Dr. Kirk Czymmek, Donald Danforth Plant Science Centre, USA
Additional information
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Related products: GFP | Green Fluorescence Protein
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