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V-ATPase | epsilon subunit of tonoplast H+ATPase (affinty purified, goat antibody)

AS09 577A Clonality: Polyclonal  |  Host: Goat  |  Reactivity: Higher plants including A.thaliana, A.strigosa, M. truncatula, N. tabacum, S. lycopersicum

V-ATPase | epsilon subunit of tonoplast H+ATPase (affinty purified, goat antibody) in the group Antibodies Plant/Algal  / Membrane Transport System / Vacuolar membrane at Agrisera AB (Antibodies for research) (AS09 577A)



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Product Information

Immunogen

KLH-conjugated synthetic peptide chosen from subunit E of plant V-ATPase including Arabidopsis thaliana At4g11150. Peptide is conserved in vacuolar H+-ATPase subunit E, isoform 1 to 3 (VHA-E1).

Host Goat
Clonality Polyclonal
Purity Immunogen affinity purified serum in PBS pH 7.4.
Format Lyophilized
Quantity 200 ĩg
Reconstitution For reconstitution add 100 µl of sterile water
Storage Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please remember to spin the tubes briefly prior to opening them to avoid any losses that might occur from material adhering to the cap or sides of the tube.
Tested applications Immunofluorescence (IF), Western blot (WB)
Recommended dilution 1: 600 (IF), 1 : 1000-1 : 3000 (WB)
Expected | apparent MW

26 | 31 kDa (Arabidopsis thaliana)

Reactivity

Confirmed reactivity Arabidopsis thaliana, Avena strigosa, Medicago truncatula, Nicotiana tabacum, Oryza sativa, Solanum lycopersicum
Predicted reactivity Algae, Chlamydomonas reinhardtii, Hordeum vulgare, Malus domestica, Mesembryanthemum sp.,   Petunia sp.,Phaseolus sp. , Physcomitrium patens,  Pteris vittata (fern), Ricinus communis, Thellungiella sp., Zea mays, Vitis vinifera      Bull frog, Chicken, Bovine, Drosophila melanogaster, Human, Mouse, Rat
Species of your interest not listed? Contact us
Not reactive in No confirmed exceptions from predicted reactivity are currently known

Application examples

Application examples Immunofluorescent localization of V-ATPase epsilon subunit of tonoplast H+ATPase using goat anti-V-ATPase antibodies

Immunofluorescent localization of V-ATPase epsilon subunit of tonoplast H+ATPase in suspension culture of Oryza sativa ssp. japonica cv. 'Unggi 9', using goat anti-V-ATPase, epsilon subunit of tonoplast antibodies (AS09 577A) and donkey anti-Goat IgG (H&L), DyLight® 488 conjugated secondary antibodies (AS10 1116, Agrisera).  Vacuolar membrane, tonoplast, is highlighted by yellow arrowheads. DAPI staining of nuclei is pseudocolored red. 
Method
Material: Suspension cultures of Oryza sativa ssp. japonica cv. 'Unggi 9
Fixation: Packed cell volume to fixer ratio: 250 µl : 5ml
Fixer composition and buffer: 4% (w/v) paraformaldehyde (freshly prepared as 8% stock and 0.2 µm filtered) 0.01% (v/v) Triton-X100 in Phosphate Buffered Saline (PBS), pH 7.4 (2x stock, 0.2 µm filtered)
Container and method: in 6 cm Petri dish, gentle shaking at room temperature (RT)
Duration: 40 min
Hydrophilization: No
Cell wall digestion: Yes Packed cell volume to enzyme ratio: 100 µl : 2 ml Enzyme composition: 1% (A) 1.2% (R) Cellulase (chromatically purified, powder, Worthington) 1% (A) 1.2% (R) Pectinase (protease free, liquid, Sigma) Buffer: 0.5% (w/v) MES buffer, pH 5.6
Container and method: in 2 ml microfuge tube by rolling at room temperature (RT)
Duration: 60 min
Membrane permeabilization: Triton-X100 (0.35%), 7 min/RT
Antigen retrieval: No
Blocking buffer: Fish gelatin (5% v/v)
Washing buffer: PBS
Primary antibody dilution and incubation time: 1:600, 4ºC/ON
Secondary antibody: donkey anti-Goat IgG (H&L), DyLight® 488 conjugated secondary antibodies (AS10 1116, Agrisera), 1:600, 1h/RT
Co-staining of the nucleus (DAPI): Yes
Cell wall and nucleus staining: 100 ng/ml DAPI

Courtesy of Dr. Ferhan Ayaydin, Hungarian Centre of Excellence for Molecular Medicine (HCEMM), Szeged, Hungary

Additional information

V-ATPase is very sensitive for the redox of the SDS buffer. We recommend using at least 50-100 mM DTT freshly prepared before handling the sample.

2 hours incubation with primary antibody is recommended over over night incubation which can contribute to increased background.

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Background

Background

Plant vacuole V-ATPase is responsible for energization of transport of ions and metabolites, and acts as well 'house-keeping' and as a stress response enzyme. V-ATPase is a multi-subunit enzyme composed of a membrane sector and a cytosolic catalytic sector. It is related to the FoF1 ATP synthase. Alternative protein names: Vacuolar proton pump subunit E, Protein EMBRYO DEFECTIVE 2448

Product citations

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