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H3 | Histone H3 (antigen affinity purified)

AS10 710A | Clonality: Polyclonal | Host: Rabbit | Reactivity: A. thaliana, S. lycopersicum, V. faba, P. patens, S. europaea, Z. mays, Ch. reinhardtii | cellular [compartment marker] of nucleoplasm

H3 | Histone H3 (antigen affinity purified) in the group Antibodies Plant/Algal  / Compartment Markers / Nuclear marker at Agrisera AB (Antibodies for research) (AS10 710A)
H3 | Histone H3 (antigen affinity purified)
H3 | Histone H3 (antigen affinity purified)
H3 | Histone H3 (antigen affinity purified)



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Product Information

Immunogen

KLH-conjugated synthetic peptide derived from known H3 sequences, inluding Arabidopsis thaliana H3.3 P59169 (At4g40030,At4g40040,At5g10980), H3.2 P59226(At1g09200,At3g27360,At5g10390,At5g10400,At5g65360), H3-like 2 Q9FXI7 (At1g19890)

Host Rabbit
Clonality Polyclonal
Purity Immunogen affinity purified serum in PBS pH 7.4.
Format Lyophilized
Quantity 50 µg
Reconstitution For reconstitution add 50 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 ChIp-qPCR (ChIp-qPCR), Immunofluorescence (IF), Western blot (WB)
Recommended dilution 2,5 µg/100 µg of chromatin (ChIp-qPCR), 1: 400 (IF), 1 : 5000 (WB)
Expected | apparent MW 15 | 17 kDa

Reactivity

Confirmed reactivity Arabidopsis thaliana, Oryza sativa
Predicted reactivity

Brassica oleracea, Capsicum annuum, Chlamydomonas acidophila, Chlamydomonas reinhardtii, Physcomitrium patens, Salicornia europaea, Solanum lycopersicum, Solanum sogarandinum, Solanum tuberosum, Vicia faba, Zea mays Brachypodium distachyon, Brassica napus, Hordeum vulgare, Nicotiana tabacum, Malus domestica, Medicago sativa, Nannochloropsis gaditana, Triticum aestivum, Pinus pinaster, Pisum sativum, Zea mays, Vitis vinifera, Volvox sp.


Species of your interest not listed? Contact us
Not reactive in No confirmed exceptions from predicted reactivity are currently known

Application examples

Application examples Application example

Immunofluorescent detection of plant histone 3 using Agrisera antigen affinity purified antibodies

Immunofluorescent localization of plant histone 3 in rice suspension culture

Immunofluorescent localization of Histone 3 on suspension culture of Arabidopsis thaliana (upper image) or Oryza sativa (bottom image), using anti-histone 3 antibodies (AS10 710A) and anti-rabbit IgG DyLight®488 conjugated secondary antibodies (AS10 1165). DAPI staining of nuclei is pseudocolored red. 

Material: Suspension cultures of Arabidopsis thaliana, ecotype Landsberg erecta cv.MM1 or 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) 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: 30 minutes (Arabidopsis thaliana) or 60 minutes (Oryza sativa). Cells were not shaken during the first 5 mins of fixation to allowed to partially recover from osmotic shock induced by formaldehyde.
Hydrophilization: no
Cell wall digestion: Yes
Packed cell volume to enzyme ratio: 100 µl : 2ml Enzyme composition: 1% (A) 1.2% (R) Cellulase (chromatically purified, powder, Worthington) 1% 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: 30 minutes (Arabidopsis thaliana) or 90 minutes (Oryza sativa)
Membrane permeabilization: Triton-X100 (0.5%), 10 min/RT
Antigen retrieval: no
Blocking buffer: Fish gelatin (5% v/v)
Washing buffer: PBS
Primary antibody dilution and incubation time: 1:400, ON/4ºC
Secondary antibody dilution and incubation time and supplier: anti-rabbit IgG DyLight®488 conjugated secondary antibodies (AS10 1165), 1:600, 1hn/RT
Co-staining of the nucleus (DAPI): Yes
Nucleus staining: 100 ng/ml DAPI

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



IP using polyclonal anti-plant H3 antibody

Chromatin Immunoprecipitation: using anti-plant Histone 3 polyclonal antibodies. Chromatin from Arabidopsis thaliana wilde type, deacetylase mutant and over-expressors was cross-linked using formaldehyde. Chromatin was isolated and DNA was sheared along with the bound protein by sonictaion. DNA-protein complex was immunoprecipitated using affinity purified, polyclonal anti-Histone 3 antibodies. Immunoprecipitated DNA was quantified using quantitative PCR and normalized to the input chromatin.
Procedure was according to a protocol described here: Saleh et al. (2008).

Courtesy of Dr. Cristián Holzmann, Catholic University of Chile, Chile

Additional information

Additional information Cellular [compartment marker] of nucleoplasm, loading control antibody for Chlamydomonas reinhardtii

Protocol for isolation of cytosolic and nuclear fractions can be found here.

Specific protol for ChIP can be found here: Saleh et al. (2008).

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Background

Background Histone 3 (H3) located in nuclei, incorporated into chromatin. Present in nucleosome together with H2A, H2B and H4.

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