Anti histone h2b

Manufactured by Santa Cruz Biotechnology

Sourced in China

Anti-histone H2B is a laboratory reagent used to detect and quantify histone H2B in various biological samples. It is a specific antibody that binds to the histone H2B protein, a core component of nucleosomes in eukaryotic cells. This reagent can be used in techniques such as Western blotting, immunoprecipitation, and immunohistochemistry to study the expression and localization of histone H2B.

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Histone H2B (6 citations) Anti-H2B (2 citations) Histones (2 citations)

5 protocols using anti histone h2b

Quantifying Extracellular DNA in BALF

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Extracellular DNA levels in BALF supernatant was determined by the Quant-iT dsDNA HS kit (Invitrogen). To visualize the EETs formation by immunofluorescence microscopy, BALF eosinophils (2 × 10 5 /ml) were plated in eight-chamber culture slides and incubated at 37°C with 5% carbon dioxide for 1 hr. Afterward, BALF eosinophils were incubated with 4% paraformaldehyde (PFA) for 45 min. Next BALF eosinophils were incubated for 45 min with primary antibodies, anti-EPO and anti-histone H2B (1:250; Santa Cruz Biotechnology). After BSA 5%; Cell Signaling Technology), or anti-β-Actin (1:1000 in BSA 5%; Cell Signaling Technology) overnight. Finally, it was incubated with secondary antibody, horseradish peroxidase anti-rabbit (1:1000 in BSA 5%; Cell signaling). The blot was developed using a Chemiluminescent photo finder (Kodak/Carestream). The bands were normalized by β-actin using Image J (Rueden et al., 2017) (link).

Silveira J.S., Antunes G.L., Kaiber D.B., da Costa M.S., Ferreira F.S., Marques E.P., Schmitz F., Gassen R.B., Breda R.V., Wyse A.T.S., Stein R.T., Pitrez P.M, & da Cunha A.A. (2020). Autophagy induces eosinophil extracellular traps formation and allergic airway inflammation in a murine asthma model. Journal of cellular physiology, 235(1).

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Lipopolysaccharide-Induced Inflammatory Signaling

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BA (purity ≥98%) was purchased from Psaitong (Beijing, China). Dimethyl sulphoxide (DMSO) and LPS were purchased from Sigma-Aldrich (St. Louis, MO). Antibodies used for western blotting were as follows: anti-iNOS/NOS Type II (#610332; BD Biosciences, San Jose, CA); anti-cyclooxygenase (COX)-2 (#160106), anti-phospho-IKKα/β (S176/180, 16A6, #2697P), anti-phospho-IκBα (Ser32, #2859), anti-IκBα (44D4, #4814), anti-phospho-ERK1/2 (Thr202/Tyr204, #9101), anti-ERK1/2 (#9102), anti-phospho-p38 MAPK (Thr180/Tyr182, #4511), anti-p38 MAPK (#9212), anti-phospho-SAPK/JNK (Thr183/Tyr185, #9251), anti-JNK2 (56G8, #9258) and anti-NF-κB p65 (D14E12, #8242) (Cell Signaling Technology, Beverly, MA); anti-IKKα (CHUK, #A2062; ABclonal Technology, Woburn, MA); anti-β-tubulin (#CW0098A) and anti-GAPDH (#CW0266A) (CWBiotech, Beijing, China); and anti-histone H2B (Santa Cruz Biotechnology, Inc., Dallas, TX).

Liu S., Che N., Ou W., Yan M., Liao Y, & Cheng Y. (2022). Bullatine A exerts anti-inflammatory effects by inhibiting the ROS/JNK/NF-κB pathway and attenuating systemic inflammatory responses in mice. Pharmaceutical Biology, 60(1), 1840-1849.

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Quantification and Visualization of EETs in RSV-induced BALF

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Extracellular DNA traps were quantified in BALF cells supernatants stimulated with RSV (10³ PFU/mL) or unstimulated using Quant-iT ds DNA HS (Invitrogen, Carlsbad, CA, USA) and measured in the Quibit 2.0 fluorimeter (Invitrogen), according to the manufacturer's recommendations. Visualization of EETs by fluorescence microscopy BALF cells (2 × 10⁵/mL) were stimulated with phorbol-12-myristate-13-acetate (50 nM). After that, cells stimulated with RSV (10³ PFU/mL) and unstimulated ones were incubated for 3 hours, fixed with 4% paraformaldehyde and stained with anti-eosinophil peroxidase (EPO) and anti-histone H2B (1:250; Santa Cruz Biotechnology, Dallas, TX, USA) for 45 minutes. Later, cells were incubated with fluorescein isothiocyanate (1:100; Santa Cruz Biotechnology) and alexa fluor 633 (1:100; Invitrogen) for 30 minutes. Next, the cells were stained with Hoechst 33342 DNA dye (1:2,000; Invitrogen) for 4 minutes. Confocal images were taken on a Leica TCS-SP8 exciter microscope (Leica Microsystem, Wetzlar, Germany).

Silveira J.S., Antunes G.L., Gassen R.B., Breda R.V., Stein R.T., Pitrez P.M, & da Cunha A.A. (2019). Respiratory syncytial virus increases eosinophil extracellular traps in a murine model of asthma. Asia Pacific Allergy, 9(4), e32.

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Cellular Signaling Pathway Regulation Analysis

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Reagents and antibodies BA (purity≥98%) was purchased from Psaitong (Beijing, China). Dimethyl sulfoxide (DMSO) and LPS was obtained from Sigma (St. Louis, MO, USA). Antibodies used for western blotting in the present study were as follows: the primary antibodies against inducible Nitric Oxide Synthase (iNOS) (#13120), anticyclooxygenase-2 (COX-2) (#12282), anti-phospho-IKKα/β (S176/180) (16A6) (#2697P), anti-phospho-IκBα (Ser32) (#2859), anti-IκBα (44D4) (#4814), anti-phospho-ERK1/2 (Thr202/Tyr204) (#9101), anti-ERK1/2 (#9102), anti-phospho-p38 MAPK (Thr180/Tyr182) (#4511), anti-p38 MAPK (#9212), antiphospho-SAPK/JNK (Thr183/Tyr185) (#9251), anti-JNK2 (56G8) (#9258), anti-NF-κB p65 (D14E12) ( #8242) were both purchased from Cell Signaling Technology (Beverly, MA, USA). Anti-IKKα (CHUK) (#A2062) was from ABclonal Technology (Wuhan, HB, China), β-tubulin (#CW0098A) and GAPDH (#CW0266A) were from CWBiotech (Beijing, China). Anti-Histone H2B were obtained from Santa Cruz Biotechnology Co., Ltd. (Shanghai, China).

Liu S., Yan M., Liao Y., & Cheng Y. (2021). Bullatine a Exerted Anti-Inflammatory Effects by Inhibiting JNK/ROS/NF-κB Pathway and Attenuates Systemic Inflammatory Response in LPS-Challenged Mice.

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Immunofluorescence Analysis of Lung Tissue

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Cross sections (9 μm) of OCT-embedded frozen lung tissues were fixed in 4% paraformaldehyde, blocked in 8% BSA in PBS and incubated with the primary antibodies: anti-Histone H2B (1:100 dilution; Cat # SC-8651; Santa Cruz Biotechnology), anti-mouse MPO (1:100 dilution; Cat # HM1051BT; Hycult Biotech) followed by the appropriate rhodamine red- or FITC-conjugated secondary antibody (1:100–1:200; Jackson ImmunoResearch Laboratories). DNA was stained with DAPI. All images were acquired with QCapture software on a Nikon Eclipse microscope.

Akk A., Springer L.E, & Pham C.T. (2016). Neutrophil Extracellular Traps Enhance Early Inflammatory Response in Sendai Virus-Induced Asthma Phenotype. Frontiers in Immunology, 7, 325.

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