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Rabbit Anti-P38 MAPK  antibody (bs-0637R)  
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產(chǎn)品編號(hào) bs-0637R
英文名稱(chēng) Rabbit Anti-P38 MAPK  antibody
中文名稱(chēng) 絲裂原活化蛋白激酶p38α抗體
別    名 CSAID Binding Protein 1; CSAID binding protein; CSAID-binding protein; Csaids binding protein; CSBP 1; CSBP 2; CSBP; CSBP1; CSBP2; CSPB 1; CSPB1; Cytokine suppressive anti inflammatory drug binding protein; Cytokine suppressive anti-inflammatory drug-binding protein; EXIP; MAP kinase 14; MAP kinase MXI2; MAP kinase p38 alpha; MAPK 14; MAPK14; MAX interacting protein 2; MAX-interacting protein 2; Mitogen Activated Protein Kinase 14; Mitogen activated protein kinase p38 alpha; Mitogen-activated protein kinase 14; Mitogen-activated protein kinase p38 alpha; MK14_HUMAN; Mxi 2; Mxi2; p38 ALPHA; p38; p38 MAP kinase; p38 MAPK; p38/MAPK; p38 mitogen activated protein kinase; p38ALPHA; p38alpha Exip; PRKM14; PRKM15; RK; SAPK 2A; SAPK2A; Stress Activated Protein Kinase 2A.  
Specific References  (106)     |     bs-0637R has been referenced in 106 publications.
[IF=20.693] Myung-Ju Lee. et al. CXCL1 confers a survival advantage in Kaposi's sarcoma-associated herpesvirus-infected human endothelial cells through STAT3 phosphorylation. J MED VIROL. 2022 Jul;:  WB ;  Human.  
[IF=17.521] Yi Yan. et al. Nanomedicines Reprogram Synovial Macrophages by Scavenging Nitric Oxide and Silencing CA9 in Progressive Osteoarthritis. Advanced Science. 2023 Feb;:2207490  WB ;  Mouse.  
[IF=12.91] Kuo-Chu Lai. et al. IFIT2-depleted metastatic oral squamous cell carcinoma cells induce muscle atrophy and cancer cachexia in mice. 2022 Feb 15  WB ;  Mouse.  
[IF=9.473] Shuting Wei. et al. Particle matters induce airway epithelial barrier dysfunction in vivo and in vitro: from a more realistic inhalation scenario. ENVIRON SCI-NANO. 2022 Jun;:  WB ;  Human.  
[IF=9.381] Zhaomin Zheng. et al. New insight into the structure-dependent two-way immunomodulatory effects of water-soluble yeast β-glucan in macrophages. CARBOHYD POLYM. 2022 Sep;291:119569  WB ;  Mouse.  
[IF=8.943] Ze Kan. et al. Respiratory exposure to graphene oxide induces pulmonary fibrosis and organ damages in rats involving caspase-1/p38MAPK/TGF-β1 signaling pathways. CHEMOSPHERE. 2022 Jun;:135181  WB ;  Rat.  
[IF=7.963] Meiqiong Wu. et al. Suppression of NADPH oxidase 4 inhibits PM2.5-induced cardiac fibrosis through ROS-P38 MAPK pathway. SCI TOTAL ENVIRON. 2022 Apr;:155558  WB ;  Mouse,Rat.  
[IF=7.675] Honghong Zhan. et al. Oxybaphus himalaicus Mitigates Lipopolysaccharide-Induced Acute Kidney Injury by Inhibiting TLR4/MD2 Complex Formation. ANTIOXIDANTS-BASEL. 2022 Dec;11(12):2307  WB ;  Mouse.  
[IF=7.129] Jin Chen. et al. Surface functionalization-dependent inflammatory potential of polystyrene nanoplastics through the activation of MAPK/ NF-κB signaling pathways in macrophage Raw 264.7. ECOTOX ENVIRON SAFE. 2023 Feb;251:114520  WB ;  Mouse.  
[IF=7.129] Tianqing Huang. et al. miR-301b-5p and its target gene nfatc2ip regulate inflammatory responses in the liver of rainbow trout (Oncorhynchus mykiss) under high temperature stress. ECOTOX ENVIRON SAFE. 2022 Sep;242:113915  WB ;  Fish.  
[IF=6.706] Najla Hajji. et al. The Role of Globularia alypum Explored Ex Vivo In Vitro on Human Colon Biopsies from Ulcerative Colitis Patients. NUTRIENTS. 2023 Jan;15(6):1457  IHC ;  Human.  
[IF=6.656] Weina Qian. et al. Exploring the mechanism of Xingpi Capsule in diarrhea predominant-irritable bowel syndrome treatment based on multiomics technology. PHYTOMEDICINE. 2023 Mar;111:154653  WB ;  Human.  
[IF=6.551] Shuming Chen. et al. Melamine induces reproductive dysfunction via down-regulated the phosphorylation of p38 and downstream transcription factors Max and Sap1a in mice testes. Sci Total Environ. 2021 May;770:144727  WB ;  Mouse.  
[IF=5.923] Junfeng Ke. et al. CTI-2 Inhibits Metastasis and Epithelial-Mesenchymal Transition of Breast Cancer Cells by Modulating MAPK Signaling Pathway. Int J Mol Sci. 2021 Jan;22(22):12229  WB,IF ;  Human.  
[IF=5.714] Xiaoxiang Peng. et al. Phasic regulation of the ATP/P2X7 receptor signaling pathway affects the function of antigen-presenting cells in experimental autoimmune uveitis. INT IMMUNOPHARMACOL. 2023 Jun;119:110241  WB ;  Mouse.  
[IF=5.714] Ruyi Qu. et al. Glucocorticoids improve the balance of M1/M2 macrophage polarization in experimental autoimmune uveitis through the P38MAPK-MEF2C axis. INT IMMUNOPHARMACOL. 2023 Jul;120:110392  WB ;  Rat.  
[IF=5.614] Zhu J et al. SPARC Promotes Self‐Renewal of Limbal Epithelial Stem Cells and Ocular Surface Restoration through JNK and p38‐MAPK Signaling Pathways. Stem Cells. 2019 Oct 23.  WB ;  Rabbit.  
[IF=5.455] Zhang, Rongrong. et al. Compound traditional Chinese medicine dermatitis ointment ameliorates inflammatory responses and dysregulation of itch-related molecules in atopic dermatitis. Chin Med-Uk. 2022 Dec;17(1):1-19  WB ;  Mouse.  
[IF=5.314] Yang Wang. et al. Role of GADD45A in myocardial ischemia/reperfusion through mediation of the JNK/p38 MAPK and STAT3/VEGF pathways. INT J MOL MED. 2022 Dec;50(6):1-11  WB ;  Rat.  
[IF=5.285] Huawei Liu. et al. Integrated multi-omics reveals the beneficial role of chlorogenic acid in improving the growth performance and immune function of immunologically-stressed broilers. ANIM NUTR. 2023 May;:  WB ;  Chicken.  
[IF=5.156] Li, Muzhe. et al. Effects of adenovirus-mediated knockdown of IRAK4 on synovitis in the osteoarthritis rabbit model. Arthritis Res Ther. 2021 Dec;23(1):1-12  WB ;  Rabbits.  
[IF=4.996] Zhao, Lei. et al. Lactobacillus plantarum S9 alleviates lipid profile, insulin resistance, and inflammation in high-fat diet-induced metabolic syndrome rats. SCI REP-UK. 2022 Sep;12(1):1-10  WB ;  Rat.  
[IF=4.927] Yanjie Wang. et al. Inhibitory Effect of Bisdemethoxycurcumin on DNCB-Induced Atopic Dermatitis in Mice. MOLECULES. 2023 Jan;28(1):293  WB ;  Mouse.  
[IF=4.868] Wang G et al. Protective?Effect?of?Methane-Rich?Saline?on?Acetic?Acid-Induced?Ulcerative?Colitis?via?Blockingthe?TLR4/NF-κB/MAPK?Pathway?and?Promoting?IL-10/JAK1/STAT3-Mediated Anti-inflammatory Response. Oxid Med Cell Longev.?2019 Apr 28;2019:7850324.  WB ;  Mouse.  
[IF=4.658] Zhang J et al. Down‐regulation of Suv39h1 attenuates neointima formation after carotid artery injury in diabetic rats. J Cell Mol Med. 2019 Nov 17.  WB ;  Rat.  
[IF=4.657] Shen, Yue. et al. Hederagenin Suppresses Inflammation and Cartilage Degradation to Ameliorate the Progression of Osteoarthritis: An In vivo and In vitro Study. INFLAMMATION. 2022 Nov;:1-24  WB ;  Human.  
[IF=4.645] Pin-Shi Ni. et al. Indirect regulation of HIPPO pathway by miRNA mediates high-intensity intermittent exercise to ameliorate aging skeletal muscle function. SCAND J MED SCI SPOR. 2023 Feb;:  WB ;  Rat.  
[IF=4.61] Yue H et al. Gestational exposure to PM2.5 impairs vascularization of the placenta. Sci Total Environ. 2019 May 15;665:153-161.  WB ;  Mouse.  
[IF=4.55] Ning, Chong, et al. "Chicory inulin ameliorates type 2 diabetes mellitus and suppresses JNK and MAPK pathways in vivo and in vitro." Molecular Nutrition & Food Research (2017).  WB ;  Rat.  
[IF=4.501] Jia-Lin Wang. et al. Silencing UBQLN2 Enhances the Radiosensitivity of Esophageal Squamous Cell Carcinoma (ESCC) via Activating p38 MAPK. J ONCOL. 2023 Jan 05;2023:2339732  IHC ;  Human,Mouse.  
研究領(lǐng)域 腫瘤  免疫學(xué)  信號(hào)轉(zhuǎn)導(dǎo)  細(xì)胞凋亡  轉(zhuǎn)錄調(diào)節(jié)因子  
抗體來(lái)源 Rabbit
克隆類(lèi)型 Polyclonal
交叉反應(yīng) Human,Mouse,Rat (predicted: Rabbit,Sheep,Dog)
產(chǎn)品應(yīng)用 WB=1:500-2000,IHC-P=1:100-500,IHC-F=1:100-500,Flow-Cyt=1μg/Test,ICC/IF=1:100,IF=1:100-500
not yet tested in other applications.
optimal dilutions/concentrations should be determined by the end user.
理論分子量 41kDa
細(xì)胞定位 細(xì)胞核 細(xì)胞漿 
性    狀 Liquid
濃    度 1mg/ml
免 疫 原 KLH conjugated synthetic peptide derived from human P38MAPK: 141-240/360 
亞    型 IgG
純化方法 affinity purified by Protein A
緩 沖 液 0.01M TBS (pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol.
保存條件 Shipped at 4℃. Store at -20℃ for one year. Avoid repeated freeze/thaw cycles.
注意事項(xiàng) This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications.
PubMed PubMed
產(chǎn)品介紹 The protein encoded by this gene is a member of the MAP kinase family. MAP kinases act as an integration point for multiple biochemical signals, and are involved in a wide variety of cellular processes such as proliferation, differentiation, transcription regulation and development. This kinase is activated by various environmental stresses and proinflammatory cytokines. The activation requires its phosphorylation by MAP kinase kinases(MKKs), or its autophosphorylation triggered by the interaction of MAP3K7IP1/TAB1 protein with this kinase. The substrates of this kinase include transcription regulator ATF2, MEF2C, and MAX, cell cycle regulator CDC25B, and tumor suppressor p53, which suggest the roles of this kinase in stress related transcription and cell cycle regulation, as well as in genotoxic stress response. Four alternatively spliced transcript variants of this gene encoding distinct isoforms have been reported.

Function:
Serine/threonine kinase which acts as an essential component of the MAP kinase signal transduction pathway. MAPK14 is one of the four p38 MAPKs which play an important role in the cascades of cellular responses evoked by extracellular stimuli such as proinflammatory cytokines or physical stress leading to direct activation of transcription factors. Accordingly, p38 MAPKs phosphorylate a broad range of proteins and it has been estimated that they may have approximately 200 to 300 substrates each. Some of the targets are downstream kinases which are activated through phosphorylation and further phosphorylate additional targets. RPS6KA5/MSK1 and RPS6KA4/MSK2 can directly phosphorylate and activate transcription factors such as CREB1, ATF1, the NF-kappa-B isoform RELA/NFKB3, STAT1 and STAT3, but can also phosphorylate histone H3 and the nucleosomal protein HMGN1. RPS6KA5/MSK1 and RPS6KA4/MSK2 play important roles in the rapid induction of immediate-early genes in response to stress or mitogenic stimuli, either by inducing chromatin remodeling or by recruiting the transcription machinery. On the other hand, two other kinase targets, MAPKAPK2/MK2 and MAPKAPK3/MK3, participate in the control of gene expression mostly at the post-transcriptional level, by phosphorylating ZFP36 (tristetraprolin) and ELAVL1, and by regulating EEF2K, which is important for the elongation of mRNA during translation. MKNK1/MNK1 and MKNK2/MNK2, two other kinases activated by p38 MAPKs, regulate protein synthesis by phosphorylating the initiation factor EIF4E2. MAPK14 interacts also with casein kinase II, leading to its activation through autophosphorylation and further phosphorylation of TP53/p53. In the cytoplasm, the p38 MAPK pathway is an important regulator of protein turnover. For example, CFLAR is an inhibitor of TNF-induced apoptosis whose proteasome-mediated degradation is regulated by p38 MAPK phosphorylation. In a similar way, MAPK14 phosphorylates the ubiquitin ligase SIAH2, regulating its activity towards EGLN3. MAPK14 may also inhibit the lysosomal degradation pathway of autophagy by interfering with the intracellular trafficking of the transmembrane protein ATG9. Another function of MAPK14 is to regulate the endocytosis of membrane receptors by different mechanisms that impinge on the small GTPase RAB5A. In addition, clathrin-mediated EGFR internalization induced by inflammatory cytokines and UV irradiation depends on MAPK14-mediated phosphorylation of EGFR itself as well as of RAB5A effectors. Ectodomain shedding of transmembrane proteins is regulated by p38 MAPKs as well. In response to inflammatory stimuli, p38 MAPKs phosphorylate the membrane-associated metalloprotease ADAM17. Such phosphorylation is required for ADAM17-mediated ectodomain shedding of TGF-alpha family ligands, which results in the activation of EGFR signaling and cell proliferation. Another p38 MAPK substrate is FGFR1. FGFR1 can be translocated from the extracellular space into the cytosol and nucleus of target cells, and regulates processes such as rRNA synthesis and cell growth. FGFR1 translocation requires p38 MAPK activation. In the nucleus, many transcription factors are phosphorylated and activated by p38 MAPKs in response to different stimuli. Classical examples include ATF1, ATF2, ATF6, ELK1, PTPRH, DDIT3, TP53/p53 and MEF2C and MEF2A. The p38 MAPKs are emerging as important modulators of gene expression by regulating chromatin modifiers and remodelers. The promoters of several genes involved in the inflammatory response, such as IL6, IL8 and IL12B, display a p38 MAPK-dependent enrichment of histone H3 phosphorylation on 'Ser-10' (H3S10ph) in LPS-stimulated myeloid cells. This phosphorylation enhances the accessibility of the cryptic NF-kappa-B-binding sites marking promoters for increased NF-kappa-B recruitment. Phosphorylates CDC25B and CDC25C which is required for binding to 14-3-3 proteins and leads to initiation of a G2 delay after ultraviolet radiation. Phosphorylates TIAR following DNA damage, releasing TIAR from GADD45A mRNA and preventing mRNA degradation. The p38 MAPKs may also have kinase-independent roles, which are thought to be due to the binding to targets in the absence of phosphorylation. Protein O-Glc-N-acylation catalyzed by the OGT is regulated by MAPK14, and, although OGT does not seem to be phosphorylated by MAPK14, their interaction increases upon MAPK14 activation induced by glucose deprivation. This interaction may regulate OGT activity by recruiting it to specific targets such as neurofilament H, stimulating its O-Glc-N-acylation. Required in mid-fetal development for the growth of embryo-derived blood vessels in the labyrinth layer of the placenta. Also plays an essential role in developmental and stress-induced erythropoiesis, through regulation of EPO gene expression. Isoform MXI2 activation is stimulated by mitogens and oxidative stress and only poorly phosphorylates ELK1 and ATF2. Isoform EXIP may play a role in the early onset of apoptosis. Phosphorylates S100A9 at 'Thr-113'.

Subunit:
Binds to a kinase interaction motif within the protein tyrosine phosphatase, PTPRR (By similarity). This interaction retains MAPK14 in the cytoplasm and prevents nuclear accumulation. Interacts with SPAG9 and GADD45A. Interacts with CDC25B, CDC25C, DUSP1, DUSP10, DUSP16, NP60, FAM48A and TAB1. Interacts with casein kinase II subunits CSNK2A1 and CSNK2B.

Subcellular Location:
Cytoplasm. Nucleus.

Tissue Specificity:
Brain, heart, placenta, pancreas and skeletal muscle. Expressed to a lesser extent in lung, liver and kidney.

Post-translational modifications:
Dually phosphorylated on Thr-180 and Tyr-182 by the MAP2Ks MAP2K3/MKK3, MAP2K4/MKK4 and MAP2K6/MKK6 in response to inflammatory citokines, environmental stress or growth factors, which a ctivates the enzyme. Dual phosphorylation can also be mediated by TAB1-mediated autophosphorylation. TCR engagement in T-cells also leads to Tyr-323 phosphorylation by ZAP70. Dephosphorylated and inactivated by DUPS1, DUSP10 and DUSP16.
Acetylated at Lys-53 and Lys-152 by KAT2B and EP300. Acetylation at Lys-53 increases the affinity for ATP and enhances kinase activity. Lys-53 and Lys-152 are deacetylated by HDAC3.
Ubiquitinated. Ubiquitination leads to degradation by the proteasome pathway.

Similarity:
Belongs to the protein kinase superfamily. CMGC Ser/Thr protein kinase family. MAP kinase subfamily.
Contains 1 protein kinase domain.

SWISS:
Q16539

Gene ID:
1432

Database links:

Entrez Gene: 1432 Human

Entrez Gene: 26416 Mouse

Entrez Gene: 81649 Rat

Entrez Gene: 403856 Dog

GenBank: NM_001315 Human

GenBank: NM_139012 Human

GenBank: NM_011951 Mouse

GenBank: NM_031020 Rat

Omim: 600289 Human

SwissProt: O02812 Dog

SwissProt: Q16539 Human

SwissProt: P47811 Mouse

SwissProt: P70618 Rat

Unigene: 485233 Human

Unigene: 311337 Mouse

Unigene: 88085 Rat



產(chǎn)品圖片
Sample: Muscle (Mouse) Lysate at 40 ug Primary: Anti-P38 MAPK (bs-0637R) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 41 kD Observed band size: 41 kD
Paraformaldehyde-fixed, paraffin embedded (rat liver tissue); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (MAPK) Polyclonal Antibody, Unconjugated (bs-0637R) at 1:400 overnight at 4°C, followed by a conjugated secondary (sp-0023) for 20 minutes and DAB staining.
Paraformaldehyde-fixed, paraffin embedded (rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-P38 MAPK (Thr180 + Tyr182) ) Polyclonal Antibody, Unconjugated (bs-0637R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (P38 MAPK) Polyclonal Antibody, Unconjugated (bs-0637R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Tissue/cell: mouse embryo tissue; 4% Paraformaldehyde-fixed and paraffin-embedded; Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min; Incubation: Anti-p38MAPK/MAPK14/p38Alpha Polyclonal Antibody, Unconjugated(bs-0637R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (P38 MAPK) Polyclonal Antibody, Unconjugated (bs-0637R) at 1:500 overnight at 4°C, followed by a conjugated secondary (sp-0023) for 20 minutes and DAB staining.
Tissue/cell: MCF7 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (P38 MAPK) polyclonal Antibody, Unconjugated (bs-0637R) 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.
Tissue/cell: HUVEC cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (P38 MAPK) polyclonal Antibody, Unconjugated (bs-0637R) 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody (bs-0295G-FITC) at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.
Blank control: HepG2(blue). Primary Antibody:Rabbit Anti-P38 MAPK antibody (bs-0637R,Green); Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA; Isotype Control Antibody: Rabbit IgG(orange) ,used under the same conditions; Secondary Antibody: Goat anti-rabbit IgG-FITC(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA. Protocol The cells were fixed with 2% paraformaldehyde for 10 min at 37℃. Primary antibody (bs-0637R, 1μg /1x10^6 cells) were incubated for 30 min at room temperature, followed by 1 X PBS containing 0.5% BSA + 1 0% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/FITC antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 40 min at room temperature. Acquisition of 20,000 events was performed.
Blank control: Raw264.7. Primary Antibody (green line): Rabbit Anti-P38 MAPK antibody (bs-0637R) Dilution: 2μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody : Goat anti-rabbit IgG-AF488 Dilution: 1μg /test. Protocol The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 90% ice-cold methanol for 20 min at-20℃. The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.
Blank control:MCF7. Primary Antibody (green line): Rabbit Anti-P38 MAPK antibody (bs-0637R) Dilution: 2μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody : Goat anti-rabbit IgG-FITC Dilution: 1μg /test. Protocol The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 90% ice-cold methanol for 20 min at-20℃. The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.
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