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活化轉錄因子4抗體

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產品編號bs-1531R
英文名稱Rabbit Anti-ATF4 antibody
中文名稱活化轉錄因子4抗體
別    名Cyclic AMP response element binding protein 2; DNA binding protein TAXREB67; Activating Transcription Factor 4; ATF 4; ATF4 protein; CREB 2; CREB2; CREB-2; Cyclic AMP dependent transcription factor ATF 4; Tax Responsive Enhancer Element B67; TAXREB67; TXREB; ATF4_HUMAN.   
Specific References  (24)     |     bs-1531R has been referenced in 24 publications.
[IF=9.274] Wiebke Sachs. et al. Distinct Modes of Balancing Glomerular Cell Proteostasis in Mucolipidosis Type II and III Prevent Proteinuria. J Am Soc Nephrol. 2020 Aug;31(8):1796-1814  WB,IF ;  Mouse.  
[IF=6.208] Yurong Fu. et al. Zearalenone Promotes LPS-Induced Oxidative Stress, Endoplasmic Reticulum Stress, and Accelerates Bovine Mammary Epithelial Cell Apoptosis. INT J MOL SCI. 2022 Jan;23(18):10925  WB ;  Bovine.  
[IF=5.793] Chunyue Wang. et al. Neuroprotective effects of verbascoside against Alzheimer’s disease via the relief of endoplasmic reticulum stress in Aβ-exposed U251 cells and APP/PS1 mice. J Neuroinflamm. 2020 Dec;17(1):1-16  WB ;  Human.  
[IF=5.778] Yuting Wang. et al. Polybrominated diphenyl ethers quinone-induced intracellular protein oxidative damage triggers ubiquitin-proteasome and autophagy-lysosomal system activation in LO2 cells. Chemosphere. 2021 Jul;275:130034  WB ;  Human.  
[IF=5.256] Shen C et al. In vitro culture expansion impairs chondrogenic differentiation and the therapeutic effect of mesenchymal stem cells by regulating the unfolded protein response.(2018)J Biol Eng. Nov 20;12:26.  WB&IF ;  Rabbit.  
[IF=5.223] Yunjung Lee. et al. Synergistic effect of n-3 PUFA and probiotic supplementation on bone loss induced by chronic mild stress through the brain–gut–bone axis. J FUNCT FOODS. 2023 Jan;100:105363  WB ;  Rat.  
[IF=5.201] Luo Liwen. et al. Cartilage Endplate Stem Cells Transdifferentiate Into Nucleus Pulposus Cells via Autocrine Exosomes. Front Cell Dev Biol. 2021 Mar;9:482  WB ;  Rat.  
[IF=5.179] Fang et al. Melatonin prevents senescence of canine adipose-derived mesenchymal stem cells through activating NRF2 and inhibiting ER stress. (2018) Aging.(Albany.NY). 10:2954-2974  WB ;  
[IF=5.085] Hsin-Ping Pao. et al. Suppression of Endoplasmic Reticulum Stress by 4-PBA Protects Against Hyperoxia-Induced Acute Lung Injury via Up-Regulating Claudin-4 Expression. Front Immunol. 2021; 12: 674316  WB ;  Mouse.  
[IF=5.057] Yin, Junhao. et al. CYP51-mediated cholesterol biosynthesis is required for the proliferation of CD4+ T cells in Sjogren’s syndrome. CLIN EXP MED. 2022 Nov;:1-21  WB ;  Mouse.  
[IF=4.86] Jiang, Mingfang, et al. "Down-regulation of miR-384-5p Attenuates Rotenone-induced Neurotoxicity in Dopaminergic SH-SY5Y Cells Through Inhibiting Endoplasmic Reticulum Stress." American Journal of Physiology-Cell Physiology (2016): ajpcell-00226.  WB ;  Human.  
[IF=4.784] Wang L et al. Radioprotective effect of Hohenbuehelia serotina polysaccharides through mediation of ER apoptosis pathway in vivo. Int J Biol Macromol. 2019 Apr 15;127:18-26.  IHC-P ;  Mouse.  
[IF=4.546] Junxiong Wang. et al. Protective Effects of Taraxasterol against Deoxynivalenol-Induced Damage to Bovine Mammary Epithelial Cells. Toxins. 2022 Mar;14(3):211  WB ;  Bovine.  
[IF=4.32] Yan et al. Cytotoxicity of CdTe quantum dots in human umbilical vein endothelial cells: the involvement of cellular uptake and induction of pro-apoptotic endoplasmic reticulum stress. (2016) Int.J.Nanomedicine. 11:529-42  WB ;  Human.  
[IF=4.15] Hayashi, Keitaro, et al. "HOXB9 acts as a negative regulator of activated human T cells in response to amino acid deficiency." Immunology and Cell Biology (2016).  WB ;  Human.  
[IF=4.145] Kunlin Wu. et al. Histone deacetylase inhibitor panobinostat in combination with rapamycin confers enhanced efficacy against triple-negative breast cancer. EXP CELL RES. 2022 Sep;:113362  WB ;  Human.  
[IF=4.014] Jian Chen. et al. Astragalus polysaccharide alleviates transport stress-induced heart injury in newly hatched chicks via ERS-UPR-Autophagy dependent pathway. POULTRY SCIENCE. 2022 Jun;:102030  WB ;  Chicken.  
[IF=3.858] Li,et al.Silver nanoparticles induce SH-SY5Y cell apoptosis via endoplasmic reticulum- and mitochondrial pathways that lengthen endoplasmic reticulum-mitochondria contact sites and alter inositol-3-phosphate receptor function.(2018) Toxicology Letters. 285:156-167.  WB ;  Human.  
[IF=3.414] Hou Y et al. Longzhibu disease and its therapeutic effects by traditional Tibetan medicine: Ershi-wei Chenxiang pills. J Ethnopharmacol. 2019 Nov 24:112426.  IHC-P&WB ;  Rat.  
[IF=3.21] Ming-Zhi Huang. et al. Exosomes from artesunate-treated bone marrow-derived mesenchymal stem cells transferring SNHG7 to promote osteogenesis via TAF15-RUNX2 pathway. REGEN MED. 2022 Oct 02  WB ;  Mouse, Human.  
[IF=3.184] Bingwei Yang et al. Polychlorinated Biphenyl Quinone Promotes Atherosclerosis through Lipid Accumulation and Endoplasmic Reticulum Stress via CD36. Chem Res Toxicol. 2020 Jun 15;33(6):1497-1507.  WB ;  Mouse.  
[IF=3.12] Jiang, H-Q., et al. "Guanabenz Delays the Onset of Disease Symptoms, Extends Lifespan, Improves Motor Performance and Attenuates Motor Neuron Loss in the SOD1 G93A Mouse Model of Amyotrophic Lateral Sclerosis." Neuroscience (2014).  WB ;  Mouse.  
[IF=2.87] Xiao-meng Wei. et al. Endoplasmic Reticulum Stress-Activated PERK-eIF2α-ATF4 Signaling Pathway is Involved in the Ameliorative Effects of Ginseng Polysaccharides against Cisplatin-Induced Nephrotoxicity in Mice. Acs Omega. 2021;XXXX(XXX):XXX-XXX  WB ;  Mouse.  
[IF=1.895] Zhang,et al.Ectopic expression of Miro 1 ameliorates seizure and inhibits hippocampal neurodegeneration in a mouse pilocarpine epilepsy model.(2018) Biochemistry and Cell Biology. :.  WB ;  Mouse.  
研究領域腫瘤  信號轉導  細胞凋亡  轉錄調節因子  
抗體來源Rabbit
克隆類型Polyclonal
交叉反應Rat,Mouse,Human (predicted: Sheep,Rabbit,Horse,Cow,Pig,Dog)
產品應用WB=1:500-2000, IHC-P=1:100-500, IHC-F=1:100-500, ICC=1:100, IF=1:100-500, Flow-Cyt=0.2μg /test, ELISA=1:5000-10000
not yet tested in other applications.
optimal dilutions/concentrations should be determined by the end user.
理論分子量38kDa
細胞定位細胞核 細胞漿 細胞膜 
性    狀Liquid
濃    度1mg/ml
免 疫 原KLH conjugated synthetic peptide derived from human ATF4: 251-351/351 
亞    型IgG
純化方法affinity purified by Protein A
緩 沖 液0.01M TBS(pH7.4) with 1% BSA, 0.03% Proclin300 and 50% Glycerol.
保存條件Shipped at 4℃. Store at -20 °C for one year. Avoid repeated freeze/thaw cycles.
注意事項This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications.
PubMedPubMed
產品介紹This gene encodes a transcription factor that was originally identified as a widely expressed mammalian DNA binding protein that could bind a tax-responsive enhancer element in the LTR of HTLV-1. The encoded protein was also isolated and characterized as the cAMP-response element binding protein 2 (CREB-2). The protein encoded by this gene belongs to a family of DNA-binding proteins that includes the AP-1 family of transcription factors, cAMP-response element binding proteins (CREBs) and CREB-like proteins. These transcription factors share a leucine zipper region that is involved in protein-protein interactions, located C-terminal to a stretch of basic amino acids that functions as a DNA binding domain. Two alternative transcripts encoding the same protein have been described. Two pseudogenes are located on the X chromosome at q28 in a region containing a large inverted duplication. [provided by RefSeq, Sep 2011]

Function:
Transcriptional activator. Binds the cAMP response element (CRE) (consensus: 5'-GTGACGT[AC][AG]-3'), a sequence present in many viral and cellular promoters. Cooperates with FOXO1 in osteoblasts to regulate glucose homeostasis through suppression of beta-cell production and decrease in insulin production. It binds to a Tax-responsive enhancer element in the long terminal repeat of HTLV-I. Regulates the induction of DDIT3/CHOP and asparagine synthetase (ASNS) in response to ER stress. In concert with DDIT3/CHOP, activates the transcription of TRIB3 and promotes ER stress-induced neuronal apoptosis by regulating the transcriptional induction of BBC3/PUMA. Activates transcription of SIRT4.

Subunit:
Binds DNA as a homo- or heterodimer. Interacts (via its leucine zipper domain) with GABBR1 and GABBR2 (via their C-termini). Interacts (via its DNA binding domain) with FOXO1 (C-terminal half); the interaction occurs in osteoblasts and regulates glucose homeostasis through suppression of beta-cell proliferation and a decrease in insulin production. Interacts with SATB2; the interaction results in enhanced DNA binding and transactivation by these transcription factors. Interacts with CEP290 (via an N-terminal region). Interacts with NEK6, DAPK2 (isoform 2) and ZIPK/DAPK3. Forms a heterodimer with TXLNG in osteoblasts. Interacts with DDIT3/CHOP.

Subcellular Location:
Cytoplasm. Cell membrane. Nucleus. Cytoplasm, cytoskeleton, microtubule organizing center, centrosome. Note=Colocalizes with GABBR1 in hippocampal neuron dendritic membranes. Co- localizes with NEK6 in the centrosome.

Post-translational modifications:
Ubiquitinated by SCF(BTRC) in response to mTORC1 signal, followed by proteasomal degradation and leading to down-regulate expression of SIRT4.
Phosphorylated by NEK6. Phosphorylated on the betaTrCP degron motif at Ser-219, followed by phosphorylation at Thr-213, Ser-224, Ser-231, Ser-235 and Ser-248, promoting interaction with BTRC and ubiquitination. Phosphorylation is promoted by mTORC1.
Phosphorylated by NEK6.

Similarity:
Belongs to the bZIP family.
Contains 1 bZIP (basic-leucine zipper) domain.

SWISS:
P18848

Gene ID:
468

Database links:

Entrez Gene: 468 Human

Entrez Gene: 11911 Mouse

Entrez Gene: 79255 Rat

NCBI: 33469976 Human

Omim: 604064 Human

SwissProt: P18848 Human

SwissProt: Q96AQ3 Human

SwissProt: Q06507 Mouse

SwissProt: Q5U4B2 Mouse

SwissProt: Q8CF69 Mouse

SwissProt: Q9ES19 Rat

Unigene: 496487 Human

Unigene: 641 Mouse

Unigene: 2423 Rat



產品圖片
Sample:
Lane 1: Mouse Cerebrum tissue lysates
Lane 2: Rat Cerebrum tissue lysates
Lane 3: Human HeLa cell lysates
Lane 4: Human Jurkat cell lysates
Lane 5: Human HL-60 cell lysates
Primary: Anti-ATF4 (bs-1531R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 38 kDa
Observed band size: 47 kDa
Sample:
Cerebrum (Rat) Lysate at 40 ug
Cerebrum (Mouse) Lysate at 40 ug
Primary: Anti-ATF4 (bs-1531R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 37’50 kD
Observed band size: 50 kD
Paraformaldehyde-fixed, paraffin embedded (rat skin); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human Abdominal skin); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human cervical carcinoma); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human lung carcinoma); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (rat intestine); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (mouse intestine); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human breast carcinoma); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Human 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Human skin cancer); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human brain glioma); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Human breast cancer); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructions 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human breast carcinoma); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) 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 spleen); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (mouse 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) 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 lung); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Mouse 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructions and DAB staining.
Tissue/cell: human rectal carcinoma; 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-ATF4 Polyclonal Antibody, Unconjugated(bs-1531R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
Tissue/cell: human gastric carcinoma; 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-ATF4/CREB-2 Polyclonal Antibody, Unconjugated(bs-1531R) 1:200, overnight at 4℃, 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Hela 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 (ATF4) polyclonal Antibody, Unconjugated (bs-1531R) 1:100, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.
U2OS 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 (ATF4) polyclonal Antibody, Unconjugated (bs-1531R) 1:100, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.
Blank control (blue line): Hela
Primary Antibody (green line): Rabbit Anti-Bid antibody (bs-1531R)
Dilution: 0.2μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG .
Secondary Antibody (white blue line): Goat anti-rabbit IgG-FITC
Dilution: 1μg /test.
Protocol
The cells were fixed with 70% ethanol (Overnight at 4℃) and then permeabilized with 90% ice-cold methanol for 30 min on ice. Cells stained with Primary Antibody for 30 min at room temperature. The cells were then incubated in 1 X PBS/2%BSA/10% goat serum to block non-specific protein-protein interactions followed by the antibody for 15 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.
Blank control:Mouse spleen.
Primary Antibody (green line): Rabbit Anti-ATF4 Alpha antibody (bs-1531R)
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.
Paraformaldehyde-fixed, paraffin embedded (mouse skin); 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 (ATF4) Polyclonal Antibody, Unconjugated (bs-1531R) at 1:100 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
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