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[KO验证抗体] Rabbit Anti-Vimentin antibody (bs-0756R)
~~~促销,代码KT2401~~~
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产品编号 bs-0756R
英文名称 [KO验证抗体] Vimentin
中文名称 波形蛋白抗体
别    名 FLJ36605; OTTHUMP00000019224; VIM; VIME_HUMAN; Vimentin.  
Specific References  (106)     |     bs-0756R has been referenced in 106 publications.
[IF=18.952] Mahmoud Labibet al. Tracking the expression of therapeutic protein targets in rare cells by antibody-mediated nanoparticle labelling and magnetic sorting. Nat Biomed Eng . 2020 Jul 27.  FCM ;  Human.  
[IF=12.822] Tian-Hao Wang. et al. Connexin-43 is a promising target for lycopene preventing phthalate-induced spermatogenic disorders. J ADV RES. 2022 Sep;:  WB,IF ;  Mouse.  
[IF=11.161] Zhipeng Jiang. et al. EIF4A3-induced circ_0084615 contributes to the progression of colorectal cancer via miR-599/ONECUT2 pathway. J Exp Clin Canc Res. 2021 Dec;40(1):1-15  WB,IHC ;  Human.  
[IF=10.633] Na Yu. et al. Comparison of fresh and preserved decellularized human corneal lenticules in femtosecond laser-assisted intrastromal lamellar keratoplasty. ACTA BIOMATER. 2022 Jul;:  IF ;  Human.  
[IF=9.584] Zheng, Bingxin. et al. Siglec-15-induced autophagy promotes invasion and metastasis of human osteosarcoma cells by activating the epithelial–mesenchymal transition and Beclin-1/ATG14 pathway. CELL BIOSCI. 2022 Dec;12(1):1-15  WB ;  Human.  
[IF=8.806] Monteiro et al. Dental cell sheet biomimetic tooth bud model. (2016) Biomaterials. 106:167-79  IHC-P,IHC-P-IF ;  Porcine.  
[IF=8.469] Que, Tianshi. et al. HMGA1 stimulates MYH9-dependent ubiquitination of GSK-3β via PI3K/Akt/c-Jun signaling to promote malignant progression and chemoresistance in gliomas. Cell Death Dis. 2021 Dec;12(12):1-12  WB ;  Human.  
[IF=8.39] Monteiro, Nelson, et al. "Dental Cell Sheet Biomimetic Tooth Bud Model."Biomaterials (2016).  IF(IHC-P) ;  Pig.  
[IF=7.666] Yufang Liu. et al. Effect of Upregulation of Transcription Factor TFDP1 Binding Promoter Activity Due to RBP4 g.36491960G>C Mutation on the Proliferation of Goat Granulosa Cells. CELLS-BASEL. 2022 Jan;11(14):2148  IF ;  Goat.  
[IF=7.43] Wang, Jing, et al. "Phosphorylation-dependent regulation of ALDH1A1 by Aurora kinase A: insights on their synergistic relationship in pancreatic cancer." BMC biology 15.1 (2017): 10.  WB ;  Human.  
[IF=7.419] Hao Wu. et al. Rutin ameliorates gout via reducing XOD activity, inhibiting ROS production and NLRP3 inflammasome activation in quail. BIOMED PHARMACOTHER. 2023 Feb;158:114175  IF ;  Quail.  
[IF=6.691] Jain, Drishti. et al. Prognostic significance of NT5E/CD73 in neuroblastoma and its function in CSC stemness maintenance. 2021 Nov 13  IF ;  Human.  
[IF=6.639] Fatima-Zohra Khadri. et al. Impact of Epithelial–Mesenchymal Transition on the Immune Landscape in Breast Cancer. Cancers. 2021 Jan;13(20):5099  IHC ;  Human.  
[IF=6.491] Nikhil K et al. Identification of LIMK2 as a Therapeutic Target in Castration Resistant Prostate Cancer. Cancer Lett. 2019 Apr 28;448:182-196.  WB ;  Mouse.  
[IF=6.304] Zhongjian Yu. et al. Circ-HMGA2 (hsa_circ_0027446) promotes the metastasis and epithelial-mesenchymal transition of lung adenocarcinoma cells through the miR-1236-3p/ZEB1 axis. Cell Death Dis. 2021 Mar;12(4):1-13  WB ;  Human.  
[IF=6.304] Peixin Du. et al. Chronic stress promotes EMT-mediated metastasis through activation of STAT3 signaling pathway by miR-337-3p in breast cancer. Cell Death Dis. 2020 Sep;11(9):1-13  WB ;  Mouse.  
[IF=6.025] Limei Yang. et al. The disruption of human trophoblast functions by autophagy activation through PI3K/AKT/mTOR pathway induced by exposure to titanium carbide (Ti3C2) MXene. FOOD CHEM TOXICOL. 2022 May;:113128  WB ;  Human.  
[IF=6.023] Ling Xie. et al. Suppression of GOLM1 by EGCG through HGF/HGFR/AKT/GSK-3β/β-catenin/c-Myc signaling pathway inhibits cell migration of MDA-MB-231. Food Chem Toxicol. 2021 Nov;157:112574  WB ;  human.  
[IF=5.696] Jiang-Tao Fan. et al. Exosomal lncRNA NEAT1 from cancer-associated fibroblasts facilitates endometrial cancer progression via miR-26a/b-5p-mediated STAT3/YKL-40 signaling pathway. Neoplasia. 2021 Jul;23:692  IF ;  Human.  
[IF=5.682] Xiaolan You. et al. Fibroblastic galectin-1-fostered invasion and metastasis are mediated by TGF-β1-induced epithelial-mesenchymal transition in gastric cancer. Aging-Us. 2021 Jul 31; 13(14): 18464–18481  WB,IHC ;  Mouse.  
[IF=5.68] Karim, A. S., et al. "Nox2 Is a Mediator of Ischemia Reperfusion Injury."American Journal of Transplantation (2015).  WB ;  Mouse.  
[IF=5.673] Bott CJ et al. Nestin Selectively Facilitates the Phosphorylation of the Lissencephaly-Linked Protein Doublecortin (DCX) by cdk5/p35 to Regulate Growth Cone Morphology and Sema3a Sensitivity in Developing Neurons. J Neurosci. 2020 May 6;40(19):3720-3740.  WB IHC IF ;  Human.  
[IF=5.646] Ye T et al. MicroRNA-7 as a potential therapeutic target for aberrant NF-κB-driven distant metastasis of gastric cancer. J Exp Clin Cancer Res. 2019 Feb 6;38(1):55.  FCM&IHC ;  Mouse&Human.  
[IF=5.211] Jin, Meiyuan. et al. MicroRNA-3935 promotes human trophoblast cell epithelial-mesenchymal transition through tumor necrosis factor receptor-associated factor 6/regulator of G protein signaling 2 axis. Reprod Biol Endocrin. 2021 Dec;19(1):1-15  WB ;  human.  
[IF=5.03] Wilkinson, Dan C., et al. "Development of a Three-Dimensional Bioengineering Technology to Generate Lung Tissue for Personalized Disease Modeling." Stem Cells Translational Medicine (2016): sctm-2016.  IHC-P ;  Human.  
[IF=5.01] Wang, Li-Ping, et al. "Angiotensin II upregulates K Ca 3.1 channels and stimulates cell proliferation in rat cardiac fibroblasts." Biochemical pharmacology 85.10 (2013): 1486-1494.  Rat.  
[IF=5.008] Gao et al. Hes1 promotes cell proliferation and migration by activating Bmi-1 and PTEN/Akt/GSK3β pathway in human colon cancer. (2015) Oncotarge. 6:38667-80  WB ;  Human.  
[IF=4.784] Zheng Wu. et al. FOXD3 suppresses epithelial–mesenchymal transition through direct transcriptional promotion of SMAD7 in esophageal squamous cell carcinoma. 2021 Sep 22  WB ;  human.  
[IF=4.76] Khayat et al. GelMA-Encapsulated hDPSCs and HUVECs for Dental Pulp Regeneration. (2017) J.Dent.Res. 96:192-199  IF(ICC) ;  Human.  
[IF=4.72] Sucre, Jennifer MS, et al. "A three-dimensional human model of the fibroblast activation that accompanies bronchopulmonary dysplasia identifies Notch-mediated pathophysiology." American Journal of Physiology-Lung Cellular and Molecular Physiology 310.10 (2016): L889-L898.  IF(IHC-P) ;  Human.  
研究领域 肿瘤  细胞生物  免疫学  发育生物学  神经生物学  信号转导  干细胞  细胞骨架  肿瘤细胞生物标志物  
抗体来源 Rabbit
克隆类型 Polyclonal
交叉反应 Human, Mouse, Rat,  (predicted: Chicken, Pig, Cow, Goat, )
产品应用 WB=1:500-2000 ELISA=1:5000-10000 IHC-P=1:100-500 IHC-F=1:100-500 Flow-Cyt=1μg/Test ICC=1:100 IF=1:100-500 (石蜡切片需做抗原修复)
not yet tested in other applications.
optimal dilutions/concentrations should be determined by the end user.
理论分子量 53kDa
细胞定位 细胞浆 细胞外基质 
性    状 Liquid
浓    度 1mg/ml
免 疫 原 KLH conjugated synthetic peptide derived from human Vimentin: 371-466/466 
亚    型 IgG
纯化方法 affinity purified by Protein A
缓 冲 液 0.01M TBS(pH7.4) with 1% BSA, 0.03% Proclin300 and 50% Glycerol.
保存条件 Shipped at 4℃. Store at -20 °C for one year. Avoid repeated freeze/thaw cycles.
注意事项 This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications.
PubMed PubMed
产品介绍 This gene encodes a member of the intermediate filament family. Intermediate filamentents, along with microtubules and actin microfilaments, make up the cytoskeleton. The protein encoded by this gene is responsible for maintaining cell shape, integrity of the cytoplasm, and stabilizing cytoskeletal interactions. It is also involved in the immune response, and controls the transport of low-density lipoprotein (LDL)-derived cholesterol from a lysosome to the site of esterification. It functions as an organizer of a number of critical proteins involved in attachment, migration, and cell signaling. Mutations in this gene causes a dominant, pulverulent cataract.[provided by RefSeq, Jun 2009]

Function:
Vimentins are class-III intermediate filaments found in various non-epithelial cells, especially mesenchymal cells. Vimentin is attached to the nucleus, endoplasmic reticulum, and mitochondria, either laterally or terminally.
Involved with LARP6 in the stabilization of type I collagen mRNAs for CO1A1 and CO1A2.

Subunit:
Homopolymer assembled from elementary dimers. Interacts with HCV core protein. Interacts with LGSN and SYNM. Interacts (via rod region) with PLEC (via CH 1 domain) (By similarity). Interacts with SLC6A4. Interacts with STK33. Interacts with LARP6. Interacts with RAB8B (By similarity).

Subcellular Location:
Cytoplasm.

Tissue Specificity:
Highly expressed in fibroblasts, some expression in T- and B-lymphocytes, and little or no expression in Burkitt's lymphoma cell lines. Expressed in many hormone-independent mammary carcinoma cell lines.

Post-translational modifications:
Filament disassembly during mitosis is promoted by phosphorylation at Ser-55 as well as by nestin (By similarity). One of the most prominent phosphoproteins in various cells of mesenchymal origin. Phosphorylation is enhanced during cell division, at which time vimentin filaments are significantly reorganized. Phosphorylation by PKN1 inhibits the formation of filaments. Phosphorylated at Ser-56 by CDK5 during neutrophil secretion in the cytoplasm. Phosphorylated by STK33.

Similarity:
Belongs to the intermediate filament family.

SWISS:
P08670

Gene ID:
7431

Database links:

Entrez Gene: 7431 Human

Entrez Gene: 22352 Mouse

Entrez Gene: 81818 Rat

Omim: 193060 Human

SwissProt: P08670 Human

SwissProt: P20152 Mouse

SwissProt: P31000 Rat

Unigene: 455493 Human



Vimentin—波形蛋白。是五种主要的中间丝之一,存在于各种正常和病理性间质来源的组织,如纤维母细胞、内皮细胞、淋巴细胞等正常细胞和肉瘤、淋巴瘤、黑色素瘤等肿瘤。波形蛋白是负责维持细胞骨架完整性的蛋白之一。
产品图片
Sample:
Hela(Human) Cell Lysate at 30 ug
Hela KO Vimentin (Human) Cell Lysate at 30 ug
Primary: Anti- Vimentin (bs-0756R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 53 kD
Observed band size: 53 kD
Sample:
Cerebrum (Mouse) Lysate at 40 ug
Primary: Anti-Vimentin (bs-0756R) at 1/2000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 53 kD
Observed band size: 53 kD
Sample:
Lane 1: Hela (Human) Cell Lysate at 30 ug
Lane 2: NIH/3T3 (Mouse) Cell Lysate at 30 ug
Lane 3: U251 (Human) Cell Lysate at 30 ug
Lane 4: SH-SY5Y (Human) Cell Lysate at 30 ug
Lane 5: A549 (Human) Cell Lysate at 30 ug
Primary: Anti-Vimentin (bs-0756R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 55 kD
Observed band size: 57 kD
Sample: Jurkat (human)Cell Lysate at 40 ug
Primary: Anti- Vimentin (bs-0756R) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 53 kD
Observed band size: 53 kD
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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 ovary); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 kidney); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 colon 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 kidney); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 tonsil); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 kidney); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 colon); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 ovary); 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 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 cervical 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) at 1:400 overnight at 4°C, followed by a conjugated secondary (sp-0023) for 20 minutes and DAB staining.
Tissue/cell: mouse kidney 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-Vimentin Polyclonal Antibody, Unconjugated(bs-0756R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
Paraformaldehyde-fixed, paraffin embedded (human cervical 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) at 1:400 overnight at 4°C, followed by a conjugated secondary (sp-0023) for 20 minutes and DAB staining.
Tissue/cell: U-87MG 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 1:100, 90 minutes at 37°C; followed by a 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.
Tissue/cell: 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 1:50, 90 minutes at 37°C; followed by a 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.
Tissue/cell: U251 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 1:50, 90 minutes at 37°C; followed by a 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.
Tissue/cell: U-2OS 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 (Vimentin) Polyclonal Antibody, Unconjugated (bs-0756R) 1:50, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody (bs-0295G-FITC) at 37°C for 90 minutes, DAPI (5ug/ml, blue, C-0033) was used to stain the cell nuclei.
Tissue/cell: mouse mesenchymal stem cells;
Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
Incubation: Anti-Vimentin Polyclonal Antibody, Unconjugated(bs-0756R) 1:300, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
Blank control: Jurkat cells(blue).
Primary Antibody:Rabbit Anti-Vimentin antibody antibody(bs-0756R), 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-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA.
Protocol
The cells were fixed with 2% paraformaldehyde (10 min) , then permeabilized with 90% ice-cold methanol for 30 min on ice. Primary antibody (bs-0756R, 1μg /1x10^6 cells) were incubated for 30 min on the ice, 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/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed.
Blank control:A549.
Primary Antibody (green line): Rabbit Anti-Vimentin antibody (bs-0756R)
Dilution: 1μg /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.
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