Cd133 pe

Manufactured by BioLegend

Sourced in United States

CD133-PE is a fluorescently-labeled monoclonal antibody that recognizes the CD133 cell surface antigen. CD133 is a pentaspan transmembrane glycoprotein that is expressed on various stem and progenitor cell populations.

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Lab products found in correlation

Cd34 alexa647, by BD (2 mentions) Cd146 alexa488, by BD (2 mentions) Cd45 fitc, by BioLegend (2 mentions) Pd l1 pe, by BioLegend (2 mentions) Egfr fitc, by BioLegend (2 mentions) Facscalibur attune nxt flow cytometer, by Thermo Fisher Scientific (2 mentions) Cd44 pe, by BioLegend (2 mentions) Cd24 pe cy7, by BioLegend (2 mentions) Cd44 apc, by BioLegend (2 mentions) Facscanto 2, by BD (2 mentions) Lsm 510, by Zeiss (2 mentions) Igg1 apc, by BD (1 mentions) Epcam apc, by BD (1 mentions) Novocyte flow cytometer system, by Agilent Technologies (1 mentions) Paraformaldehyde (pfa), by Merck Group (1 mentions) Stain buffer, by BD (1 mentions) Cd90 fitc, by Miltenyi Biotec (1 mentions) Cd44 fitc, by Miltenyi Biotec (1 mentions) Cd86 percp cy5, by BioLegend (1 mentions) Cd206 pe dazzle594, by BioLegend (1 mentions)

Spelling variants of this product

CD133 (15 citations) Anti-CD133-PE (8 citations) PE anti-human CD133 (6 citations) PE-conjugated anti-human CD133 antibody (6 citations) PE‐conjugated anti‐CD133 antibody (5 citations) PE) anti-human CD133 antibody (4 citations) PE-conjugated CD133 antibody (3 citations) CD133-PE-Cy7 (3 citations) PE anti-mouse CD133 (2 citations) PE-Cy7 anti-mouse CD133 antibody (1 citations)

17 protocols using cd133 pe

Immunophenotypic Characterization of Cells

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To determine the cell-surface antigens, the cells at the fourth passage were subjected to staining with monoclonal antibodies specific to various proteins. Specifically, the following antibodies were used: CD90-FITC, CD45-FITC, CD133-PE, CD44-FITC, CD34-FITC, CD133-PE, and CD105-FITC (BioLegend, San Diego, CA, USA). In order to establish appropriate controls, cells were also treated with corresponding isotype control antibodies. Following staining, the cells were suspended in PBS and analyzed using a FACS Calibur flow cytometer (Becton Dickinson, NJ, USA). A minimum of 10,000 events were recorded for each sample.
First, the cells attached to the bottom of the flask were separated by trypsin/EDTA and centrifuged for 5 min at 1500 rpm at room temperature. After counting the cells, 105–106 cells were poured into the test and negative control tubes and centrifuged again for 5 min at 1500 rpm at room temperature. Subsequently, the supernatant was discarded and 100 μL of PBS were added to the pellet at the bottom of each tube and were pipetted. Then, 3 μL of specific antibodies were added to the test tubes in a dark environment. Antibodies were not added to control tubes. Hence, the tubes were incubated for 45–60 min at 4°Celsius. Finally, the expression rate of CD34, CD90, CD44, and CD105 markers were read by flow cytometry.

Vazirzadeh M., Azarpira N., Vosough M, & Ghaedi K. (2023). Galactosylation of rat natural scaffold for MSC differentiation into hepatocyte-like cells: A comparative analysis of 2D vs. 3D cell culture techniques. Biochemistry and Biophysics Reports, 35, 101503.

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Multiparametric Profiling of Stem Cell Markers

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Trypsinized (Biological Industries, Cat: BI03-052-1B) cells were collected and washed with ice-cold PBS once. Next, cells were fixed with 4% Paraformaldehyde (Sigma, Cat:158127) in PBS for 20 minutes at room temperature and centrifuged at 1500 rpm for 5 minutes; following by resuspension in stain buffer (BD, Cat: 554656) in a concentration scale of 1 × 106 cells/ml. Following antibodies were used as described; EpCAM-APC (BD, 347200) (1:100 v/v), CD133-PE (BioLegend, 372804) (1:100 v/v), CD44-FITC (Miltenyi Biotec, Cat: 130-095-195) (1:10 v/v) and CD90-FITC (Miltenyi Biotec, Cat: 130-095-403) (1:10 v/v). For unstained controls, IgG1 Isotypes; IgG1-FITC (Immunostep, Cat: ICIGG1F-100), IgG1-PE (Immunostep, Cat: ICIGG1PE-50), and IgG1-APC (BD, Cat: 555751); were used as 1:20 (v/v). For staining, cells were incubated with antibodies for 30 minutes in room temperature at dark and washed once with staining buffer. Stained cells were analyzed on NovoCyte Flow Cytometer System (Acea) and analysis was performed via NovoExpress Software (Supplementary Fig. S1).

Sahin I.D., Christodoulou M.S., Guzelcan E.A., Koyas A., Karaca C., Passarella D, & Cetin-Atalay R. (2020). A small library of chalcones induce liver cancer cell death through Akt phosphorylation inhibition. Scientific Reports, 10, 11814.

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Kidney Cell Isolation and Characterization

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Kidneys were placed into Stomacher bags containing 10 ml of RPMI buffer and ground using a Stomacher 80 Biomaster (Seward Lab Systems). The samples were centrifuged at 1,800 rpm for 10 min, and then, the cell pellet was resuspended in 40% Percoll (GE Healthcare, Sweden), placed on top of a layer of 80% Percoll and centrifuged at 3,000 rpm for 30 min (stopped without using the break braking). The cell layer located the middle of tube was collected and PBS was added for washing. The cell pellet was then resuspended in FACS buffer. The following antibodies were used for FACS: CCR7-Alexa 488, CD206-PE-Dazzle 594, CD133-PE, Ly6G-Cy7, CD68-FITC, CD86-PerCP/Cy5.5, CD169-Cy7 (Biolegend, CA, USA), CD163-PE (Invitrogen, CA, USA), CD11b-PE, CD309-PE, CD309-APC, CD31-FITC, CD29-FITC, CD105-APC, CD45-PE, CD45-APC, and Sca-1-PE (Miltenyi Bio, CA, USA). The cells were analyzed using a FACSCalibur flow cytometry system (BD Biosciences, CA, USA) and Cytomics FC 500 (Beckman Coulter, CA, USA), and data analysis was performed using FlowJo software version 10 (TreeStar, OR, USA).

Kim D.J., Moon J.Y., Kim S.M., Seo J.W., Lee Y.H., Jung S.W., Kim K., Kim Y.G., Lim S.J., Lee S., Son Y, & Lee S.H. (2020). Substance P Improves Renal Ischemia Reperfusion Injury Through Modulating Immune Response. Frontiers in Immunology, 11, 600.

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Quantifying Apoptosis in Intracranial and Subcutaneous Tumor Models

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In vivo tumor response was assessed by determining the level of apoptosis. At the indicated time, intracranial U87-luc2 tumors and subcutaneous T98G tumors were harvested, weighed, and the tumor cells isolated [6 (link)]. For the cell cycle assay, cells were fixed in cold ethanol (70% vol/vol) and stained with propidium iodide (BD Biosciences). For the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, cells were double-labeled with antibody to human CD133-PE (Miltenyi Biotec), and TUNEL using the in situ cell death detection kit (Roche Applied Science). Apoptosis was further evaluated using cCASP3 and cleaved PARP (cPARP) (Cell Signaling Technology) in combination with CD133-PE and SSEA-1-Alexa Fluor 647 (BioLegend). Samples were analyzed by BD FACS Aria flow cytometer (BD Biosciences).

Kim S.S., Rait A., Kim E., DeMarco J., Pirollo K.F, & Chang E.H. (2015). Encapsulation of temozolomide in a tumor-targeting nanocomplex enhances anti-cancer efficacy and reduces toxicity in a mouse model of glioblastoma. Cancer letters, 369(1), 250-258.

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Flow Cytometry Analysis of Cell Markers

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To determine the cell surface marker expression, cells were harvested with trypsin for flow cytometry analysis. After being blocked with 2.5% FBS, a total of 5 × 10⁵ cells were re-suspended in 200 μl PBS for each reaction. Cells were incubated on ice for 30 min with the following anti-human antibodies, which were conjugated with fluorescein phycoerythrin (PE): CD133-PE, Ki67-PE and CXCR4-PE (BioLegend, USA). Rabbit IgG1-PE (Beyotime, China) was used as the isotype control. Cells were analyzed by flow cytometry (Beckman Coulter, USA).
To determine the intracellular marker expression, cells were harvested and then fixed in a 0.5 ml/tube Fixation Buffer in the dark for 20 min at RT, followed by a 1x wash with Cell Staining Buffer. Cells were then re-suspended in Cell Staining Buffer and stored in 90% FCS/10% DMSO at − 80 °C. The fixed/permeabilized cells were re-suspended in residual Intracellular Staining Perm Wash Buffer and were added a predetermined optimum concentration of PE-conjugated antibody of Ki67 (BioLegend, USA) or rat IgG2b-PE isotype (BioLegend, USA) for 20 min in the dark at RT. Cells were analyzed by flow cytometry (Beckman Coulter, USA).

Guo X.R., Wu M.Y., Dai L.J., Huang Y., Shan M.Y., Ma S.N., Wang J., Peng H., Ding Y., Zhang Q.F., Tang J.M., Ruan X.Z, & Li D.S. (2019). Nuclear FAM289-Galectin-1 interaction controls FAM289-mediated tumor promotion in malignant glioma. Journal of Experimental & Clinical Cancer Research : CR, 38, 394.

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Quantification of CD34+CD133+CD146+ Hematopoietic Stem Cells

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BMCs were harvested freshly from mice for the experiment and cultured in DMEM high glucose media with 10% FBS. 100 ng/ml of SCF, tmSCF, tmSCFPLs, tmSCFNDs were added to the BMCs media, and cells were incubated for 30 min. Treatment was stopped by placing the cell plate on ice. The cells were stained for CD34-Alexa647 (BD Biosciences), CD133-PE (BioLegend), and CD146-Alexa488 (BD Biosciences). CD34-CD133+CD146+ cells are quantified by FlowJo software. Gating was performed similarly to the analysis of peripheral blood.

Takematsu E., Massidda M., Auster J., Chen P.C., Im B., Srinath S., Canga S., Singh A., Majid M., Sherman M., Dunn A., Graham A., Martin P, & Baker A.B. (2022). Transmembrane stem cell factor protein therapeutics enhance revascularization in ischemia without mast cell activation. Nature Communications, 13, 2497.

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EPC Quantification in Blood and Tissues

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For quantification of EPCs in the BM, circulation, and retina, fluorescence‐activated cells sorter analysis (FACS) was used as described 36. Samples were transferred into 12 mm × 75 mm polystyrene round‐bottom tubes and incubated with CD45‐FITC (no. 103108; Biolegend, San Diego, CA), CD133‐PE (no. 141204; Biolegend), CD34‐BV421 (no. 119321; Biolegend), and VEGFR2‐APC (no. 136406; Biolegend) antibodies for 1 hour in the dark. The red blood cells were lysed by adding 2 ml of red blood cell lysis buffer (BD Biosciences, Faranklin Lakes, NZ) for 15 minutes. The remaining cells were washed in 2 ml PBS containing 10% fetal bovine serum and pelleted by centrifugation at 300g for 5 minutes and finally resuspended in 250 ml paraformaldehyde (4%). The sample tubes were stored at 4°C in the dark until analysis. The samples were measured by Stratedigm S1300Ex (Stratedigm, San Jose, CA) and CD34+/CD45−/VEGFR2+/CD133+ population was analyzed by FlowJo software (Life Science Software Company, Ashland, Oregon).

Shao Y., Chen J., Freeman W., Dong L., Zhang Z., Xu M., Qiu F., Du Y., Liu J., Li X, & Ma J. (2019). Canonical Wnt Signaling Promotes Neovascularization Through Determination of Endothelial Progenitor Cell Fate via Metabolic Profile Regulation. Stem Cells (Dayton, Ohio), 37(10), 1331-1343.

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Quantifying EPC Mobilization Treatments

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To measure the ability of treatments to induce EPCs to peripheral blood, 240 µg/kg of SCF, tmSCF, tmSCFPLs, and tmSCFNDs were injected subcutaneously for consecutive four days. At the end of day 4, peripheral blood was collected for EPC marker analysis. As EPCs markers, FLK1, CD146, CD34, and CD133 were used for flow cytometry analysis. Then cells are stained for CD34-Alexa647 (BD Biosciences), CD133-PE (BioLegend), CD146-Alexa488 (BD Biosciences), and Flk-1-APC-Cy7 (BD Biosciences). An example of the gating strategy used for the studies is shown in Supplementary Fig. 14.

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Tumor Cell Immunophenotyping and Apoptosis

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The 2 × 10⁵ trypsinized tumor cells were resuspended in 100 μL of RPMI medium and incubated with fluorescent reagents (EGFR-FITC, PDL1-PE, CD133-PE, and CD44-PE antibodies, BioLegend, San Diego, CA, USA) for 30 min at room temperature. Cells were consequently added with 900 μL of PBS buffer containing 1% of FBS and analyzed using an FACSCalibur Attune NxT Flow Cytometer (Invitrogen, Waltham, MA, USA). A commercial kit containing Annexin V-FITC and Propidium Iodide was used (Strong Biotech Corporation, Taiwan) to detect cell apoptosis in tumor cells treated by 0, 10, 20 Gy of irradiation and co-cultured with healthy PBMCs.

Wang C.I., Chang Y.F., Sie Z.L., Ho A.S., Chang J.S., Peng C.L, & Cheng C.C. (2021). Irradiation Suppresses IFNγ-Mediated PD-L1 and MCL1 Expression in EGFR-Positive Lung Cancer to Augment CD8+ T Cells Cytotoxicity. Cells, 10(10), 2515.

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Assessing AhR Agonist FICZ Effects

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The affinity purified AhR antibody (SA-210) was from Enzo, CD133-PE, CD44-PerCP and CD29-FITC were from Biolegend. Antibodies for Aldh1a1 were from Becton-Dickinson and Santa Cruz Biotechnology. The antibody for β-actin was obtained from Sigma-Aldrich. Matrigel-coated transwells were from Becton-Dickinson. The iScript™ Reverse Transcription Supermix was from Bio-Rad and the SYBR® Select Master Mix for real-time PCR from Life Technologies. The AhR agonist 6-formylindolo[3,2-b]carbazole (FICZ) was from Enzo and it was used at a 5 nM concentration.

Contador-Troca M., Alvarez-Barrientos A., Merino J.M., Morales-Hernández A., Rodríguez M.I., Rey-Barroso J., Barrasa E., Cerezo-Guisado M.I., Catalina-Fernández I., Sáenz-Santamaría J., Oliver F.J, & Fernandez-Salguero P.M. (2015). Dioxin receptor regulates aldehyde dehydrogenase to block melanoma tumorigenesis and metastasis. Molecular Cancer, 14, 148.

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