50 000 viable B16-F10 cells (trypan blue exclusion test) dissociated from one-week-old tumorspheres or from trypsinized adherent monolayers were incubated for 30 minutes at 4°C in the dark with 0.5 µg of rat anti-mouse monoclonal CD133-APC, CD44-FITC or CD24-PE antibodies (eBiosciences, Paris, France) in 100 µL of a buffer solution consisting in PBS containing 3% bovine serum albumin (Sigma). Immunostaining was also performed for HT-29, MCF-7 and MDA-MB-231 cell lines using mouse anti-human monoclonal CD133-APC, CD44-PE, CD44-APC or CD24-FITC antibodies (Miltenyi Biotec, Paris, France) according to the manufacturer’s instructions. Cells were then washed and analyzed with an Accuri C6 flow cytometer (BD biosciences, USA).
Cd133 apc
Manufactured by Thermo Fisher Scientific
Sourced in United States
CD133-APC is a fluorescently-labeled antibody that targets the CD133 cell surface antigen. It is used for the identification and isolation of CD133-positive cells through flow cytometry or cell sorting applications.
Automatically generated - may contain errors
Lab products found in correlation
Cd31 fitc, by BioLegend (2 mentions)
Cd45 percp, by BioLegend (2 mentions)
Hoechst 33342 stain, by Merck Group (2 mentions)
Cd146 pe, by BioLegend (2 mentions)
Lsr 2, by BD (2 mentions)
Goat anti rabbit igg alexa fluor 700, by Thermo Fisher Scientific (2 mentions)
Cd44 fitc, by Thermo Fisher Scientific (1 mentions)
Cd44 pe, by Miltenyi Biotec (1 mentions)
Cd133 apc, by Miltenyi Biotec (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Cd24 pe, by Thermo Fisher Scientific (1 mentions)
Accuri c6 flow cytometer, by BD (1 mentions)
Cd44 apc, by Miltenyi Biotec (1 mentions)
Cd24 fitc, by Miltenyi Biotec (1 mentions)
Ng2 chondroitin sulfate proteoglycan, by Merck Group (1 mentions)
Cd140a, by BioLegend (1 mentions)
Ab5320, by Merck Group (1 mentions)
5 protocols using cd133 apc
1
Flow Cytometry Analysis of Stem Cell Markers
2
Multiparametric Cytometry of Stem Cells
3
Flow Cytometry Analysis of Cell Markers
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Flow cytometry was performed as described by Buono et al. (2015) (link) using the following antibodies specified in Tables S3 and S4 ). Lewis-X (1/20; BD Biosciences), CD133-APC (1/50; eBioscience), CD140a (1/400; BioLegend), and NG2 chondroitin sulfate proteoglycan (1/50; Millipore). EdU experiments were performed as described previously (Chen et al., 2015 ). Goat anti-rabbit IgG Alexa Fluor 700 (1/100; Invitrogen) was used for NG2. DAPI was used for dead cell exclusion. IL-6Rα was analyzed using the flow panel described above with the addition of an antibody against IL-6Rα (1/20, R&D Systems). All sample data were collected on a BD LSR II (BD Biosciences) and analyzed using FlowJo.
4
Characterization of Endothelial Progenitor Cells
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The antibodies CD31‐FITC, CD45‐PerCP, and CD146‐PE were purchased from BioLegend (San Diego, CA, USA) and CD133‐APC from eBiosciences (San Diego, CA, USA). Hoechst 33342 stain was obtained from Sigma (St. Louis, MO, USA). The ELISA kits used were ST2 (DST200; R&D Systems Quantikine®, Minneapolis, MN, USA), VCAM1, ICAM1, and TIM3 (DY809, DY720, and DY2365; R&D Systems Duosets®, Minneapolis, MN, USA) and REG3α (Cat. No. 5323; Ab‐Match Assembly Human PAP1 [REG3α] kit, MBL International Corp, Japan).
5
Characterization of Stem Cell Subpopulations
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Spheres were dissociated and counted as previously described (Buono et al., 2012) (see supplemental methods). For surface marker analysis, cells were incubated in PGB for 25 min with antibodies against Lewis-X (1:20, LeX/CD15 FITC, MMA; BD Bioscience), CD133-APC (1:50,13A4; eBioscience), CD140a (1:400, APA5; BioLegend) and NG2 Chondroitin Sulfate Proteoglycan (1:50, AB5320; Millipore). Goat anti-rabbit IgGAlexa Fluor 700 (1:100; Invitrogen) was used for NG2. All sample data were collected on the BD LSR II (BD Biosciences Immunocytometry Systems). Post-acquisition analysis was performed using FlowJoX (Tree Star Inc, Ashland, OR).
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