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Fitc conjugated cd24

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FITC-conjugated CD24 is a laboratory reagent used to detect the presence and expression levels of the CD24 cell surface antigen. CD24 is a glycosylphosphatidylinositol-anchored sialoglycoprotein that is expressed on a variety of cell types. The FITC (fluorescein isothiocyanate) conjugate allows for the fluorescent labeling of CD24-positive cells, enabling their identification and analysis using flow cytometry or other fluorescence-based techniques.

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5 protocols using fitc conjugated cd24

1

CD44 and CD24 Expression Analysis

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1X 106 cells were labeled with PE-conjugated anti-CD44 (Biolegend, San Diego, USA) and FITC-conjugated CD24 (BD Pharmingen, Mississauga, Canada) for 20 min, washed twice, resuspended in PBS and analyzed on a BD FACSCalibur™ platform (Beckon Dickinson, Quebec, Canada) or MACS Quant Analyzer (Miltenyi Biotec, Cologne, Germany). Data was analyzed by FCS Express (BD Bioscience) or MACS Quantify (Miltenyi Biotec) softwares using floating quadrants to enumerate negative, single-positive and double-positive populations. CD44 and CD24 double-labeling studies always included double negative and single positive staining controls for compensation. Immunocytochemistry was performed as described before. Staining was visualized by Zeiss LSM700 confocal laser scanning microscopy system or Nikon inverted microscope.
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2

Lineage Tracing of Mammary Tumor Cells

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Intraductal injections of PNA.Met1 cells labeled with DiR
(LifeTechnologies) were carried out on age-matched, cage-matched FVB mice. Cells
were stained with the APC-conjugated lineage cocktail (Lin) (BD Biosciences)
containing CD45, Ter119, and CD31. Cells were also stained with PE-conjugated
CD49f and FITC-conjugated CD24 (BD Biosciences).
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3

Phenotypic Characterization of Colon Cancer Cells

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HT-29, LS174T and SW480 colon cancer cells were stained using FITC-conjugated CD24, PE-coniugated CD166, APC-coniugated CD133 (BD Biosciences), and PE-Cy7-conjugated CD44 (BioLegend). Samples were analyzed on a BD LSRII flow-cytometer (Bekton Dikinson, Franklin Lakes, NJ, USA). Fluorescence-activated cell sorting of HT-29 cells was performed using BD FACSAria II (Bekton Dikinson). Analysis of cytometric data was performed using FACSDiva software (Bekton Dikinson) (see Supplemental Information).
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4

Flow Cytometry Analysis of NPC Markers

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NPC surface markers were analyzed by flow cytometry as described previously [48 (link)]. After PQ treatment, cells were washed three times with PBS, and incubated on ice for 1 h in the dark with antibodies against APC-conjugated Tie-2 (FAB3131A, R&D Systems, MN, USA), PE-conjugated GD2 (562,100, BD Biosciences), and FITC-conjugated CD24 (555,427, BD Biosciences). For the isotype control, the antibody was replaced with IgG1 mouse APC (IM2475, Beckman Coulter, CA, USA), IgG1 mouse PE (A07796, Beckman Coulter), and IgG1 mouse FITC (A07795, Beckman Coulter). Only viable cells were analyzed using the PI-negative gate.
For ECM analysis, cells were fixed and permeabilized with IntraPrep Permeabilization Reagent (A07803, Beckman Coulter). Cells were then subjected to three cycles of freezing (−80 °C) and thawing (37 °C). Subsequently, cells were incubated overnight at 4 °C with primary antibodies against collagen II (NB600-844, Novus Biologicals, CO, USA) and proteoglycan (MAB2015, Sigma-Aldrich). After washing with PBS, cells were reacted with goat anti-mouse Alexa Flour 488-conjugated secondary antibody (A28175, Invitrogen) on ice for 1 h in the dark, and then analyzed by flow cytometry.
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5

Pancreatic Cancer Cell Immunophenotyping

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Human pancreatic cancer cells were dissociated with trypsin, washed twice with HBSS containing 2% fetal bovine serum and 10 mM HEPES (staining buffer). Cells were stained and incubated on ice for 30 minutes with the following primary antibodies: ABCG2 (R&D Systems Inc, 1:50), FITC-conjugated CD24 (BD Biosciences, 1:100), APC-conjugated CD44 (BD Biosciences, 1:100), biotinylated SNA lectin (Vector Laboratories, 1:100), or MALII lectin (Vector Laboratories, 1:100). After stained with appropriated secondary antibody, the stained cells were analyzed by FACSCalibur (BD Biosciences).
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