i, j Quantification of percentage of ( i) ALDH1A1-positive and ( j) β-catenin-positive cells compared to parental BM-MSCs by intracellular flow cytometry staining. For comparison, isotype control (black) used to define the positive and negative population for each marker. g, h Flow cytometry overlay histogram analysis of ( g) ALDH1A1 and ( h) β-catenin in BM-MSCs and MCF7, Hela, and HepG2 CiSCs. f Confocal immunofluorescence images for β-catenin (green) of control BM-MSCs and MCF7, Hela, and HepG2 CiSCs. e Confocal immunofluorescence images for ALDH1A1 (green) of control BM-MSCs and MCF7, Hela, and HepG2 CiSCs. d Flow cytometry overlay histogram analysis of CD24 expression in BM-MSCs and MCF7, Hela, and HepG2 CiSCs. c Flow cytometry plots for cell surface markers CD44 and CD24 in BM-MSCs and MCF7, Hela, and HepG2 CiSCs. Data reported on a log-10 scale as mean ± SD. β-actin mRNA used to normalize variability in template loading. b Real-time qRT-PCR analysis of CSC marker genes. a Expression levels of mRNAs encoding KRAS, HER2, CDK4, BRCA2, E2F3, SMAD7, ABCB1, APC, and TP53 in MCF7, Hela, and HepG2 CiSCs relative to the parental BM-MSCs determined by real-time qRT-PCR. 566385).Expression of cancer and CSC markers in CiSCs and side population (SP) properties of CiSCs.
563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. Performing fewer than the recommended wash steps may lead to increased spread of the negative population.įor optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes are used in the same experiment. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescence staining, prior to acquisition on a flow cytometer. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and CompBead to ensure that BD Comp beads are appropriate for your specific cellular application.įor optimal results, it is recommended to perform 2 washes after staining with antibodies. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. When fluorochrome conjugated antibodies are bound to CompBeads, they have spectral properties very similar to cells. BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation).
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