MCF7 breast cancer cells were obtained from ATCC (Manassas, VA). MCF7 cells were cultured in EMEM (ATCC) supplemented with 10% FBS, 10 μg/mL insulin (Sigma I02643), 50 units/mL pencillin and 50 μg/mL Streptomycin (Invitrogen). Anti-Mouse Ig CompBead Plus (#560497), anti-Rat Ig CompBead Plus (#560499), EpCAM-PE (#347198), CD44-APC (#559942), CD49f-PE-Cy5 (#551129), Isotype-PE (#349043), Isotype-APC (#340442), Isotype PE-Cy5 (#553931) and Isotype-PE-Cy7 (#348788) antibodies were from BD Bio-sciences (San Jose, CA). CD24 PE-Cy7 (#311120) was from Biolegend (San Diego, CA). Aldefluor reagent (BAAA-DA) was synthesized by the Vahlteich Medicinal Chemistry Core (University of Michigan) from BODIPY FL succinimidyl ester (Invitrogen #D2184), purified by TLC, structure verified by HPLC, aliquoted at 50μg per micro-centrifuge tube and stored at −20°C(unpublished data). DAPI (D8417) and Live-Dead near-IR (L10119) were from Invitrogen and resuspended at 1 mg/mL in dd H2O or 200 μL DMSO, respectively. The ArC Bead Kit (A10346) was from Invitrogen. Aldefluor assay buffer (#01072) and DEAB (#01705) were from STEM-CELL Technologies (Vancouver, BC, Canada).
Bodipy fl succinimidyl ester
Manufactured by Thermo Fisher Scientific
BODIPY FL succinimidyl ester is a fluorescent dye used for protein labeling. It contains a reactive succinimidyl ester group that can form covalent bonds with primary amine groups, allowing it to be conjugated to proteins and other biomolecules. The BODIPY FL fluorophore has a green-fluorescent emission spectrum.
Automatically generated - may contain errors
Lab products found in correlation
Pencillin, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
Epcam pe, by BD (1 mentions)
Cd49f pe cy5, by BD (1 mentions)
Cd24 pe cy7, by BioLegend (1 mentions)
Aldefluor assay buffer, by STEMCELL (1 mentions)
Cd44 apc, by BD (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Insulin, by Merck Group (1 mentions)
Total e coli trna, by Roche (1 mentions)
Hitrap q hp 5 ml column, by GE Healthcare (1 mentions)
Nucleotide triphosphates, by Jena Biosciences (1 mentions)
2 protocols using bodipy fl succinimidyl ester
1
MCF7 Breast Cancer Cell Culture
2
Preparation of Fluorescent-labeled Met-tRNA for Bacterial Translation Assays
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All experiments were carried out in HiFi buffer [50 mM tris-HCl (pH 7.5), 70 mM NH4Cl, 30 mM KCl, 3.5 mM MgCl2, 8 mM putrescine, and 0.5 mM spermidine] (34 (link)). Biochemicals were from Merck, and nucleotide triphosphates were from Jena Bioscience. [3H]methionine and [14C]leucine were from Hartmann Analytics. Bodipy-FL-succinimidyl ester was from Invitrogen. Total E. coli tRNA was from Roche, and oligonucleotides were from IBA. 70S ribosomes, EF-Tu, EF-G, IF1, IF2, IF3, [3H]Met-tRNAfMet, and f[3H]Met-tRNAfMet were prepared from E. coli (35 (link)–37 (link)). Site-directed mutagenesis of the WT gene 60 construct (9 (link)) was performed using the QuikChange polymerase chain reaction protocol with the appropriate oligonucleotide primers. mRNAs were produced by T7 RNA polymerase in vitro transcription and purified by ion-exchange chromatography on a HiTrap Q HP 5-ml column (GE Healthcare). Fluorescence-labeled [3H]Met-tRNAfMet (Bpy-Met-tRNAfMet) was prepared, as previously described (38 (link)), with modifications (39 (link)).
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