Journal: Journal of visualized experiments : JoVE

Article Title: Tumorsphere derivation and treatment from primary tumor cells isolated from mouse rhabdomyosarcomas

doi: 10.3791/59897

Figure Lengend Snippet: Boscolo et al. Table of Materials

Article Snippet: ​ Name of Material/Equipment Company Catalog Number Comments/Description DMEM/F12 Media Gibco 11320033 Component of tumosphere media DMEM high glucose media Gibco 11965092 Component of tumor cells media Ham’s F10 Media Life Technologies 11550043 Component of FACS buffer Penicillin - Streptomyocin Life Technologies 15140163 Component of tumosphere and tumor cells media Fetal Bovine Serum Omega Scientific FB-11 Component of tumor cells media Goat Serum Life Technologies 16210072 Component of FACS buffer Recombinant Human βFGF-basic Peprotech 10018B Component of tumosphere media EGF recombinant mouse protein Gibco PMG8041 Component of tumosphere media N-2 Supplements (100X) Gibco 17502048 Component of tumosphere media PBS Gibco 10010023 Component of FACS buffer and used for washing cells Fluriso (Isofluornae) anesthetic agent MWI Vet Supply 502017 Anesthetic reagent for animals Accutase cell dissociation reagent Gibco A1110501 Detach adherent cells and dissociate tumorspheres Matrigel membrane matrix Corning CB40234 Provides support to trasplanted cells Horse Serum Life Technologies 16050114 Component of cell isolation media Collagenase, Type II Life Technologies 17101015 Tissue digestion enzyme Dispase II, protease Life Technologies 17105041 Tissue digestion enzyme EDTA ThermoFisher S312500 Component of FACS buffer FxCycle™ Violet Stain Life Technologies F10347 Discriminate live and dead cells Trypan blue ThermoFisher 15250061 Discriminate live and dead cells Lipofectamine 3000 transfection reagent ThermoFisher L3000015 Transfection Reagent Recombinant mouse Flt-3 Ligand Protein R&D Systems 427-FL-005 Recombinant protein pEGFP-C1 Addgene 6084–1 GFP plasmid Celigo Nexcelom Celigo Microwell plate based image cytometer for adherent and suspension cells FACSAria II Flow Cytometry BD Biosciences 650033 Fluorescent activated cell sorter Neomycin-Polymyxin B Sulfates-Bacitracin Zinc Ophthalmic Ointment MWI Vet Supply 701008 Eyes ointment Open in a separate window Boscolo et al. Table of Materials

Techniques: Recombinant, Cell Isolation, Staining, Transfection, Plasmid Preparation, Cytometry, Flow Cytometry

Journal: Biology

Article Title: Identification of a Unique Cytotoxic Thieno[2,3-c]Pyrazole Derivative with Potent and Selective Anticancer Effects In Vitro

doi: 10.3390/biology11060930

Figure Lengend Snippet: The intrinsic apoptosis pathway mediates Tpz-1-induced cell death. ( a ) Analysis of phosphatidylserine externalization by flow cytometry showed apoptosis induction in HL-60 cells after 24 h of exposure to CC 50 (0.95 μM; p < 0.00001) and 2xCC 50 (1.9 μM; p < 0.00001) concentrations of Tpz-1. The percentages displayed represent total (early and late) apoptosis values. ( b ) Flow cytometry analysis indicated that Tpz-1 CC 50 (0.95 μM; p = 0.00646) and 2xCC 50 (1.9 μM; p = 0.005941) concentrations induced significant caspase-3/7 activation after 6 h of incubation in HL-60 cells. Positive fluorescence for NucView 488 Caspase-3/7 substrate indicated caspase-3/7 activation. ( c ) Tpz-1 causes reactive oxygen species accumulation in HL-60 cells at CC 50 (0.95 μM; p = 0.000139) and 2xCC 50 (1.9 μM; p < 0.00001) concentrations. ROS was quantified using the carboxy-H 2 DCFDA reagent. ( d ) Tpz-1 disrupted mitochondrial membrane potential in HL-60 cells after 5 h of incubation. A modest but significant increase in depolarized mitochondria was seen in cells treated with Tpz-1 2xCC 50 (1.9 μM; p = 0.029193). Mitochondrial depolarization was quantified using the JC-1 dye; an increase in green fluorescent signal denoted loss of membrane potential. ( a – d ) Cells were treated with the 24 h CC 50 (0.95 μM) or 2xCC 50 (1.9 μM) for the HL-60 cell line, and 1% v / v DMSO or 1 mM H 2 O 2 as vehicle and positive controls for cytotoxicity, respectively. Untreated (UNT) controls were also included. The percentages and standard deviations represent the average of three technical replicates. Representative density plots for each experiment are shown. Statistical analysis was achieved by two-tailed Student’s paired t -test, and the asterisk annotations represent the statistical significance of the treatments against the vehicle control; (*) p < 0.05, (**) p < 0.01, (***) p < 0.001.

Article Snippet: Therefore, to further validate that apoptosis is the cell death mechanism of Tpz-1, caspase-3/7 activation (NucView 488 Caspase-3/7 Substrate Assay Kit, Biotium) was measured in compound-treated HL-60 acute myeloid leukemia and CCRF-CEM T cell leukemia cells [ ].

Techniques: Flow Cytometry, Activation Assay, Incubation, Fluorescence, Membrane, Two Tailed Test

Journal: Cell Reports Medicine

Article Title: DNAJB1-PRKACA fusion neoantigens elicit rare endogenous T cell responses that potentiate cell therapy for fibrolamellar carcinoma

doi: 10.1016/j.xcrm.2024.101469

Figure Lengend Snippet:

Article Snippet: NucView 530 caspase-3 substrate , Biotium , 10408.

Techniques: Blocking Assay, Recombinant, Plasmid Preparation, Protease Inhibitor, Cell Stimulation, Binding Assay, Multiplex Assay, Clone Assay, Software