Tc20 cell counter

Manufactured by Bio-Rad

Sourced in United States

The TC20 cell counter is a compact and automated instrument designed for quickly and accurately counting cells. It utilizes a microfluidic chip and imaging technology to provide cell count and viability data. The TC20 is a versatile tool suitable for a variety of cell types and applications in the life sciences research and biopharmaceutical industries.

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TC20 automated cell counter (941 citations) Cell counter (74 citations) TC20 automatic cell counter (49 citations) TC20TM (27 citations) TC20 counter (16 citations) TC20 automated cell counter systems (2 citations) TC20 cell counter slide (2 citations) TC20 Automated Cell Counter with Trypan Blue reagent (2 citations)

170 protocols using tc20 cell counter

Evaluating small-molecule inhibitors in mouse ESCs

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For hairpin-transduced v6.5 mouse ESCs, 100K cells per well were plated on a gelatin-coated 12-well plate 2 days after puromycin selection. Cells were counted using Trypan Blue exclusion method on the TC20 Cell Counter (Biorad) after 2 or 4 days. Cells were passaged at day 2 to inhibit contact-induced differentiation.
For the small-molecule inhibitor experiments, v6.5 mouse ESCs were plated at 100K cells per well on a gelatin-coated 12-well plate. Twenty-four hours later, the cells were treated with either vehicle (DMSO) or varying concentrations of the following inhibitors: I-BRD9, BI-9564, PFI-3, or TP472. Cells were counted using Trypan Blue exclusion method on the TC20 Cell Counter (Biorad) every 48 h, during which the cells were re-plated at the same cell density on a new 12-well plate.
Two-tailed t-test was performed to obtain the p values from biological replicates, n = 9 for I-BRD9, n = 6 for BI-9564 and TP472, n = 3 for PFI-3, and n = 3 for each shBrd9 hairpin. *p < 0.05, **p < 0.01, ***p < 0.001.

Gatchalian J., Malik S., Ho J., Lee D.S., Kelso T.W., Shokhirev M.N., Dixon J.R, & Hargreaves D.C. (2018). A non-canonical BRD9-containing BAF chromatin remodeling complex regulates naive pluripotency in mouse embryonic stem cells. Nature Communications, 9, 5139.

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Whole Blood Cell Isolation Protocols

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Whole blood samples preserved in SmartTube solution were kept on ice for 15 minutes, followed by thawing in a water bath set to 20°C. Red blood cells were lysed by addition of SmartTube Lysis buffer #1 and incubated at 20°C for 10 minutes. The white blood cells were washed twice with CyFACS (1% BSA, 0.5% Na-Azide and 0.5M EDTA in PBS), filtered through a 35μm mesh and counted using a Bio-Rad TC20 cell counter.
Whole blood samples preserved in Cytodelics Whole Blood Cell Stabilizer were thawed at 20°C. Cytodelics Fix/Lyse buffer was added at a blood:buffer concentration of 1:10 and samples were incubated at 20°C for 5 minutes. Samples were then diluted 1:4 with Cytodelics Wash buffer #1 and left to lyse for 15 minutes. Cells were then washed twice with Cytodelics Wash buffer #2, filtered through a 35μm mesh and counted using a Bio-Rad TC20 cell counter.

Olin A., Henckel E., Chen Y., Lakshmikanth T., Pou C., Mikes J., Gustafsson A., Bernhardsson A.K., Zhang C., Bohlin K, & Brodin P. (2018). Stereotypic Immune System Development in Newborn Children. Cell, 174(5), 1277-1292.e14.

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Cell Nuclei Preparation and Mixing for Paired-seq

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HEK293T (human, ATCC CRL-11268), HepG2 (human, ATCC HB-8065) and NIH/3T3 (murine, ATCC CRL-1658) cells were cultured according to standard procedures in Dulbecco’s Modified Eagles’ Medium (Thermo Fisher Scientific, 10569010) supplemented with 10% fetal bovine serum (FBS, Thermo Fisher Scientific, 16000044) and 1% penicillin–streptomycin (Thermo Fisher Scientific, 10378016) at 37 °C with 5% CO₂. Cells were not authenticated nor tested for mycoplasma. To prepare nuclei, HepG2 and 3T3 cells were harvested by centrifugation, washed with PBS (Thermo Fisher Scientific, 10010-23) and counted using BioRad TC20 cell counter. The percentage of live cells in the samples were higher than 95%. The cells were then resuspended in cold Lysis Buffer (10 mM Tris-HCl pH 7.4 [Sigma, T4661], 10 mM NaCl [Sigma, S7653], 3 mM MgCl₂ [Sigma, 63069], 0.1% IGEPAL CA-630 [Sigma, I8896]) and centrifuged for 15 min at 600 g, 4 °C. For the species mixing experiment, nuclei were then washed with PBS and resuspended, counted using BioRad TC20 cell counter. HepG2, HEK293T and 3T3 nuclei were then mixed in equal proportions and applied to Paired-seq.

Zhu C., Yu M., Huang H., Juric I., Abnousi A., Hu R., Lucero J., Behrens M.M., Hu M, & Ren B. (2019). An ultra high-throughput method for single-cell joint analysis of open chromatin and transcriptome. Nature structural & molecular biology, 26(11), 1063-1070.

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Maintenance of hiPSC and wild-type stem cells

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For maintenance, HiTCA and non-gene edited wild type (WT) SC cells were cultured in media composed of 88% Gibco RPMI (Thermo Fisher Scientific, Waltham, MA, USA), 10% FBS, 1% Sodium Pyruvate (100 mM), and 1% Pen-Strep antibiotic. The cell cultures were propagated in 25 cm² canted neck cell flasks with vented caps at a density between 2.0 × 10⁶ and 2.0 × 10⁷ cells/mL. The flasks were incubated at 37 °C and 5% CO₂. While maintaining the number of cells in culture, they were passaged every 2–3 days by decanting half of the culture media and adding an equal volume of fresh culture media. Cells were tested periodically for viability with trypan blue and counted with a Bio-Rad TC20 Cell Counter (Bio-Rad Laboratories, Hercules, CA, USA) to ensure viability over 90%.

Gottlieb C., Henrich M., Liu P.T., Yacoubian V., Wang J., Chun R, & Adams J.S. (2023). High-Throughput CAMP Assay (HiTCA): A Novel Tool for Evaluating the Vitamin D-Dependent Antimicrobial Response. Nutrients, 15(6), 1380.

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Microfluidic Viable Cell Separation

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An epifluorescence microscope (IX83, Olympus, Japan), connected with a CCD camera (QIClick, QImaging, Canada), and controlled by Olympus cellSens Dimension™ software, was used to observe and record the motion of the beads or cell separation process, and to take images of beads or cells inside the ViaChip. A three-channel peristaltic pump (EW-78001–70, Cole-Parmer, USA) was used to control the fluidic system. In all studies in this paper, the flow rates pumping to two inlets were 25 μL/min. Several other flow rates were also investigated, and the cell size distribution under different flow rates is shown in Figure S2. We found that 25 μL/min is the optimal flow rate in our microfluidic system for viable cell separation. Dulbecco’s phosphate-buffered saline (DPBS, 21–031-CV, Corning, USA) was used as diluent and buffer inlet when analyzing beads in the ViaChip, while for cell or mouse-derived bone marrow samples, we used cell culture media in the buffer inlet. All tubes connected to inlets or outlets were incubated with cell medium for 10 minutes before sample loading to reduce non-specific binding. The number of cells or polystyrene beads was visualized under the microscope and counted by a BioRad TC20 cell counter (BioRad, USA).

Yeh P.Y., Snijders A.M, & Wang D. (2021). ViaChip for Size-based Enrichment of Viable Cells. Sensors and actuators. B, Chemical, 353, 131159.

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Quantifying Cell Proliferation with NDRG4 Expression

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FLO-1 and JH-eso-ad1 cells were infected with 10 MOI control or NDRG4 expression adenoviral particles or transfected with PcDNA control or NDRG4 expression plasmids, followed by selection under 600 mg/mL G418 for two weeks. In total, 5 × 10⁴ cells per well in the 6-well plates were seeded and the cell numbers were counted at 24 h interval using trypan blue exclusion assay and Bio-rad TC20 Cell Counter (Bio-Rad). The experiments were repeated in triplicates and each sample was counted three times. Then, the readings were averaged for each sample.

Cao L., Hu T., Lu H, & Peng D. (2020). N-MYC Downstream Regulated Gene 4 (NDRG4), a Frequent Downregulated Gene through DNA Hypermethylation, plays a Tumor Suppressive Role in Esophageal Adenocarcinoma. Cancers, 12(9), 2573.

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Metastatic Appendiceal Cancer: Single-Cell Dissociation

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Patients with metastatic appendiceal cancer were consented on IRB protocol 44036 approved by Stanford University. Tumor tissues were obtained from surgery and stored in RPMI medium before dissociation. Single-cell suspensions were obtained from tissue fragments using enzymatic and mechanical dissociation. Cells were washed twice in RPMI + 10% fetal bovine serum (FBS), filtered through 70 μm (Flowmi, Bel-Art SP Scienceware, Wayne, NJ), followed by 30 μm (Miltenyi) or 40 μm filter (Flowmi). Cryofrozen cells were rapidly thawed in a bead bath at 37°C followed by above washing and filtering steps. Live cell counts were obtained on a Bio-Rad TC20 Cell Counter (Bio-Rad, Hercules, CA) or a Countess II FL Automated Cell Counter (Thermo Fisher Scientific) using 1:1 trypan blue dilution. Cells were concentrated between 500 and 1500 live cells/μl for subsequent single-cell library preparation. The patient with follicular lymphoma was consented on IRB protocol 13500 approved by Stanford University. Fine needle aspirate specimens from two spatially separated nodal tumor sites were obtained and prepared as previously described (11 (link)).

Grimes S.M., Kim H.S., Roy S., Sathe A., Ayala C.I., Bai X., Almeda-Notestine A.F., Haebe S., Shree T., Levy R., Lau B.T, & Ji H.P. (2023). Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer. NAR Cancer, 5(3), zcad034.

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Single-cell RNA Sequencing of Tumor Samples

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This study was conducted in compliance with the Helsinki Declaration. All patients were enrolled according to a study protocol approved by the Stanford University School of Medicine Institutional Review Board (IRB-11886 and IRB-44036). Written informed consent was obtained from all patients.
Tissue samples came from surgical resections or matched normal tissue from sites displaced at least several centimeters from the tumor. Tissues were collected in plain RPMI on ice immediately after resection and dissected with iris scissors. Single-cell suspensions were obtained from tissue fragments using enzymatic and mechanical dissociation and from peripheral blood using peripheral blood mononuclear cell (PBMC) isolation as described previously (10 (link)). Briefly, cells were washed twice in RPMI + 10% FBS, filtered through 70 μm (Flowmi, Bel-Art SP Scienceware), followed by 40-μm filter (Flowmi). Cryofrozen cells were rapidly thawed in a bead bath at 37°C followed by above washing and filtering steps. Live-cell counts were obtained on a Bio-Rad TC20 cell counter (Bio-Rad) or a Countess II FL Automated Cell Counter (Thermo Fisher Scientific) using 1:1 trypan blue dilution. Cells were concentrated between 500 and 1,500 live cells/μL for scRNA-seq.

Sathe A., Mason K., Grimes S.M., Zhou Z., Lau B.T., Bai X., Su A., Tan X., Lee H., Suarez C.J., Nguyen Q., Poultsides G., Zhang N.R, & Ji H.P. (2022). Colorectal Cancer Metastases in the Liver Establish Immunosuppressive Spatial Networking between Tumor-Associated SPP1+ Macrophages and Fibroblasts. Clinical Cancer Research, 29(1), 244-260.

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Cell Viability Assay with OmpC-CD154

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The cell viability was determined using cell lines treated with the OmpC-CD154 protein by trypan blue staining (Invitrogen). The cell lines were cultured at a density of 1 Â 10 6 cells/well in 1 mL of culture medium in 12-well, flat-bottom cell culture microplates. The treated cells were incubated with the OmpC, OmpC-CD154 or CD154rh proteins at a concentration of 2 lg/ml for 12, 18, 36 or 48 h at 37 C in a 5% CO 2 incubator. The percentage of viable cells was determined using a Bio-Rad TC20 cell counter (Bio-Rad, Hercules CA).

Pantoja-Escobar G., Morales-Martínez M., Vega G.G., Castro-Escarpulli G, & Vega M.I. (2019). Cytotoxic effect caspase activation dependent of a genetically engineered fusion protein with a CD154 peptide mimetic (OmpC-CD154_p) on B-NHL cell lines is mediated by the inhibition of bcl-6 and YY1 through MAPK p38 activation. Leukemia & lymphoma, 60(4).

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Neuro-2a Cell Preparation for Lipid Extraction

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Neuro-2a cells were seeded at 1.25 × 10⁶ cells/dish in a 10 cm dish. One day after seeding, medium was aspirated and cells were cultured in 2% serum and vehicle (0.1% DMSO). After 48 h, cells were washed with PBS and harvested by resuspension in PBS. Cells were pelleted (200 g, 5 min, rt) and resuspended in PBS. Cell count and viability were measured by Trypan blue staining and automated cell counting (TC20^TM Cell Counter, Bio-Rad) and 1 × 10⁶ cells were pelleted (1000 g, 3 min, rt). Pellets were flash frozen in liquid nitrogen and stored at −80°C until lipid extraction.

van Esbroeck A.C., Kantae V., Di X., van der Wel T., den Dulk H., Stevens A.F., Singh S., Bakker A.T., Florea B.I., Stella N., Overkleeft H.S., Hankemeier T, & van der Stelt M. (2019). Identification of α,β-Hydrolase Domain Containing Protein 6 as a Diacylglycerol Lipase in Neuro-2a Cells. Frontiers in Molecular Neuroscience, 12, 286.

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