Cell survival fraction was measured using the Trypan blue exclusion assay (Beyotime institute of Biotechnology). Cell suspension solution and Trypan Blue solution were mixed to stain non-viable cells. The viable and non-viable cells were counted under light microscope, and cell survival fraction (%) was calculated as [viable cells/(viable cells + non-viable cells)] × 100%.
Trypan blue exclusion assay
Manufactured by Beyotime
Sourced in China
The Trypan blue exclusion assay is a widely used technique for determining cell viability. It is a simple and affordable method that relies on the principle that viable cells with intact cell membranes do not take up the Trypan blue dye, while non-viable cells with compromised membranes allow the dye to enter and stain the cells blue. This assay provides a direct count of live and dead cells, enabling researchers to quantify the proportion of viable cells in a given sample.
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5 protocols using trypan blue exclusion assay
1
Trypan Blue Viability Assay
2
Bufalin Inhibits Cancer Cell Viability
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The cell viability was determined using the CCK-8 kit based on the manufacturer's instructions. BGC823, SGC7901 and GES-1 were plated at a density of 3×103 cells per well in 96-well microtiter plates and cultured overnight at 37°C in a humidified incubator containing 5% CO2. After treatment with 20, 50, 80, 100, 150 and 200 nmol/l of bufalin for 48 h, or 80 nmol/l of bufalin for 0, 1, 2, 3 and 4 days, the culture medium was replaced with 100 μl of fresh medium followed by the addition of 10 μl of CCK-8 solution. The cells were further incubated for 2 h before the optical density at 450 nm was recorded. The effect of bufalin on proliferation was assessed by Trypan Blue exclusion assay (Beyotime, Haimen, China). Trypan Blue staining was evaluated under the microscope and the number of viable cells (unstained blue) was counted using a hemocytometer.
3
Quantifying Cell Viability and Cell Cycle
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Cell viability was assessed by MTS assay using a CellTiter 96 Kit (Promega, USA) according to the manufacturer's instructions. Trypan blue exclusion assay (C0011, Beyotime, China) were performed according to the manufacturer's instructions. The percentages of viable cells were measured by Countess II (Life technologies, USA). For FACS analyses, cells were grown to approximately 80% confluence in 6-well plates, washed with cold PBS and then fixed in 70% ethanol at 4°C overnight. The cells were subsequently stained with 50 μg/mL propidium iodide (PI) supplemented with 80 μg/mL RNase A at 37°C in dark for 1 h. The cells were then subjected to FACS analyses using a FACSCalibur Low Cytometer (Becton Dickson).
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Trypan Blue Cell Viability Assay
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Trypan blue is cell membrane impermeable and only enters cells with damaged membrane. Therefore, the cell viability can be determined using trypan blue exclusion assay (Beyotime Institute of Biotechnology, Haimen, Jiangsu, China). Briefly, the cells were stained with trypan blue solution and counted using an inverted microscope (Olympus, Tokyo, Japan) within 5 minutes. The dead cells were stained with blue colour and the cell viability was calculated by the following formula: cell viability (%) = (1 − the dead cell number/the total number) × 100%.
5
Cell Viability and Antiproliferative Assays
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Cell viability was tested with MTT kit (Sigma) according to the manufacturer's instruction. The antiproliferative effect was assessed by trypan blue exclusion assay (Beyotime, Haimen, China). Trypan blue staining was evaluated under the microscope and the number of viable ells (unstained blue) was counted using a hemocytometer. For colony formation assay, a certain number of transfected cells were placed in each well of 6-well plates and maintained in proper media containing 10% FBS for two weeks, during which the medium was replaced every 4 days. Colonies were then fixed with methanol and stained with 0.1% crystal violet (Sigma) in PBS for 15 minutes. Colony formation was determined by counting the number of stained colonies.
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