Cell Viability Percentage Calculator
Percent cell viability from trypan blue live/dead counts, automated-counter concentrations or an MTT/MTS/resazurin plate assay — with viable and dead cells per mL, total viable cells, a 95% confidence range, seeding volume and replicate statistics.
How it works
Viability % = live ÷ (live + dead) × 100 · cells/mL = cells counted ÷ squares × dilution × 10⁴ · total viable cells = viable cells/mL × volume · Plate assay: viability % = (treated − blank) ÷ (control − blank) × 100, inhibition = 100 − viability · 95% range: Wilson score interval on live ÷ counted
Trypan blue enters cells with damaged membranes, so blue cells are dead and clear ones are alive. Viability is the live share of all cells counted. Each large haemocytometer square holds 0.1 µL, so the count per square times the dilution times 10⁴ gives cells per millilitre — for 1:1 with trypan blue the dilution is 2. If an automated counter or flow cytometer gave you live and dead concentrations instead, switch to “Concentrations” and the units menu takes them as cells/mL or × 10⁴, 10⁵ or 10⁶. For a 96-well MTT, MTS, XTT, resazurin or ATP assay, paste the treated and untreated-control readings and the medium blank: viability is the blank-corrected treated signal as a percentage of the control, with the spread between replicate wells. Counts carry sampling error — with 200 cells counted the 95% range on a 92% viability is about 87.4–95.0%, which the calculator shows so you know whether a small drop is real. Count within a few minutes of mixing, because trypan blue slowly stains live cells too, and aim for 100–400 cells in total.
Worked example
184 live + 16 dead over 4 squares with a 1:1 trypan blue mix: viability = 184 ÷ 200 = 92.0%; viable cells = 184 ÷ 4 × 2 × 10⁴ = 0.92 × 10⁶ per mL, so 9.2 × 10⁶ in 10 mL; the 95% range is 87.4–95.0%. To seed 1 × 10⁶ viable cells take 1.09 mL. In a plate assay, treated wells reading 0.62, 0.65 and 0.60 against controls of 0.95, 0.98 and 0.93 with a 0.05 blank give (0.623 − 0.05) ÷ (0.953 − 0.05) = 63.5% viability, 36.5% inhibition.
Sources: Thermo Fisher Gibco — Trypan blue exclusion protocol · AAT Bioquest — Trypan blue dye exclusion assay
Frequently asked questions
What viability is needed for experiments?
Over 90% for most assays, passaging and transfections; at least 95% for healthy log-phase cultures, flow cytometry and single-cell work. After thawing, 70–90% is common for the first passage.
How do I calculate percent viability from an MTT assay?
Subtract the blank (medium plus reagent, no cells) from every reading, then divide the mean treated value by the mean untreated control and multiply by 100. Choose “Plate assay”, paste the replicate readings and the calculator gives viability, inhibition and the replicate spread.
How accurate is a trypan blue viability count?
It is a sample, so it carries counting error: with 100 cells the 95% range is roughly ±6 percentage points at 90% viability, with 400 cells about ±3. Count more squares or both chambers when a few points matter.
Automated counter gives a different number?
Automated counters size-gate debris and clumps differently from the eye, so they rarely match a manual count exactly. Keep to one method within an experiment, and enter the counter’s live and dead concentrations under “Concentrations”.
Why can an MTT result differ from trypan blue?
MTT and similar assays measure metabolic activity, while trypan blue tests membrane integrity. A treatment that slows metabolism lowers the MTT signal without killing cells, and dying cells can still exclude dye for a while — so the two numbers answer different questions.
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For teaching and planning — verify against your lab protocol and instrument documentation.