Plasmid MapTurn your cell count into a concentration
Record the four large corner squares in one chamber, or all eight across two chambers. Choose live and dead separately if you recorded both populations. Choose total cells only if you did not distinguish them. A blank means the observation is missing; enter 0 when you counted a square and found no cells.
This calculator uses a 1 mm × 1 mm square and a 0.1 mm chamber depth. Each large square therefore samples 0.1 µL, or 0.0001 mL. Include all the subdivisions within that large square. The same conversion does not apply to a smaller central square or a chamber with a different depth.
Hemocytometer formula and the 10⁴ factor
Cells/mL = cells counted ÷ large squares counted × total dilution factor × 10⁴.
The factor 10⁴ converts one large square's 0.0001 mL volume into a per-milliliter concentration. With separate live and dead counts, the main result uses live cells. Total and dead-cell concentrations are shown alongside it. With two chambers, the calculator pools all eight equally sized squares.
For a published arithmetic check, the STEMCELL Technologies counting protocol gives four counts of 21, 15, 20 and 17, with a dilution factor of 100. Their mean is 18.25 cells per square. Multiplying 18.25 × 100 × 10⁴ gives 18,250,000 cells/mL in the original suspension. This verifies the arithmetic; it is not a recommended count density or dilution for every sample.
Include the full dilution factor
The factor describes final counting-mixture volume divided by the original sample volume in that mixture. Mixing equal volumes of sample and stain gives a factor of 2, not 1. If the sample was diluted tenfold first and then mixed with an equal volume of stain, the combined factor is 10 × 2 = 20.
The initial field shows 2 for an equal-volume mixture. Change it to match your actual preparation; use 1 for undiluted material. The result estimates the concentration before these counting dilutions. An optional original suspension volume converts that concentration into an estimated number of cells available.
Cell viability: live divided by live plus dead
Viability (%) = live cells ÷ (live cells + dead cells) × 100. The same dilution factor applies to both populations and cancels out of this ratio. For 200 live cells and 10 dead cells, viability is approximately 95.24%. Under a trypan-blue exclusion protocol, intact cells exclude the dye while cells with compromised membranes take it up; follow your laboratory's identification and counting method.
If every recorded count is zero, the observed concentration is zero and viability is undefined because its denominator is zero. An empty observed sample does not establish that the original suspension contains no cells. Total-only counts cannot establish viability either.
Plan the next cell dilution
Choose Add a dilution plan, then enter a target concentration in cells/mL and a final volume in mL. The plan uses V₁ = C₂ × V₂ / C₁. When live and dead counts are separate, C₁ and C₂ refer to viable cells. Otherwise they refer to total cells.
For a viable concentration of 1,000,000 cells/mL, a target of 200,000 viable cells/mL and a final volume of 2 mL, take 400 µL of suspension and add 1.6 mL of medium. The final dilution contains 400,000 viable cells under the model's assumptions. If you enter an available suspension volume, the tool checks that the required transfer fits within it. A target above the measured concentration cannot be reached by adding medium.
This is volume arithmetic, assuming representative counts, thorough mixing, additive volumes and no cell loss. It does not choose a seeding density, culture conditions or a counting protocol, and it does not estimate measurement uncertainty. Save the text calculation or Worksheet CSV to retain your original square counts, dilution factor, chamber geometry, results and assumptions. All inputs and exports stay in your browser.
For solution concentrations in molar or mass units, use the dilution calculator. For successive dilution steps and retained volumes, use the serial dilution calculator. The molarity calculator handles preparation from a solute mass and final volume.