A serial dilution is a chain of small, identical dilutions. It gets you from a dense culture or a concentrated standard down to a countable or measurable range without ever pipetting a tiny volume.
Why dilute in series
Getting from 10⁹ cells/mL to 100 cells/mL in one step would need a 1:10,000,000 dilution. No pipette does that. Six 1:10 steps do it with 100 µL into 900 µL each time. Each step is easy to pipette and easy to check.
Dilution factor
100 µL into 900 µL is a 1:10 dilution, factor 10. 50 µL into 50 µL is 1:2. The total dilution after n identical steps is the step factor raised to n. Six 1:10 steps give 10⁶.
"1:10" in most biology protocols means 1 part in 10 total, not 1 part plus 10 parts. A few older chemistry texts use the second meaning. If a protocol is ambiguous, check what total volume it expects.
Want more options, saved recipes and warnings? Open the full Serial dilution table in BenchMate.
Ready-made series
Tenfold series (100 µL into 900 µL)
| Tube | Dilution | From 10 mM stock |
|---|---|---|
| 1 | 10⁻¹ | 1 mM |
| 2 | 10⁻² | 100 µM |
| 3 | 10⁻³ | 10 µM |
| 4 | 10⁻⁴ | 1 µM |
| 5 | 10⁻⁵ | 100 nM |
Twofold series (for IC₅₀ and MIC plates)
Put 100 µL diluent in each well, add 100 µL of the top concentration to well 1, mix, and carry 100 µL across. Discard 100 µL from the last well so every well ends at the same volume. Eight wells span a 128-fold range. Twelve wells span 2048-fold.
Back-calculating CFU/mL
Say 10⁻⁶ gives 54 colonies from 100 µL plated. CFU/mL = 54 / (0.1 × 10⁻⁶) = 5.4 × 10⁸ CFU/mL. Count plates in the 30-300 colony range. Below that the count is noisy. Above it colonies merge.
How errors compound
Each step carries its own pipetting error, and the errors multiply down the chain. A 2% error per step becomes about 13% after six steps (1.02⁶ ≈ 1.13). Three habits keep it down:
- Change tips every step. Carry-over from a dense tube inflates every tube after it.
- Mix properly. Pipette up and down 5-8 times or vortex briefly. Unmixed tubes are the biggest source of bad series.
- Pre-wet the tip for viscous or protein-rich samples.
BenchMate builds a serial dilution table for any start concentration, step factor and number of tubes, and prints it as a bench sheet.
Frequently asked questions
How do you calculate a dilution factor?
Divide the final volume by the sample volume. 100 µL of sample in 900 µL of diluent gives 1000 µL total, a dilution factor of 10.
How do I calculate the total dilution of a serial dilution?
Multiply the dilution factors of each step. Six 1:10 steps give a total dilution of 10⁶.
How do I calculate CFU/mL from a plate count?
Divide the colony count by the volume plated in mL times the dilution. 54 colonies from 0.1 mL of a 10⁻⁶ dilution gives 5.4 × 10⁸ CFU/mL.
Which plates should I count?
Plates with roughly 30 to 300 colonies give the most reliable counts.
22 core bench techniques, from molarity to sterilisation. Volume 1 of The Biotechnology Techniques Bible.