How It Works
This calculator works in 3 steps.
Enter the starting concentration
Type the highest concentration in the series, such as 128 µg/mL.
Enter steps and well volume
Type the number of dilutions and the volume left per well, such as 100 µL.
Read the series
The calculator halves the concentration at each step and lists cumulative DF from 2 to 2ⁿ.
Follow the transfer plan
Transfer a volume equal to the diluent volume at each step, then discard the same volume from the last well.
The Formula
The formula divides the starting concentration by 2 raised to the step number.
Cₙ = C₀ / 2ⁿn = log₂(C₀ / C_target)V_t = V_d (equal volumes)- Cₙ:
- Concentration after n halving steps
- C₀:
- Starting concentration
- n:
- Step number
- V_t / V_d:
- Transfer and diluent volumes
Starting at 128 µg/mL with 100 µL broth per well, transfer 100 µL from well to well. Well 8 holds 128 / 2⁸ = 0.5 µg/mL.
Conversion Table
The conversion table lists 9 concentrations from 128 µg/mL with the matching DF and ratio.
| Step | Cumulative DF | Ratio | Concentration |
|---|---|---|---|
| 0 | 1 | Neat | 128 µg/mL |
| 1 | 2 | 1:2 | 64 µg/mL |
| 2 | 4 | 1:4 | 32 µg/mL |
| 3 | 8 | 1:8 | 16 µg/mL |
| 4 | 16 | 1:16 | 8 µg/mL |
| 5 | 32 | 1:32 | 4 µg/mL |
| 6 | 64 | 1:64 | 2 µg/mL |
| 7 | 128 | 1:128 | 1 µg/mL |
| 8 | 256 | 1:256 | 0.5 µg/mL |
What Is the Twofold Serial Dilution Calculator?
The Twofold Serial Dilution Calculator is a tool that halves the concentration at every step and returns the concentration and volumes for each tube or well. Each step mixes equal volumes of the previous dilution and diluent, so tube n holds 1/2ⁿ of the starting concentration.
Twofold series are the standard format for minimum inhibitory concentration (MIC) testing, hemagglutination assays, and antibody titers. The Clinical and Laboratory Standards Institute (CLSI) broth microdilution method uses twofold steps, such as 128, 64, 32 down to 0.125 µg/mL. Ten twofold steps cover a 1,024-fold range.
Common Uses
There are 5 main uses of this calculator.
MIC panels
Clinical microbiologists test antibiotic activity on twofold broth series.
Antibody titers
Serologists dilute serum 1:2, 1:4, 1:8 to find the last positive dilution.
Hemagglutination assays
Virologists titrate influenza virus stocks in twofold steps.
Checkerboard rows
Researchers create twofold axes for drug combination plates.
Dose ranging
Toxicologists start a range-finding study with twofold spacing.