EWOD device with calibrated serial dilution function

1. A method of performing dilution of a droplet in an electro-wetting on dielectric (EWOD) device comprising the steps of:

providing a parent droplet on a first portion of an electrode array of the EWOD device, wherein the parent droplet has a first concentration of a species;

providing a diluent droplet on a second portion of the electrode array of the EWOD device;

controlling actuation voltages applied to the electrode array of the EWOD device to join the parent droplet and the diluent droplet into a product droplet having a second concentration of the species different from the first concentration of the species in the parent droplet;

calibrating a dilution ratio, wherein the dilution ratio is based on a ratio of the second concentration of the species in the product droplet to the first concentration of the species in the parent droplet, by measuring a volume of the parent droplet and a volume of the diluent droplet.

2. The dilution method of claim 1 , wherein a concentration of the species in the diluent droplet is smaller than the concentration of the species in the parent droplet by a factor of one of 2, 10, 1000, or 1,000,000, or where the concentration of the species in the diluent droplet is zero.

3. The dilution method of claim 1 , wherein the diluent droplet is one of water or a buffer solution, and/or includes a surfactant.

4. The dilution method of claim 1 , wherein the species is at least one of a chemical species, a solute, a molecule or bio-molecule, a particle, or a cell.

5. The dilution method of claim 1 , wherein the EWOD device includes a sensor, and the dilution method further comprises measuring the volumes of the parent and diluent droplets with the sensor.

6. The dilution method of claim 5 , wherein the sensor is an integrated sensor that is integrated into array element circuitry of the EWOD device.

7. The dilution method of claim 6 , wherein the integrated sensor is an integrated capacitance sensor.

8. The dilution method of claim 5 , wherein the sensor is an optical sensor.

9. The dilution method of claim 1 comprising the further step of controlling the actuation voltages applied to the electrode array to split the product droplet into one or more daughter droplets, the one or more daughter droplets having the second concentration of the species.

10. The dilution method of claim 1 , wherein the dilution ratio is between 2 and 10.

11. The dilution method of claim 1 , further comprising:

measuring volumes of each of the parent droplet and the diluent droplet;

measuring a volume of the product droplet; and

outputting an error condition when the volume of the product droplet does not equal a sum of the volumes of the parent droplet and the diluent droplet.

12. The dilution method of claim 1 , further comprising:

maintaining the parent droplet at one position on the electrode array;

merging the diluent droplet into the parent droplet at the one position to form the product droplet; and

splitting the daughter droplet from the product droplet at the one position to another different position on the electrode array.

13. The dilution method of claim 1 , comprising:

performing serial dilution steps by serially repeating the method of claim 1 , wherein a daughter droplet of a dilution step is the parent droplet in a next dilution step; and

calibrating dilution ratios by measuring the volumes of the parent and diluent droplets participating in each dilution step.

14. The dilution method of claim 13 , wherein an overall dilution factor is a product of the dilution ratios at each dilution step.

15. The dilution method of claim 12 , wherein the dilution ratios are different at each dilution step, or wherein the dilution ratios are the same at each dilution step.

16. The dilution method of claim 13 wherein each serial dilution step produces a reagent droplet, wherein the reagent droplets comprise a multiplicity of droplets of different concentrations.

17. The dilution method of claim 16 , wherein the dilution factor of the multiplicity of droplets at each serial dilution step is one of a factor of two or a factor of ten.

18. A method of performing a digital assay comprising the steps of: diluting a parent droplet into one or more daughter droplets of different concentrations in accordance with the dilution method of claim 1 ;

splitting the one or more daughter droplets into a plurality of smaller reaction droplets;

performing a digitized reaction with at least one of the reaction droplets.

19. The method of claim 18 , wherein the digitized reaction of one of a nucleic acid amplification reaction, an immunoassay, or an enzyme turnover assay.

20. A method of measuring a concentration of species in a sample comprising the steps of:

inputting the sample as a parent droplet to an EWOD device, the sample having a starting concentration of the species;

diluting the sample in accordance with the dilution method of claim 1 ;

determining the dilution ratio by measuring the volumes of the parent and diluent droplets;

measuring a second concentration of the species in the daughter droplet; and

calculating the starting concentration of the species based on the second concentration and the dilution ratio.