Quantitative cell dispensing apparatus using liquid drop manipulation

Abstract

Provided is a quantitative cell dispensing apparatus using liquid droplet manipulation, for quantitatively dispensing cells from hydrophilic suspension containing a plurality of cells. The quantitative cell dispensing apparatus includes hydrophilic suspension, a liquid droplet manipulation unit, a dispensing unit, a sensing unit, and a control unit. The hydrophilic suspension includes a plurality of cells and electrolyte, and the liquid droplet manipulation unit. The liquid droplet manipulation unit includes an inlet and a liquid droplet outlet portion. The dispensing unit discharges a liquid droplet received from the liquid droplet outlet portion to a target portion. The sensing unit optically senses liquid droplets distributed on a panel of the liquid droplet manipulation unit. The control unit analyses the number of cells contained in each liquid droplet using a single sensed by the sensing unit, and controls the liquid droplet manipulation unit to transfer a liquid droplet containing a predetermined number of cells to the liquid droplet outlet portion.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

The above and other features and advantages of the present invention will become more apparent by describing in detail exemplary embodiments thereof with reference to the attached drawings in which:

FIG. 1 is a schematic perspective view of a quantitative cell dispensing apparatus using liquid droplet manipulation according to an embodiment of the present invention;

FIG. 2 is a schematic cross-sectional view of a liquid droplet manipulation unit according to an embodiment of the present invention;

FIG. 3 is a plan view illustrating portion of an electrode array of the liquid droplet manipulation unit illustrated in FIG. 2;

FIG. 4 is a plan view illustrating an inlet of a liquid droplet manipulation unit according to an embodiment of the present invention;

FIG. 5 is a cross-sectional view of a liquid droplet outlet portion of a liquid droplet manipulation unit and a dispensing unit according to an embodiment of the present invention;

FIG. 6 is a cross-sectional view of a liquid droplet outlet portion of a liquid droplet manipulation unit and a dispensing unit according to another embodiment of the present invention;

FIG. 7 is a view illustrating a liquid droplet discharging principle of the dispensing unit illustrated in FIG. 5 and 6:

FIG. 8 is a view illustrating a liquid droplet discharging process of the dispensing unit illustrated in FIG. 5 and 6;

FIG. 9A and 9B are photos illustrating suspension of cells is discharged using the dispensing unit illustrated in FIG. 5 and 6;

FIG. 10A and 10B are photos illustrating a liquid droplet discharged by the dispensing unit illustrated in FIG. 5 and 6;

FIG. 11 is a cross-sectional view illustrating an example of experiment apparatus for measuring a change in a contact angle caused by electrowetting and a critical voltage;

FIG. 12A through 12C are photos showing a change in a contact angle depending on a change in an external voltage having an influence on a sample solution;

FIG. 13 is a graph illustrating a critical voltage Vc vs. electrolyte density of a sample solution in an experiment performed by the experiment apparatus illustrated in FIG. 11; and

FIG. 14 is a graph illustrating a critical voltage Vc vs. electrolyte density of a sample solution containing E. coli in an experiment performed by the experiment apparatus illustrated in FIG. 11.

Claims

1. A quantitative cell dispensing apparatus using liquid droplet manipulation, the apparatus comprising: a hydrophilic suspension containing a plurality of cells and to which electrolyte is added;a liquid droplet manipulation unit manipulating a liquid droplet of the hydrophilic suspension on an electrode array consisting of a plurality of electrode pads using electrowetting, and including an inlet through which the hydrophilic suspension is supplied and at least one liquid droplet outlet portion discharging the manipulated liquid droplet;a dispensing unit discharging a liquid droplet received from the liquid droplet outlet portion to a target portion;a sensing unit optically sensing liquid droplets distributed on a panel of the liquid droplet manipulation unit; anda control unit analyzing the number of cells contained in each liquid droplet using a signal detected by the sensing unit and controlling the liquid droplet manipulation unit to provide liquid droplets containing a predetermined number of cells to the liquid droplet outlet portion through dividing, transferring, and merging the liquid droplets.

2. The apparatus of claim 1, wherein the electrolyte is one material selected from the group consisting of halogenide of alkali metal, alkali metal sulfate, and alkali metal nitrate.

3. The apparatus of claim 2, wherein the electrolyte is added at concentration of 1-100 mM.

4. The apparatus of claim 3, wherein the electrolyte is added at concentration of 10-30 mM.

5. The apparatus of claim 1, wherein the liquid droplet manipulation unit further includes at least one drain portion draining a liquid droplet not containing a cell.

6. The apparatus of claim 1, wherein the liquid droplet manipulation unit includes: a lower substrate;the electrode array located on the lower substrate and having a plurality of electrode pads electrically isolated from each other, the plurality of electrode pads including at least one liquid droplet outlet pad;a first film located on a surface of the electrode array and having insulation property and hydrophobicity;a transparent upper substrate facing an upper surface of the lower substrate, and located substantially in parallel to the lower substrate such that a predetermined space is provided between the lower substrate and the upper substrate;a transparent electrode located on a lower surface of the upper substrate;a second film provided on a surface of the transparent electrode to contact the predetermined space and having transparency and hydrophobicity;a non-polar fluid filling the predetermined space; andan electrode selection portion connected to each of the plurality of electrode pads, and selecting some of the electrode pads of the electrode array according to a control signal to apply a predetermined voltage thereto.

7. The apparatus of claim 6, wherein the electrode array is arranged and formed such that each of the plurality of electrode pads has an unevenness formed in its edge and thus is engaged with adjacent electrode pads.

8. The apparatus of claim 6, wherein the non-polar fluid comprises air or oil.

9. The apparatus of claim 6, wherein the transparent electrode is integrally formed over an entire surface of the transparent upper substrate.

10. The apparatus of claim 6, wherein the dispensing unit comprises: a receiving portion connected with the liquid droplet outlet pad to receive a liquid droplet containing a predetermined amount of cells; anda dispensing port provided vertically below the receiving portion to discharge the liquid droplet containing the predetermined amount of cells that has fallen down due to gravity toward the target portion.

11. The apparatus of claim 6, wherein the liquid droplet outlet pad has an upper surface covered with a layer having insulation property and hydrophilicity, or covered with an insulation layer whose surface has been hydrophilic-processed, and oil fluid is used for the non-polar fluid to prevent the non-polar fluid from penetrating a space above the liquid droplet outlet pad.

12. The apparatus of claim 6, wherein the plurality of electrode pads includes at least one drain pad, which is connected with a drain passage connecting the space with the outside.

13. The apparatus of claim 12, wherein the drain pad has an upper surface covered with a layer having insulation property and hydrophilicity, or covered with a layer whose surface has been hydrophilic-processed, and oil fluid is used for the non-polar fluid to prevent the non-polar fluid from penetrating a space above the drain pad.

14. The apparatus of claim 1, wherein the sensing unit comprises a digital camera.

15. The apparatus of claim 14, wherein the digital camera is fixed at a predetermined distance from the upper surface of the liquid droplet manipulation unit and has a lens suitable for close-up shot of the liquid droplet manipulation unit.

16. A quantitative cell dispensing apparatus using liquid droplet manipulation, the apparatus comprising: a hydrophilic suspension containing a plurality of cells and to which electrolyte is added;a liquid droplet manipulation unit manipulating a liquid droplet of the hydrophilic suspension on an electrode array consisting of a plurality of electrode pads using electrowetting, and including an inlet through which the hydrophilic suspension is supplied and at least one liquid droplet outlet portion discharging the manipulated liquid droplet;a dispensing unit discharging a liquid droplet received from the liquid droplet outlet portion to a target portion;a sensing unit optically sensing liquid droplets distributed on a panel of the liquid droplet manipulation unit; anda control unit analyzing the number of cells contained in each liquid droplet using a signal detected by the sensing unit and controlling the liquid droplet manipulation unit to provide liquid droplets containing a predetermined number of cells to the liquid droplet outlet portion through dividing, transferring, and merging the liquid droplets,wherein the dispensing unit of the quantitative cell dispensing apparatus using the liquid droplet manipulation includes:an electric field generating electrode having a vertically long syringe needle shape, an upper end connected with the liquid droplet outlet portion, an inside constituting a receiving portion receiving liquid droplets containing a cell, and a lower end where a dispensing port is formed;a target plate grounded and located below the electric field generating electrode to support a target portion that faces the dispensing port; andan open type voltage applying device applying a predetermined voltage to the electric field generating electrode to allow a liquid droplet containing a cell to be discharged to the target portion by Coulomb force originated from charges concentrated on the dispensing port and the target portion.

17. The apparatus of claim 16, wherein the dispensing unit further includes a support member supporting an upper portion of the electric field generating electrode, and the support member is fixed in the liquid droplet manipulation unit.

18. The apparatus of claim 16, wherein the open type voltage applying device applies a voltage having superposition of a DC component and an AC component.

19. The apparatus of claim 18, wherein each of the DC component and the AC component has an amplitude ranging from 5 V to 100 kV.

20. The apparatus of claim 19, wherein the frequency of the AC component is in a range of 10-1 kHz.

21. The apparatus of claim 16, wherein the target plate is formed of one material selected from the group consisting of silicon, glass, and polymer.

22. The apparatus of claim 16, wherein the electric field generating electrode has vicinity of the dispensing port that is hydrophobic-processed.

23. The apparatus of claim 16, wherein the electrolyte is one material selected from the group consisting of halogenide of alkali metal, alkali metal sulfate, and alkali metal nitrate.

24. The apparatus of claim 23, wherein the electrolyte is added at concentration of 1-100 mM.

25. The apparatus of claim 24, wherein the electrolyte is added at concentration of 10-30 mM.

26. The apparatus of claim 16, wherein the liquid droplet manipulation unit further includes at least one drain portion draining a liquid droplet not containing a cell.

27. The apparatus of claim 16, wherein the liquid droplet manipulation unit further includes: a lower substrate;an electrode array located on the lower substrate and having a plurality of electrode pads electrically isolated from each other, the plurality of electrode pads including at least one liquid droplet outlet pad;a first film located on a surface of the electrode array and having insulation property and hydrophobicity;a transparent upper substrate facing the upper surface of the lower substrate, and located substantially in parallel to the lower substrate such that a predetermined space is provided between the lower substrate and the upper substrate;a transparent electrode located on a lower surface of the upper substrate;a second film provided on a surface of the transparent electrode to contact the predetermined space and having transparency and hydrophobicity;a non-polar fluid filling the predetermined space; andan electrode selection portion connected to each of the plurality of electrode pads, and selecting some of the electrode pads of the electrode array according to a control signal to apply a predetermined voltage thereto.

28. The apparatus of claim 27, wherein the electrode array is arranged and formed such that each of the plurality of electrode pads has an unevenness formed in its edge and engaged with adjacent electrode pads.

29. The apparatus of claim 27, wherein the non-polar fluid comprises air or oil.

30. The apparatus of claim 27, wherein the transparent electrode is integrally formed over an entire surface of the transparent upper substrate.

31. The apparatus of claim 27, wherein the liquid droplet outlet pad has an upper surface covered with a layer having insulation property and hydrophilicity, or covered with an insulation layer whose surface has been hydrophilic-processed, and oil fluid is used for the non-polar fluid to prevent the non-polar fluid from penetrating a space above the liquid droplet outlet pad.

32. The apparatus of claim 31, wherein the plurality of electrode pads further includes at least one drain pad, which is connected with a drain passage connecting the space with the outside.

33. The apparatus of claim 27, wherein the plurality of electrode pads further includes at least one drain pad, which is connected with a drain passage connecting the space with the outside.

34. The apparatus of claim 33, wherein the drain pad has an upper surface covered with a layer having insulation property and hydrophilicity, or covered with a layer whose surface has been hydrophilic-processed, and oil fluid is used for the non-polar fluid to prevent the non-polar fluid from penetrating a space above the drain pad.

35. The apparatus of claim 16, wherein the sensing unit comprises a digital camera.

36. The apparatus of claim 35, wherein the digital camera is fixed at a predetermined distance from the upper surface of the liquid droplet manipulation unit and has a lens suitable for close-up shot of the liquid droplet manipulation unit.