Hanging droplet plate

Patent metadata

Field	Value
Publication number	US-9267933-B2
Application number	US-201214002269-A
Country	US
Kind code	B2
Filing date	Mar 2, 2012
Priority date	Mar 3, 2011
Publication date	Feb 23, 2016
Grant date	Feb 23, 2016

Abstract

A hanging droplet plate ( 1 ) comprises a predetermined number of droplet compartments ( 10 ) each being capable of receiving a droplet of a liquid. The respective droplet compartment ( 10 ) comprises a circumferential microfluidic wetting barrier ( 102 ) which is arranged to surround a respective cavity ( 100 ) and which prevents a droplet from spreading beyond the microfluidic wetting barrier ( 102 ). The respective compartment ( 10 ) comprises a closed bottom ( 101 ) and at least one additional circumferential microfluidic wetting barrier ( 104 ), each additional circumferential microfluidic wetting barrier ( 104 ) which is arranged to surround a preceding circumferential microfluidic wetting barrier ( 102 ). A wettable area ( 103 ) is arranged between two adjacently arranged microfluidic wetting barriers ( 102, 104 ).

First claim

The invention claimed is:

1. A hanging droplet plate comprising:
a predetermined number of droplet compartments each being capable of receiving a droplet of a liquid, each droplet compartment having a cavity and a circumferential microfluidic wetting barrier surrounding the cavity such that a droplet is prevented from spreading beyond the circumferential microfluidic wetting barrier, wherein
each droplet compartment comprises a separate and distinct closed bottom within the circumference of the circumferential microfluidic wetting barrier and at least one additional circumferential microfluidic wetting barrier, each at least one additional circumferential microfluidic wetting barrier surrounding a preceding circumferential microfluidic wetting barrier, with a wettable area being arranged between two adjacently arranged microfluidic wetting barriers.

2. The hanging droplet plate according to claim 1 , wherein each of the droplet compartments comprise a well.

3. The hanging droplet plate according to claim 1 , wherein the circumferential microfluidic wetting barrier comprises a circumferential edge, the at least one additional circumferential microfluidic wetting barrier comprises at least one additional circumferential edge, each at least one additional circumferential edge being arranged to surround a preceding circumferential edge, with the wettable area being arranged between two adjacently arranged circumferential edges.

4. The hanging droplet plate according to claim 3 , wherein two adjacent circumferential edges are arranged in a stepped manner.

5. The hanging droplet plate according to claim 1 , wherein the circumferential microfluidic wetting barrier comprises a circumferential rim, the at least one additional microfluidic wetting barrier comprises an additional circumferential rim, each additional circumferential rim surrounding a preceding circumferential rim with the wettable area being arranged between two adjacently arranged circumferential rims.

6. The hanging droplet plate according to claim 5 , wherein the adjacently arranged circumferential rims are arranged in a stepped manner.

7. The hanging droplet plate according to claim 2 , wherein the hanging droplet plate is made of a separately manufactured plate having a predetermined number of holes therein and of a corresponding predetermined number of separately manufactured wells forming the droplet compartments, with each separately manufactured well being press-fitted into a respective hole of the separately manufactured plate.

8. The hanging droplet plate according to claim 7 , wherein the separately manufactured wells are press-fitted into the holes from the side of the plate forming the outer surface of the plate.

9. The hanging droplet plate according to claim 7 , comprising 24 wells, 96 wells or 384 wells.

10. The hanging droplet plate according to claim 5 , wherein the circumferential rims are arranged on a plane surface.

11. A hanging droplet plate assembly, comprising:
a hanging droplet plate according to claim 1 , the plate having a predetermined number of droplet compartments; and
a receiving plate having a number of wells corresponding to the predetermined number of droplet compartments of the hanging droplet plate, the hanging droplet plate and the receiving plate being assembled in a manner such that in the assembled state the wells of the receiving plate are aligned with the droplet compartments of the hanging droplet plate.

12. A method of testing a substance for its toxicity to cells, comprising:
introducing a predetermined number of liquid droplets into a corresponding number of droplet compartments of a hanging droplet plate, each droplet containing a predetermined volume of the substance to be tested and of a liquid culture medium as well as a plurality of cells;
inverting and incubating the hanging droplet plate for a predetermined time interval with the hanging droplet plate carrying the droplets in a manner such that they hang down from the respective droplet compartments to allow the cells to form three-dimensional cellular aggregates in the respective droplets;
supplying additional liquid culture medium to the droplets in the respective droplet compartments to promote additional growing of the three-dimensional cellular aggregates; and
analyzing the three-dimensional cellular aggregates in order to assess whether the substance to be tested is toxic to the three-dimensional cellular aggregates, wherein a hanging droplet plate according to any one of the preceding claims is used; and wherein supplying the additional liquid culture medium step is performed by re-inverting the hanging droplet plate, adding a droplet of the additional liquid culture medium to the respective droplets containing the three-dimensional cellular aggregates so as to form respective larger droplets in the respective droplet compartments, and then inverting the hanging droplet plate again to allow the three-dimensional cellular aggregates to grow in the respective larger droplets hanging down from the droplet compartments.

13. The method according to claim 12 , further comprising:
transferring the grown three-dimensional cellular aggregates from the droplet compartments of the hanging droplet plate into a corresponding number of wells of a receiving plate;
incubating the receiving plate with the wells containing the three-dimensional cellular aggregates for a further predetermined time interval; and
after incubation, analyzing the three-dimensional cellular aggregates in order to assess whether the substance to be tested is toxic to the three-dimensional cellular aggregates.

14. The method according to claim 13 , wherein the transferring step is performed by assembling the hanging droplet plate and the receiving plate in a manner such that the respective wells of the receiving plate are arranged opposite to the respective droplet compartments of the hanging droplet plate, and by subsequent centrifugation of the assembled plates.

15. The method according to claim 13 , wherein the cells are embryonic stem cells and the three-dimensional cellular aggregates are embryoid bodies, and wherein the step of analyzing the three dimensional cellular aggregates to assess whether the substance to be tested is toxic to the three-dimensional cellular aggregates is performed by analyzing whether the embryoid bodies contain myocardial cells.

Assignees

• Fattinger Christof
• Iaiza Patrick
• Kissling Tom
• Voegelin Dieter
• Zumstein Thomas
• Mcginnis Claudia
• Hoffmann La Roche

Inventors

• Fattinger Christof
• Iaiza Patrick
• Kissling Tom
• Voegelin Dieter
• Zumstein Thomas
• Mcginnis Claudia

Classifications

• B01L2300/0829

  Multi-well plates; Microtitration plates · CPC title

• B01L3/06

  Crystallising dishes · CPC title

• C12M25/01

  Drops · CPC title

• B01L2300/042

  Caps; Plugs · CPC title

• B01L3/50853

  with covers or lids (closures for test tubes B01L3/50825) · CPC title