Miniaturized thermal cycler

• US 20020173032A1
• Filed: 07/03/2002
• Published: 11/21/2002
• Est. Priority Date: 02/16/2001
• Status: Active Grant

• Current Assignee: Agency For Science Technology and Research
• Original Assignee: Agency For Science Technology and Research
• Inventors: Chen, Yu, Sridhar, Uppili, Yan, Tie, Lim, Tit Meng, Zou, Quanbo, Zachariah, Emmanuel Selvanayagam
• Granted Patent: US 6,521,447 B2
• Time in Patent Office: Days
• US Class Current: 435/287.2
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0877 Flow chambers
• CPC Class Codes: B01L 2300/1827 using resistive heater
• CPC Class Codes: B01L 2300/1883 using thermal insulation
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/0688 surface tension valves, capillary stop, capillary break
• CPC Class Codes: B01L 3/5025 for parallel transport of multiple samples
• CPC Class Codes: B01L 3/5027 by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: B01L 7/52 with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046

Abstract

The invention describes a thermal cycler which permits simultaneous treatment of multiple individual samples in independent thermal protocols, so as to implement large numbers of DNA experiments simultaneously in a short time. The chamber is thermally isolated from its surroundings, heat flow in and out of the unit being limited to one or two specific heat transfer areas. All heating elements are located within these transfer areas and at least one temperature sensor per heating element is positioned close by. Fluid bearing channels that facilitate sending fluid into, and removing fluid from, the chamber are provided. The chambers may be manufactured as integrated arrays to form units in which each cycler chamber has independent temperature and fluid flow control Two embodiments of the invention are described together with a process for manufacturing them.

Claims

1. A thermal cycling unit, comprising:

a chamber, thermally isolated from its surroundings except for one or more heat transfer areas through which all heat that flows in and out of the chamber passes;

within the chamber, at least one heating element per transfer area, each such heating element being located within a transfer area,

a first fluid bearing channel that connects to the chamber through a first orifice located within a transfer area;

a second fluid bearing channel that connects to the chamber through a collection orifice located within a transfer area;

within the chamber, at least one temperature sensor per heating element located close to that heating element; and

means for sending fluid into, and removing fluid from, the chamber through said channels and orifices.

2. The thermal cycling unit described in claim 1 wherein there is a plurality of chambers each of which can be independently heated and cooled.

3. The thermal cycling unit described in claim 1 wherein there is a plurality of chambers and thermal cross-talk between the chambers is less than about 0.5°

C. at temperatures ranging from about 20 to 95°

C.

4. A structure for thermal cycling, comprising:

a frame made of a thermally conductive material and having an open interior area,

suspended within said open area, a plurality of chambers in the form hollow bodies having first and second opposing ends each of which is connected to the frame through a thermally conductive beam;

within each chamber, at each end, a heat transfer area through which all heat that flows in and out of the chamber passes;

within each chamber, two heating elements, one each symmetrically disposed around one each of said transfer areas;

for each chamber, a first fluid bearing channel that enters the chamber at its first end through a first orifice located within the transfer area at that end;

for each chamber, a second fluid bearing channel that enters the chamber at its second end through a second orifice located within the transfer area at that end;

within each chamber, at least one temperature sensor per heating element, said sensor being located close to said heating element, and

means for sending fluid into, and removing fluid from, each chamber through said channels and orifices whereby fluid flow into and out of each chamber is individually controllable.

5. The structure described in claim 4 wherein the frame is thermally connected to a heat sink.

6. The structure described in claim 4 wherein the frame and the beams are thermally conductive materials selected from the group consisting of monocrystalline silicon germanium and gallium arsenide, metals, and ceramics.

7. The structure described in claim 4 wherein each chamber further comprises a silicon membrane between about 30 and 100 microns thick surrounded by sidewalls that have been anodically bonded to a sheet of glass, whereby the chamber has low thermal capacitance.

8. The structure described in claim 4 wherein the heating elements in each chamber are independently controllable.

9. The structure described in claim 4 wherein the said fluid bearing channels are located inside the beams.

10. The structure described in claim 4 wherein said means for sending fluid into each chamber further comprises a source of compressed gas connected to the first channel, said compressed gas causing liquid to flow into the chamber from a common reservoir..

11. The structure described in claim 10 wherein said means for removing fluid from each chamber further comprises a local reservoir into which gas from the chamber is forced when, under pressure from said compressed gas or from hydraulic or pneumatic interaction with a gas-liquid interface at the valve, the liquid fills the chamber.

12. The structure described in claim 4 wherein the fluid bearing channels include at least one pressure barrier capable of stopping both hydrophillic and hydrophobic liquid flow.

13. The structure described in claim 12 wherein said pressure barrier further comprises a section of the fluid-bearing channel that is narrower than other parts of the channel.

14. The structure described in claim 4 wherein each chamber has an interior volume that is less than about 100 micro-liters.

15. The structure described in claim 4 wherein thermal cross-talk between the chambers is less than about 0.5°

C. at temperatures ranging from about 20 to 95°

C.

17. The structure described in claim 16 wherein the frame is thermally connected to a heat sink.

18. The structure described in claim 16 wherein the frame and the beam are monocrystalline silicon.

19. The structure described in claim 16 wherein each chamber further comprises a silicon membrane between about 30 and 100 microns thick, surrounded by sidewalls that have been anodically bonded to a sheet of glass, whereby the chamber has low thermal capacitance.

20. The structure described in claim 16 wherein the heating elements in each chamber are independently controllable

21. The structure described in claim 16 wherein the said fluid bearing channels are located inside the beam.

22. The structure described in claim 16 wherein said means for sending fluid into each chamber further comprises a source of compressed gas connected to the first channel, said compressed gas causing liquid to flow into the chamber from a common reservoir

23. The structure described in claim 22 wherein said means for removing fluid from each chamber further comprises a local reservoir into which gas from the chamber is forced when, under pressure from said compressed gas, the liquid fills the chamber.

24. The structure described in claim 16 wherein the fluid bearing channels include at least one pressure barrier capable of stopping both hydrophillic and hydrophobic liquid flow.

25. The structure described in claim 24 wherein said pressure barrier further comprises a section of the fluid-bearing channel that is narrower than other parts of the channel.

26. The structure described in claim 16 wherein each chamber has an interior volume that is less than about 100 micro-liters

27. The structure described in claim 16 wherein thermal cross-talk between the chambers is less than about 0.5°

C. at temperatures ranging from about 20 to 95°

C.

29. The process described in claim 28 wherein, at the start of the process the silicon wafer has a thickness between about 350 and 700 microns.

30. The process described in claim 28 wherein said first trench depth is between about 0.1 and 1 microns, said inner trenches'"'"' first width is between about 20 and 500 microns and said outer trenches'"'"' second width is between about 50 and 500 microns.

31. The process described in claim 28 wherein the first dielectric layer is selected from the group consisting of silicon oxide, phosphosilicate glass, silicon nitride, polymers, and plastics.

32. The process described in claim 28 wherein the layer of material suitable for use as a sensor and as a resistive heater is selected from the group consisting of monocrystalline silicon germanium and gallium arsenide, metals, and ceramics.

33. The process described in claim 28 wherein said second depth of the two top preliminary trenches is between about 30 and 100 microns and their third width is between about 20 and 100 microns and each top preliminary trench is about 100 microns from an inner trench

34. The process described in claim 28 wherein said fourth width of the chamber trench is between about 100 and 10,000 microns and said increased fourth depth of the top preliminary trenches is between about 60 and 600 microns.

35. The process described in claim 28 wherein the second dielectric layer is formed to a thickness between about 0.1 and 0.5 microns.

36. The process described in claim 28 wherein said under-trench has a width between about 200 and 12,000 microns and a depth between about 50 and 500 microns.

37. The process described in claim 28 wherein said sheet of dielectric material is glass and bonding of the sheet to the wafer is achieved by means of anodic bonding

38. The process described in claim 28 wherein said sheet of dielectric material is selected from the group consisting of rigid plastics, fused quartz, silicon, elastomers, and ceramics, and bonding is by means of glue or epoxy.

16. A structure for thermal cycling, comprising:

a frame made of a thermally conductive material and having an open interior area,

suspended within said open area, a plurality of chambers in the form hollow bodies each being connected to the frame through a single thermally conductive beam;

within each chamber a heat transfer area, having an interior surface, through which all heat that flows in and out of the chamber passes;

within each chamber, a heating element symmetrically disposed inside the transfer area;

for each chamber, a first fluid bearing channel that enters the chamber through a first orifice located within the transfer area;

for each chamber, a second fluid bearing channel that enters the chamber through a second orifice located within the transfer area;

within each chamber, a baffle that is parallel to the surface of the transfer area and that is orthogonally connected to the transfer area by a sheet of material that comes between the first and second orifices;

within each chamber, at least one temperature sensor per heating element, said sensor being located close to said heating element; and

means for sending fluid into, and removing fluid from, each chamber through said channels and orifices whereby fluid flow into and out of- each chamber is individually controllable.

28. A process for manufacturing a thermal cycler, comprising the sequential steps of:

providing a silicon wafer having upper and lower surfaces,

in said upper surface, etching two inner and two outer trenches to a first depth, said inner tenches having a first width and said outer trenches having second width;

forming a first dielectric layer on said upper surface, including said trenches;

depositing a layer of material suitable for use as a sensor and as a resistive heater,

patterning and etching the material layer to form temperature sensors and heater elements;

in said upper surface etching, to a second depth, two top preliminary trenches having a third width, each being located between an inner trench and an outer trench;

patterning and etching said upper surface whereby a chamber trench, having a fourth width and located between said inner trenches, is formed to a third depth and the top preliminary trenches have their depth increased to a fourth depth;

forming a second dielectric layer on said upper surface, including all trenches;

patterning and etching the lower surface of the wafer to form an under-trench that is wide enough to slightly overlap the top preliminary trenches, to a depth such that the top preliminary trenches extend through said lower surface and, within the chamber trench, the wafer has a thickness that is between about 30 and 100 microns,

providing a sheet of dielectric material and micro-machining said sheet to form holes in selected locations; and

bonding the sheet to the wafer thereby forming a hermetically sealed chamber that is thermally isolated from the wafer.