Nucleic acid amplification apparatus

• US 5,725,831 A
• Filed: 03/24/1995
• Issued: 03/10/1998
• Est. Priority Date: 03/14/1994
• Status: Expired due to Fees

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Becton, Dickinson & Co
• Inventors: Bourdelle, Peter A., Reichler, Allen S., Cracauer, Raymond F.
• Primary Examiner(s): Pyon, Harold Y.
• Application Number:
  US08/409,805
• Time in Patent Office: 1,082 Days
• Field of Search: 422/50, 422/58, 422/61, 422/68.1, 422/73, 422/56, 422/82.09, 422/102, 422/104, 436/63, 436/94, 436/86, 436/180, 436/501, 436/508, 436/807, 435/6, 435/91, 435/172.3, 435/301
• US Class Current: 422/417
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/0825 Test strips
• CPC Class Codes: B01L 2300/087 Multiple sequential chambers
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 3/502 with fluid transport, e.g. in multi-compartment structures centrifugal-type cuvettes G01N21/07; analysis by separation into components G01N30/00; automatic analysers G01N35/00
• 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 3/50273 characterised by the means or forces applied to move the fluids micropumps F04B19/006, of the membrane type F04B43/043
• 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
• CPC Class Codes: C12Q 1/6848 characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

An apparatus for containing a liquid biological sample and for performing a biological process thereon comprises a sample area for receiving the sample, at least one reaction area in fluid communication with the sample area, a pneumatic area in pneumatic communication with the reaction area and the sample area, and a pneumatic port in the pneumatic area for connection of the apparatus to a pneumatic aspiration/dispensing pipette. The pneumatic aspiration/dispensing pipette provides for controlled flow of the liquid biological sample between the sample area and the reaction area. To reduce evaporative loss of the sample from the apparatus, a sample tower is provided in fluid communication with the sample area. A similar tower may be provided at the pneumatic port to reduce evaporative loss through the pneumatic area. A restricted orifice between the sample tower and the sample area holds the liquid sample in place during removal of the sample from the apparatus, counteracting the tendency of the liquid bolus to flow back toward the reaction area due to capillary forces. Also disclosed is a novel sealing arrangement which provides an effective pneumatic seal between the pneumatic port and a pneumatic aspiration/dispensing pipette without requiring the apparatus to include a separate "O" ring or other type of discrete sealing device.

Claims

1. An apparatus for containing a liquid biological sample and for performing a biological process thereon, comprising:

a sample area for receiving a liquid biological sample, said sample area having an orifice for admitting said sample into said sample area;

at least one reaction area in fluid communication with said sample area;

a pneumatic area in pneumatic communication with said reaction area and said sample area;

a pneumatic port in said pneumatic area for connection of said apparatus to a pneumatic aspiration/dispensing means, said aspiration/dispensing means causing the flow of said liquid biological sample between said sample area and said reaction area; and

a sample tower located above said sample area and in fluid communication with said sample area through said orifice for reducing evaporative loss of said liquid biological sample from said sample area and said reaction area, said sample tower having a sample port at an upper end thereof for admitting said liquid biological sample into said sample area through said orifice, said sample tower having said orifice at a lower end thereof;

wherein said orifice is of smaller size than the adjacent portion of said sample area to provide a capillary lock for inhibiting capillary flow of said liquid biological sample from said sample area to said reaction area;

wherein said pneumatic area includes a pneumatic tower for reducing evaporative loss of said liquid biological sample through said pneumatic area, said pneumatic tower having said pneumatic port at an upper end thereof, and wherein said pneumatic port is encircled by a pair of rigid, concentric sealing rings adapted to make sealing contact with a resilient portion of said pneumatic aspiration/dispensing means.

2. The apparatus of claim 1, wherein said sample tower has interior walls tapering inwardly in the direction from said sample port to said orifice.

3. The apparatus of claim 1, wherein said sample tower, said sample port and said orifice are each circular in cross-section.

4. The apparatus of claim 1, wherein said sample area and said reaction area are provided in the form of a continuous channel through which said liquid biological sample flows in the form of a liquid bolus.

5. The apparatus of claim 1, wherein said apparatus is elongate in shape with said sample area and said pneumatic area at opposite ends thereof, and said reaction area positioned therebetween.

6. The apparatus of claim 1, further comprising a microchannel positioned between said reaction area and said pneumatic area for substantially preventing liquid flow by hydrostatic force therethrough.

7. The apparatus of claim 6, wherein said apparatus is devoid of liquid flow control means between said orifice and said microchannel.

8. The apparatus of claim 6, wherein said microchannel has an enlarged opening extending partially across said microchannel.

9. The apparatus of claim 1, wherein said apparatus is formed from non-wettable material to substantially prevent liquid flow by capillary action.

10. The apparatus of claim 9, wherein said apparatus is formed from polypropylene.

11. The apparatus of claim 1, wherein at least a first dried reagent necessary for said biological process is affixed to an internal surface of said reaction area.

12. The apparatus of claim 11, wherein a second dried reagent necessary for said biological process is affixed to an internal surface of said reaction chamber at a location different from that of said first dried reagent.

13. The apparatus of claim 12, wherein said biological process has a decontamination step utilizing said first dried reagent and a nucleic acid amplification step utilizing said second dried reagent, and wherein the locations of said first and second dried reagents in said reaction chamber are such that said liquid biological sample is subjected to said decontamination step prior to said amplification step.

14. A system for performing a biological process, comprising:

an apparatus for containing a liquid biological sample, said apparatus including a sample area for receiving said liquid biological sample, at least one reaction area in fluid communication with said sample area, a pneumatic area in pneumatic communication with said reaction area and said sample area, and a pneumatic port in said pneumatic area; and

pneumatic aspiration/dispensing means adapted to be brought into contact with said pneumatic port to cause said liquid biological sample to flow between said sample area and said reaction area;

wherein said pneumatic port of said apparatus is encircled by at least one rigid sealing ring and said pneumatic aspiration/dispensing means has a resilient portion adapted to be deformed by contact with said sealing ring to create a pneumatic seal around said pneumatic port.

15. The system of claim 14, wherein said pneumatic aspiration/dispensing means comprises a rigid aspiration/dispensing pipette and said resilient portion comprises a resilient tip affixed to said pipette, said resilient tip having an opening therein communicating with the lumen of said pipette and with the pneumatic port of said apparatus.

16. The system of claim 15, wherein said resilient tip is made of silicone rubber.

17. The system of claim 15, wherein said sealing ring is one of a pair of rigid, concentric sealing rings encircling said pneumatic port and adapted to be brought into contact with said resilient tip.