Mesoscale polynucleotide amplification device and method

A device for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction, the device comprising:

• a solid substrate microfabricated to define:
• a sample inlet port;
• and
• a mesoscale flow system comprising:
• a sample flow channel extending from said inlet port;
• and
• a polynucleotide polymerization reaction chamber, in fluid communication with said flow channel, containing reagents for the polymerization reaction;
• and
• means for thermally regulating the contents of said chamber whereby the temperature is controlled to amplify said preselected polynucleotide;
• and
• wherein the sample is a cell sample, the device further comprising:
• cell lysing means, in fluid communication with said reaction chamber in said mesoscale flow system, for lysing a cell sample;
• and
• means for inducing flow of said sample to said cell lysing means then to said reaction chamber.

A device for amplifying a preselected polynucleotide in a sample by conducting a polymerase chain reaction (PCR), the device comprising:

• a solid substrate microfabricated to define:
• a sample inlet port;
• and
• a mesoscale flow system comprising:
• a sample flow channel extending from said inlet port;
• and
• a PCR chamber, in fluid communication with said flow channel, for receiving said preselected polynucleotide and PCR reagents;
• an appliance for use in combination with said substrate, said appliance comprising:
• a nesting site for holding said substrate, which comprises fluid input means interfitting with an inlet port on said substrate;
• electrical contacts fabricated in said substrate, said nesting site further including an electrical connection for interfitting with said electrical contact in said substrate;
• means for thermally cycling the contents of said chamber whereby, in each cycle, temperature is controlled to separate double stranded polynucleotide, and to synthesize polynucleotide, thereby to amplify said preselected polynucleotide;
• and
• said flow system further comprising a detection chamber in fluid communication with said PCR chamber.

A method for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction, the method comprising:

• (i) providing a device comprising:
• a solid substrate microfabricated to define:
• a sample inlet port;
• and
• a mesoscale flow system comprising:
• a sample flow channel having a cross-section and extending from said inlet port;
• and
• a polynucleotide polymerization reaction chamber having a cross-section and being in fluid communication with said flow channel, said flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is between about 0.1 to 500 .mu.m said reaction chamber cross-section and said flow channel cross-section being dissimilar;
• means for thermally regulating the contents of said chamber at a temperature controlled to amplify said preselected polynucleotide;
• and
• means for detecting said amplified polynucleotide, which means is effective to detect resistance to flow of liquid in said flow channel caused by polynucleotide aggregation;
• (ii) delivering, through said inlet port and said mesoscale flow system to said reaction chamber, said sample polynucleotide and reagents required for the polymerization reaction;
• (iii) thermally controlling the contents of said chamber to polymerize said polynucleotide;
• and
• (iv) detecting the occurrence of resistance to flow in said polynucleotide flow channel.

A method for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction, the method comprising:

• (i) providing a device comprising:
• a solid substrate microfabricated to define:
• a sample inlet port;
• and
• a mesoscale flow system comprising:
• a sample flow channel extending from said inlet port;
• and
• a polynucleotide polymerization reaction chamber in fluid communication with said flow channel;
• and
• means for thermally regulating the contents of said chamber at a temperature controlled to amplify said preselected polynucleotide;
• (ii) delivering, through said inlet port and said mesoscale flow system to said reaction chamber, said sample polynucleotide and reagents required for the polymerization reaction;
• (iii) thermally controlling the contents of said chamber to polymerize said polynucleotide;
• wherein the sample is a cell sample, and the device further comprises:
• cell lysing means, in fluid communication with said reaction chamber, in said mesoscale flow system, for lysing a cell sample prior to delivery of the sample to the reaction chamber;
• and
• means for inducing flow of said sample through said cell lysing means to said reaction chamber;
• and
• wherein step (ii) includes the step of delivering said sample to said lysis means then to said reaction chamber.

A device for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction, said device comprising at least

• one silicon substrate and including:
• a sample inlet port;
• and
• a mesoscale flow system comprising:
• a sample flow channel having a cross-section and extending from said inlet port;
• and
• a polynucleotide polymerization reaction chamber having a cross-section and in fluid communication with said flow channel, said flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is between about 0.1 to 500 .mu.m said reaction chamber cross-section and said flow channel cross-section being dissimilar;
• means for thermally regulating the contents of said chamber whereby the temperature is controlled to amplify said preselected polynucleotide;
• and
• means for detecting said amplified polynucleotide.