Analysis devices, kits, and related methods for digital quantification of nucleic acids and other analytes
Provided are devices and methods for effecting processing of samples, including essentially isothermal amplification of nucleic acids, at multiple reaction locations in a single device. In some embodiments, the disclosed devices and methods provide for effecting parallel sample processing in several hundred locations on a single device.
1. A method of amplifying a nucleic acid molecule, comprising:
(a) introducing a sample comprising at least one nucleic acid molecule within a plurality of first areas in a first substrate and a plurality of first areas in a second substrate, wherein said first and second substrates are configured in a first position with the plurality of first areas in said first substrate overlapping with the plurality of first areas in said second substrate to form a continuous fluidic path within said first and second substrates;
(b) contacting the sample disposed in the plurality of first areas in the first substrate with at least one component of an amplification reagent disposed in a plurality of second areas in the second substrate,
the contacting being effected by relative motion between said first substrate and said second substrate to a second position, wherein said plurality of first areas in said first substrate are isolated from the plurality of first areas in said second substrate, thereby placing at least some of the first areas in said first substrate into direct fluid communication with at least some of the plurality of second areas so as to form a plurality of closed reaction chambers; and
(c) exposing the plurality of closed reaction chambers having the at least one nucleic acid molecule to conditions effective for amplification of the at least one nucleic acid molecule.
2. The method of claim 1 , wherein the amplification is essentially isothermal.
3. The method of claim 1 , wherein at least two of the plurality of first areas differ from one another in volume, wherein at least two of the plurality of second areas differ from one another in volume, wherein at least one first area differs in volume from at least one second area, or any combination thereof.
4. The method of claim 1 , wherein the relative motion comprises rotation, linear translation, or both.
5. The method of claim 1 , wherein at least 10 first areas are placed into direct fluid communication with at least 10 second areas essentially simultaneously.
6. The method of claim 1 , wherein at least one of the first and second areas has a volume of from 0.1 nL to about 1000 nL.
7. The method of claim 1 , further comprising introducing the at least one component of the amplification reagent to the plurality of second areas.
8. The method claim 1 , wherein introducing the sample to the plurality of first areas comprises exerting the sample through a conduit in fluid communication with the plurality of first areas.
9. The method of claim 8 , wherein the conduit is formed in said second substrate.
10. The method of claim 7 , wherein introducing the at least one component of the amplification reagent to the plurality of second areas comprises exerting the amplification reagent through a conduit in fluid communication with the plurality of second areas.
11. The method of claim 10 , wherein the conduit is formed in said first substrate.
12. The method of claim 1 , further comprising distributing the sample between the plurality of first areas and a first control area, wherein said sample at the first control area remains uncontacted with the at least one component of the amplification reagent during exposure to said conditions effective for amplification.
13. A method of effecting amplification of at least one nucleic acid target molecule, comprising:
(a) introducing a sample material comprising a nucleic acid target within a plurality of first areas in a first substrate and a plurality of first areas in a second substrate, wherein said first and second substrates are configured in a first position with the plurality of first areas in said first substrate overlapping with the plurality of first areas in said second substrate to form a continuous fluidic path within said first and second substrates;
(b) contacting
(1) the sample material disposed in the plurality of first areas in the first substrate, at least one of the first areas in the first substrate containing one molecule of the nucleic acid target, with
(2) a reactant material disposed in a plurality of second areas in the second substrate, the contacting being effected by relative motion between said first substrate and said second substrate to a second position, wherein said plurality of first areas in said first substrate are isolated from the plurality of first areas in said second substrate, resulting in pairwise placement of at least some of the first areas and at least some of the second areas into direct fluid communication with one another so as to form a plurality of closed reaction chambers,
the contacting effecting amplification of at least one nucleic acid target molecule.
14. The method of claim 13 , wherein the sample material comprises a reagent.
15. The method of claim 13 , wherein the reactant material comprises an amplification reagent, and wherein the method further comprises exposing the at least one of the first areas containing one molecule of the nucleic acid target to conditions effective for amplification of the one nucleic acid target so as to give rise to an amplification product.