Multiplexed digital assays
System, including methods and apparatus, for performing a multiplexed digital assay.
1. A method of performing a multiplexed digital assay, the method comprising:
forming partitions that collectively contain R targets;
amplifying the R targets in the partitions;
collecting data representing amplification of each of the R targets in the partitions, all of the data being collected in fewer than R optical channels; and
determining a respective level of each of the R targets from the data, wherein each level is specific for a single target of the R targets, and wherein the level determined for at least one of the R targets is based in part on a partition count for a partition population positive for two of the R targets;
wherein the step of determining is based on at least R+2 identified partition populations.
2. The method of claim 1 , wherein R is at least three.
3. The method of claim 1 , wherein the data is collected in only two optical channels.
4. The method of claim 1 , wherein each optical channel represents a different waveband of emitted light.
5. The method of claim 1 , wherein each optical channel represents a different waveband of excitation light.
6. The method of claim 1 , wherein a different plurality of the partitions contains each different pair of the R targets.
7. The method of claim 1 , wherein at least 10% of the partitions contain a pair of the R targets.
8. The method of claim 1 , further comprising a step of identifying from the data a plurality of partition populations each positive for a different combination of the R targets.
9. The method of claim 1 , wherein the step of determining is based on at least 2R identified partition populations.
10. The method of claim 8 , wherein the step of identifying includes a step of resolving each partition population that is positive for exactly two of the R targets from each partition population that is positive for only one of the R targets.
11. A method of performing a multiplexed digital assay, the method comprising:
forming partitions that collectively contain R targets;
amplifying the R targets in the partitions;
collecting data representing amplification of each of the R targets in the partitions, all of the data being collected in fewer than R optical channels;
identifying from the data a plurality of partition populations each positive for a different combination of the R targets, wherein the step of identifying includes a step of resolving each partition population that is positive for exactly two of the R targets from each partition population that is positive for only one of the R targets; and
determining a respective level of each of the R targets from the data, wherein each level is specific for a single target of the R targets, and wherein the level determined for at least one of the R targets is based in part on a partition count for a partition population positive for two of the R targets.
12. The method of claim 11 , wherein R is at least three.
13. The method of claim 11 , wherein the data is collected in only two optical channels.
14. The method of claim 11 , wherein each optical channel represents a different waveband of emitted light.
15. The method of claim 11 , wherein a different plurality of the partitions contains each different pair of the R targets.
16. The method of claim 11 , wherein at least 10% of the partitions contain a pair of the R targets.
17. The method of claim 11 , wherein the step of determining is based on at least 2R identified partition populations.
18. A method of performing a multiplexed digital assay, the method comprising:
forming partitions that collectively contain R targets;
amplifying the R targets in the partitions;
collecting data representing amplification of each of the R targets in the partitions, all of the data being collected in fewer than R optical channels, wherein each optical channel represents a different waveband of excitation light; and
determining a respective level of each of the R targets from the data, wherein each level is specific for a single target of the R targets, and wherein the level determined for at least one of the R targets is based in part on a partition count for a partition population positive for two of the R targets.
19. A method of performing a multiplexed digital assay, the method comprising:
forming partitions that collectively contain R targets;
amplifying the R targets in the partitions;
collecting data representing amplification of each of the R targets in the partitions, all of the data being collected in fewer than R optical channels;
plotting the data such that partition populations containing different combinations of the R targets form a rectangular array, wherein the step of plotting is performed with respect to a pair of orthogonal axes, and wherein each partition population that is positive for only one of the R targets is arranged along a line that is at least generally parallel to one of the axes; and
determining a respective level of each of the R targets from the data, wherein each level is specific for a single target of the R targets, and wherein the level determined for at least one of the R targets is based in part on a partition count for a partition population positive for two of the R targets.
20. The method of claim 19 , wherein R is at least three.
21. The method of claim 19 , wherein the data is collected in only two optical channels.
22. The method of claim 19 , wherein each optical channel represents a different waveband of emitted light.
23. The method of claim 19 , wherein a different plurality of the partitions contains each different pair of the R targets.
24. The method of claim 19 , wherein at least 10% of the partitions contain a pair of the R targets.
25. The method of claim 19 , further comprising a step of identifying from the data a plurality of partition populations each positive for a different combination of the R targets.
26. The method of claim 25 , wherein the step of identifying includes a step of resolving each partition population that is positive for exactly two of the R targets from each partition population that is positive for only one of the R targets.
27. The method of claim 19 , wherein the step of determining is based on at least 2R identified partition populations.
28. The method of claim 19 , wherein the array is composed of a plurality of rows and columns, with each row and each column including at least two of the partition populations.