Detection and digital quantitation of multiple targets

A method comprising:

• (a) generating a plurality of droplets comprising at least 100,000 droplets,
• wherein the plurality of droplets comprises:
• (i) a set of target nucleic acid molecules;
• and
• (ii) a set of processing materials,
• wherein a droplet of the plurality of droplets comprises two or fewer target nucleic acid molecules of the set of target nucleic acid molecules;
• (b) providing the plurality of droplets in a well plate;
• (c) in the plurality of droplets, reacting the set of processing materials with the plurality of target nucleic acid molecules;
• (d) using a detection subsystem to scan the plurality of droplets across a set of channels to detect signals from at least a subset of the plurality of droplets, wherein each channel of the set of channels comprises a different filter configuration, wherein the set of channels comprises at least six channels, and wherein the number of target nucleic acid molecules of the set of target nucleic acid molecules is greater than the number of channels of the set of channels;
• and
• (e) using the signals detected in (d) to identify at least a subset of the plurality of target nucleic acid molecules.

A method comprising:

• (a) generating a plurality of droplets comprising at least 100,000 droplets, wherein the plurality of droplets comprises:
• (i) a set of target nucleic acid molecules from a sample;
• and
• (ii) a set of processing materials, wherein the set of processing materials comprises a first probe comprising a fluorescein amidite (FAM) fluorophore, a second probe comprising an Alexa Fluor™ fluorophore, and a third probe comprising a Cyanine 5™ (Cy5™) fluorophore,
• wherein a droplet of the plurality of droplets comprises two or fewer target nucleic acid molecules of the set of target nucleic acid molecules;
• (b) providing the plurality of droplets in a well plate;
• (c) in the plurality of droplets, reacting the set of processing materials with the set of target nucleic acid molecules;
• (d) using a detection subsystem to scan the plurality of droplets across a set of channels to detect signals from at least a subset of the plurality of droplets,
• wherein each channel of the set of channels comprises a different filter configuration,
• wherein the set of channels comprises at least four channels, and
• wherein the number of target nucleic acid molecules of the set of target nucleic acid molecules is greater than the number of channels of the set of channels;
• and
• (e) using the signals detected in (d) to identify at least a subset of the set of target nucleic acid molecules.