Methods and systems for detecting biological components
Methods for the detection of components from biological samples are provided. In certain aspects, the methods may be used to detect and/or quantify specific components in a biological sample, such as tumor cells (e.g., circulating tumor cells). Systems and devices for practicing the subject methods are also provided.
1. A method of synthesizing a target polynucleotide, the method comprising:
contacting on a first device a polynucleotide-containing component from a sample with lysis reagents in a lysate droplet, the lysis reagents comprising an enzyme having protease activity, wherein the lysate droplet is encapsulated in an immiscible carrier fluid;
moving the lysate droplet into a collection chamber of a second device;
incubating the lysate droplet in the collection chamber for a first duration and at a temperature sufficient to inactivate the enzyme having protease activity;
returning the lysate droplet to the first device;
adding to the lysate droplet a nucleic acid synthesis reagent to form a nucleic acid synthesis droplet in the immiscible carrier fluid; and
synthesizing the target polynucleotide within the nucleic acid synthesis droplet.
2. The method of claim 1 , wherein the first device is a microfluidic device.
3. The method of claim 1 , wherein the second device is a syringe.
4. The method of claim 1 , wherein the first duration is a period longer than necessary for lysis.
5. The method of claim 1 , wherein the step of adding to the lysate droplet a nucleic acid synthesis reagent further comprises adding a plurality of nucleic acid synthesis reagents.
6. The method of claim 1 , wherein the nucleic acid synthesis droplet has a volume of 0.001 to 1000 picoliters.
7. The method of claim 1 , wherein the nucleic acid synthesis droplet has a diameter of between 0.1 microns to 1000 microns.
8. The method of claim 1 , wherein the step of adding to the lysate droplet a nucleic acid synthesis reagent further comprises:
contacting the lysate droplet with a continuous stream of fluid comprising the nucleic acid synthesis reagent, and
forming the nucleic acid synthesis droplet from a portion of the continuous stream of fluid and the lysate droplet.
9. The method of claim 1 , further comprising detecting the target polynucleotide by determining a sequence of a nucleic acid synthesis product of the nucleic acid synthesis droplet or by forming a double-emulsion comprising the nucleic acid synthesis droplet within an outer droplet, and sorting the double-emulsion based on at least one of droplet size and fluorescence.