Device and method for performing digital PCR

Patent metadata

Field	Value
Publication number	US-11266990-B2
Application number	US-201816059576-A
Country	US
Kind code	B2
Filing date	Aug 9, 2018
Priority date	Nov 29, 2013
Publication date	Mar 8, 2022
Grant date	Mar 8, 2022

Abstract

A micro-fluidic device 100 for performing digital PCR is presented. The device comprises: a semiconductor substrate; a first micro-fluidic channel 104, comprising an inlet 102 and an outlet 103, embedded in the semiconductor substrate; a heating element 101 thermally coupled to the first micro-fluidic channel 104; a droplet generator 107 connected to the inlet 102 of the first micro-fluidic channel 104 for generating droplets and pumping generated droplets at a flow rate into the first micro-fluidic channel 104; characterized in that: the heating element 101 is a single heating element connected to a temperature control unit 111 configured to cycle the temperature of the complete first micro-fluidic channel 104 through at least two temperature values; and wherein the flow rate of the droplet generator 107 is adaptable. Further, a method to perform digital PCR is presented using the micro-fluidic device 100.

First claim

The invention claimed is:

1. A method for performing continuous PCR on a fluid sample using a micro-fluidic device, the fluid sample comprising an analyte, the method comprising:
continuously providing the fluid sample in the micro-fluidic device, the micro-fluidic device comprising
a semiconductor substrate;
a first micro-fluidic channel, comprising an inlet and an outlet,
embedded in the semiconductor substrate;
a single heating element configured for cycling a temperature of the first micro-fluidic channel through at least two temperature values, the heating element thermally coupled to the first micro-fluidic channel;
a droplet generator connected to the inlet of the first micro-fluidic channel and configured for generating droplets of the fluid sample and pumping the generated droplets into the first micro-fluidic channel, the droplet generator is reconfigurable for changing a number of copies of the analyte in the droplets;
a detector located at the outlet of the first micro-fluidic channel and configured for detecting droplets containing PCR products following PCR in the first micro-fluidic channel, the PCR products comprising the copies of the analyte; and
a computing unit connected to the detector and to the droplet generator and configured for continuously monitoring and determining a percentage of droplets containing PCR products based on input data from the detector and for reconfiguring the droplet generator to change the number of the copies of the analyte in the droplets to a pre-determined level based on the percentage of droplets;

continuously generating droplets of the fluid sample and pumping the droplets into the first micro-fluidic channel using the droplet generator;
continuously cycling the temperature of the first micro-fluidic channel through the at least two temperature values using the heating element while continuously pumping the droplets into the first micro-fluidic channel;
continuously detecting droplets containing the PCR products using the detector; and
continuously monitoring and determining the number of droplets containing the copies of the analyte and reconfiguring the droplet generator to change the number of copies of the analyte in the droplets using the computing unit to continuously adjust a carrier flow rate and an analyte flow rate and optionally a diluent flow rate of the droplet generator, depending on the percentage of droplets containing PCR products as determined by the computing unit.

2. The method according to claim 1 , wherein said generating droplets and pumping the droplets is stopped when the first micro-fluidic channel is completely filled with the droplets.

3. The method according to claim 1 , wherein the droplets are pumped at a flow rate adapted to a duration of the temperature cycling of the first micro-fluidic channel.

4. The method according to claim 1 , further comprising: pre-heating the generated droplets before pumping the droplets into the first micro-fluidic channel.

5. The method according to claim 1 , wherein the micro-fluidic device further comprises a heating element present at the inlet of the first micro-fluidic channel and a second micro-fluidic channel, the second micro-fluidic channel connected on one side to an outlet of the droplet generator and on the other side to the heating element present at the inlet of the first micro-fluidic channel, wherein the heating element located at the inlet of the first micro-fluidic channel pre-heats the droplets prior to being pumped into the first micro-fluidic channel.

6. The method according to claim 1 , wherein the micro-fluidic device further comprises at least one through-substrate trench at least partially surrounding the first micro-fluidic channel.

Assignees

• Imec Vzw

Inventors

• Fiorini Paolo
• Stakenborg Tim
• Colle Frederik

Classifications

• B01L2300/0867

  Multiple inlets and one sample wells, e.g. mixing, dilution · CPC title

• B01L2300/0816

  Cards, e.g. flat sample carriers usually with flow in two horizontal directions · CPC title

• B01L2200/0673

  Handling of plugs of fluid surrounded by immiscible fluid · CPC title

• B01L7/52

  with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples · CPC title

• B01L3/502784 Primary

  specially adapted for droplet or plug flow, e.g. digital microfluidics · CPC title