MICROFLUIDICS BASED ASSAY DEVICE

• US 20140017709A1
• Filed: 02/07/2012
• Published: 01/16/2014
• Est. Priority Date: 02/07/2011
• Status: Active Grant

• Current Assignee: Lumiradx UK Ltd
• Original Assignee: Multi-Sense Technologies Limited (LumiraDx Ltd. (Cayman Islands))
• Inventors: Lowe, Phillip, Keatch, Steven Alexander, Mcguigan, Brian
• Granted Patent: US 9,341,620 B2
• Time in Patent Office: Days
• US Class Current: 435/7.92
• CPC Class Codes: B01L 2200/04 Exchange or ejection of cartridges, containers or reservoirs
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/161 Control and use of surface tension forces, e.g. hydrophobic, hydrophilic
• CPC Class Codes: B01L 2400/043 magnetic forces
• CPC Class Codes: B01L 2400/0688 surface tension valves, capillary stop, capillary break
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: C12Q 1/001 Enzyme electrodes
• CPC Class Codes: C12Q 1/006 for glucose
• CPC Class Codes: C12Y 101/03004 Glucose oxidase (1.1.3.4)
• CPC Class Codes: G01N 2333/904 acting on CHOH groups as donors, e.g. glucose oxidase, lactate dehydrogenase (1.1)
• CPC Class Codes: G01N 27/3277 being a redox reaction, e.g. detection by cyclic voltammetry voltammetry per se G01N27/42, G01N27/48
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/5438 Electrodes

Abstract

A subtractive corrective assay device and methodology, whereby ail required binding and label detection reagents are initially located within the detection zone. Application of a magnetic field is used to selectively remove bound label from the detection zone by means of paramagnetic particles. The relationship between measured label concentration before and after the application of a magnetic field within the detection zone is used to accurately measure analyte concentration within the sample.

Claims

1. A microfluidic assay cartridge for use in a subtraction assay for detecting an analyte in a sample of fluid, the cartridge comprising:

a sample port for introducing said fluid sample into the cartridge,

a substrate comprising one or more microfluidic channels disposed therein and comprising a binding agent disposed within said channel(s) for binding any of said analyte within the sample and a label for use in detecting an amount of the analyte present in the sample; and

a detection area within or wholly comprising an area where the analyte and the label binding occurs, said detection area from which bound analyte is removed allowing determination of said analyte concentration indirectly by measurement of any remaining unbound label.

2. The microfluidic assay cartridge according to claim 1, wherein the binding agent and the label are deposited within the detection area prior to a sample being introduced into the cartridge.

3. The microfluidic assay cartridge according to claim 2, wherein one or more additional reagents are present or deposited upstream or within the detection area.

4. The microfluidic assay cartridge according to claim 1, wherein the cartridge is adapted for carrying out more than one assay.

5. The microfluidic assay cartridge according to claim 4, wherein the cartridge is for carrying out the same assay measurement multiple times on a single sample or for carrying out different assay measurements on a single sample.

6. The microfluidic assay cartridge according to claim 1, wherein the binding agent is attached or otherwise bound to a particle, such as a paramagnetic particle.

7. The microfluidic assay cartridge according to claim 1, wherein the label is glucose oxidase.

8. The microfluidic assay cartridge according to claim 1, wherein the cartridge further comprises:

a working electrode and

a counter/reference electrode or a counter electrode and a reference electrode, wherein the working electrode is present in the detection area of the cartridge.

9. The microfluidic assay cartridge according to claim 8, wherein a separate counter/reference or the counter electrode and the reference electrode are provided for each corresponding working electrode, or a single counter/reference electrode or a single counter electrode and a single reference electrode is provided to work in conjunction with a plurality of working electrodes.

10. The microfluidic assay cartridge according to claim 3, wherein said one or more additional reagents are glucose, ferrocyanide, and peroxidase.

11. A method of conducting an assay on a sample, the method comprising the steps of:

introducing a sample into a microfluidic cartridge according to claim 1 such that an analyte present in the sample is capable of being bound by the analyte binding agent present in the cartridge;

detecting a level of a label that is complexed or uncomplexed to the analyte present in the detection area in order to obtain a first reference/control value;

removing the analyte/label complex from the detection area; and

detecting the uncomplexed label or an unreacted label that remains in the detection area after removal of said analyte/label complex.

12. The method according to claim 11, wherein introduction of the sample to the cartridge causes at least a portion of the binding agent and the label to be brought into solution or suspension within the detection area.

13. The method according to claim 12, wherein the detection is carried out electrochemically.

14. The method according to claim 13, wherein an oxidising potential or a reduction potential is used to measure a level of the label.

15. The method according to claim 14,

wherein the oxidising potential or the reduction potential is applied to reduce any background effects, such as to eliminate or minimise effect of electrochemical interferents which may be present in the sample,

wherein the effect of the electrochemical interferents may include generating a background signal or minimising a current generated by an enzyme during the binding incubation phase.

16. The method according to claim 13, wherein glucose oxidase is employed as the label.

17. The method according to claim 16, further comprising the step of:

using a peroxidise, such as HRP, and a mediator, such as ferrocyanide or ABTS, which is able to be converted between an oxidised form and a reduced form and which can be electrochemically detected.

18. The method according to claim 11, wherein the binding agent is coupled or otherwise bound to a particle which is capable of being complexed with the analyte present in the sample and which can be removed, when complexed or uncomplexed with the analyte, from the detection area, by suitable means.

19. The method according to claim 18, wherein the particle is a paramagnetic particle and the suitable means is a magnet or an electromagnet.

20. The method according to claim 11, wherein the sample is a sample of whole blood.

21. The method according to claim 11, wherein detection is carried out directly on the sample, without any separation, washing step, or dilution of the sample by a further fluid being carried out.

22. The method according to claim 11, wherein a decrease in a signal before and after removal of the bound analyte/label from the detection area is detected, and wherein the signal is proportional to the amount of the analyte present in the sample.

23. An assay system for conducting an assay on a fluid sample, the assay system comprising:

a) a microfluidic cartridge according to claim 1, wherein the cartridge includes a plurality of paramagnetic particles; and

b) a reader device, wherein the reader device includes;

i) a receiving port for introducing the cartridge into the reader;

ii) a magnet or magnetic force generating means, capable of applying a magnetic force to the cartridge and to remove the paramagnetic particles from the detection area within the cartridge; and

iii) a detection means for detecting a label present within a detection area of the cartridge, both before removal and following removal of an analyte/binding agent complex from the detection area of the cartridge.

24. The assay system according to claim 23, wherein a decrease from a first signal detected following detection of a reconstituted label in the detection area to a second signal detected following removal of an analyte/binding agent complex from the detection area, is detected by the reader and a level of an analyte present in a sample is calculated by the reader.

25. The assay system according to claim 24, wherein the first signal and the second signal is detected electrochemically.

26. The assay system according to claim 23, wherein the reader further comprises means, such as a display or a printing device, for identifying the level of an analyte present in the sample to a user of the system, or a signalling means to transmit the level of the analyte to a remote location or a remote practitioner.