Real-time detection of influenza virus

• US 9,885,715 B2
• Filed: 01/14/2014
• Issued: 02/06/2018
• Est. Priority Date: 05/10/2006
• Status: Active Grant

• Current Assignee: Labrador Diagnostics LLC (FIG LLC (d/b/a Fortress Investment Group LLC))
• Original Assignee: Theranos IP Company, LLC
• Inventors: Holmes, Elizabeth A., Gibbons, Ian
• Primary Examiner(s): Blumel, Benjamin P
• Application Number:
  US14/155,150
• Publication Number:
  US 20140193806A1
• Time in Patent Office: 1,484 Days
• Field of Search: None
• CPC Class Codes: B01L 2200/10 Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
• CPC Class Codes: B01L 2200/142 Preventing evaporation
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/044 pierceable, e.g. films, membranes
• CPC Class Codes: B01L 2300/0672 Integrated piercing tool
• CPC Class Codes: B01L 2300/0874 Three dimensional network
• CPC Class Codes: B01L 2400/0481 squeezing of channels or chambers
• CPC Class Codes: B01L 2400/049 vacuum
• CPC Class Codes: B01L 2400/0683 mechanically breaking a wall or membrane within a channel or chamber
• CPC Class Codes: B01L 3/5027 by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: B01L 3/502738 characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: C12Q 1/701 Specific hybridization probes
• CPC Class Codes: G01N 2333/11 Orthomyxoviridae, e.g. influenza virus
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/56983 Viruses

Abstract

The present invention provides system and methods for detecting an analyte indicative of an influenza viral infection in a sample of bodily fluid. The present invention also provides for systems and method for detection a plurality of analytes, at least two of which are indicative of an influenza viral infection in a sample of bodily fluid.

Claims

1. A method of performing a trend analysis on the concentration of an analyte comprising two influenza proteins on the same influenza viral particle, said analyte concentration being indicative of an influenza viral infection in samples obtained from a subject, comprising:

a) providing a fluidic device comprising at least one sample collection unit and an immunoassay assembly containing immunoassay reagents comprising a first antibody that binds specifically to a hemagglutinin, and a second antibody that binds specifically to a neuraminidase;

b) actuating said fluidic device and directing said immunoassay reagents within said fluidic device;

c) allowing a sample of bodily fluid obtained from said subject to react with said immunoassay reagents contained within said assay immunoassay assembly to yield a detectable signal indicative of the simultaneous presence of a hemagglutinin and a neuraminidase on said same influenza viral particle in said sample;

d) detecting said detectable signal generated from the simultaneous presence of a hemagglutinin and of a neuraminidase on the same influenza viral particle, said analyte collected in said sample of bodily fluid comprising said influenza viral particle, and determining the concentration of said analyte in said sample from said detectable signal; and

e) repeating steps a) through d) for one or more further bodily fluid samples obtained from said subject over a period of time to detect concentrations of said analyte over said period of time, thereby performing said trend analysis.

2. The method of claim 1, wherein the hemagglutinin is selected from the group consisting of H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, and H16, and the neuraminidase is selected from the group consisting of N1, N2, N3, N4, and N5.

3. The method of claim 1, wherein the hemagglutinin is H5 and the neuraminidase is N1, or wherein the hemagglutinin is H1 and the neuraminidase is N1.

4. The method of claim 1, wherein detecting said detectable signal is indicative of an influenza viral infection selected from an influenza type A viral infection, an influenza type B viral infection, and an influenza type C viral infection.

5. The method of claim 1, wherein said immunoassay assembly further comprises a plurality of channels in fluid communication with said sample collection unit or with said immunoassay assembly.

6. The method of claim 1, wherein said one or more further bodily fluid samples are obtained at a frequency selected from once a week and twice a week.

7. A method of performing a trend analysis on the concentration of an analyte comprising an influenza viral particle in each of a plurality of bodily fluid samples obtained from a subject suspected of containing influenza viral particles, wherein each of said influenza viral particles comprises a first antigen and a second antigen present on the same influenza viral particle, said method comprising:

a) providing an immunoassay assembly, wherein said assembly comprises a first antibody that binds specifically to said first antigen present on influenza viral particles, wherein the first antibody is bound to a surface of the assembly;

b) reacting a bodily fluid sample obtained from said subject in the immunoassay assembly in the presence of immunoassay reagents comprising a second antibody that binds specifically to said second antigen present on influenza viral particles which is distinct from the first antigen present on influenza viral particles, wherein the second antibody is labeled with a detectable label, wherein either the first antigen comprises a hemagglutinin and the second antigen comprises a neuraminidase, or the first antigen comprises a neuraminidase and the second antigen comprises a hemagglutinin, and wherein said reaction yields an immune complex comprising the first antibody bound to the first antigen, and the second antibody bound to the second antigen, wherein both antibodies are bound to antigens on the same influenza viral particle, said analyte comprising said influenza viral particle;

c) washing the assembly to remove excess immunoassay reagents;

d) detecting a signal generated by the detectable label from the immune complex, thereby indicating the simultaneous presence of the first antigen and the second antigen on the same influenza viral particle; and

e) repeating steps a) through d) for one or more further bodily fluid samples obtained from the subject over a period of time to detect concentrations of said analyte over said period of time, thereby performing said trend analysis on the concentration of an analyte indicative of an influenza viral infection in a subject.

8. The method of claim 7, wherein the hemagglutinin is selected from the group consisting of H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, and H16, and the neuraminidase is selected from the group consisting of N1, N2, N3, N4, and N5.

9. The method of claim 7, wherein the hemagglutinin is H5 and the neuraminidase is N1, or wherein the hemagglutinin is H1 and the neuraminidase is N1.

10. The method of claim 7, wherein detecting said detectable signal is indicative of an influenza viral infection selected from an influenza type A viral infection, an influenza type B viral infection, and an influenza type C viral infection.

11. The method of claim 7, wherein said immunoassay assembly further comprises a plurality of channels in fluid communication with said sample collection unit or with said immunoassay assembly.

12. The method of claim 7, wherein said one or more further bodily fluid samples are obtained at a frequency selected from once a week and twice a week.

13. A method of detecting a trend in the concentration of influenza viral particles in a plurality of bodily fluid samples obtained from a subject, comprising:

a) allowing a bodily fluid sample suspected of containing an influenza viral particle to react with a first immunoassay reagent and a second immunoassay reagent, both of which immunoassay reagents are contained within an immunoassay assembly of a cartridge, wherein said first immunoassay reagent binds to a hemagglutinin molecule to form a first immune complex on said influenza viral particle, and wherein said second immunoassay reagent binds to a neuraminidase molecule to form a second immune complex on the same influenza viral particle, said immune complexes yielding one or more detectable signals that indicate the simultaneous presence of hemagglutinin and neuraminidase on said same influenza viral particle wherein either the first or second immunoassay reagent is immobilized on a solid support;

b) detecting said one or more detectable signals generated from said immune complexes, wherein said simultaneous presence of hemagglutinin and neuraminidase is indicative of the presence of said influenza viral particle simultaneously having hemagglutinin and neuraminidase, and is indicative of concentrations of influenza viral particles in the sample; and

c) repeating steps a) and b) for one or more further bodily fluid samples obtained from the subject over a period of time to detect concentrations of said influenza viral particles over said period of time, thereby performing said trend analysis.

14. The method of claim 13, wherein the volume of each bodily fluid sample is less than about 500 microliters.

15. The method of claim 13, wherein detection of said hemagglutinin and neuraminidase is indicative of an influenza viral infection selected from an influenza type A viral infection, an influenza type B viral infection, and an influenza type C viral infection.

16. The method of claim 13, wherein said hemagglutinin is selected from the group consisting of H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, and H16, and said neuraminidase is selected from the group consisting of N1, N2, N3, N4, and N5.

17. The method of claim 13, wherein said hemagglutinin is H5 and said neuraminidase is N1, or wherein said hemagglutinin is H1 and said neuraminidase is N1.

18. The method of claim 13, wherein said cartridge comprises at least one sample collection unit and an immunoassay assembly containing immunoassay reagents.

19. The method of claim 13 further comprising actuating said cartridge and directing said immunoassay reagents within said cartridge.

20. The method of claim 13, wherein said sample of bodily fluid suspected of containing said influenza viral particle reacts with said immunoassay reagents contained within said immunoassay assembly based on a protocol transmitted from an external device.

21. The method of claim 20, wherein the protocol is wirelessly transmitted from said external device.

22. The method of claim 13, further comprising the step of transmitting said detectable signal to an external device.

23. The method of claim 22, wherein the external device is a server.

24. The method of claim 13, wherein the cartridge comprises a plurality of channels providing fluid communication between said sample collection unit and said immunoassay assembly.

25. The method of claim 13, wherein said cartridge comprises an identifier, a sample collection unit, and an immunoassay assembly comprising said immunoassay reagents;

an identifier detector reads said identifier and communicates to a communication assembly that transmits information of said identifier to an external device;

a protocol associated with said identifier, said protocol being transmitted from said external device in response to the transmission of said information of said identifier to said external device; and

wherein said sample of bodily fluid collected in said sample collection unit reacts with said immunoassay reagents based on said protocol to yield said detectable signal indicative of the presence of said viral particle.

26. The method of claim 13, wherein said one or more further bodily fluid samples are obtained at a frequency selected from once a week and twice a week.