Compositions and methods for detecting mycobacterium

Patent metadata

Field	Value
Publication number	US-9835624-B2
Application number	US-201214111677-A
Country	US
Kind code	B2
Filing date	Apr 16, 2012
Priority date	May 2, 2011
Publication date	Dec 5, 2017
Grant date	Dec 5, 2017

Abstract

The present disclosure provides methods of detecting mycobacterium in an individual, generally involving detecting antibody to a mycobacterial lipid in a biological sample obtained from the individual. The present disclosure further provides compositions and kits for carrying out the methods.

First claim

What is claimed is:

1. A method of determining response in a human individual to treatment for a non-cavitary Mycobacterium tuberculosis infection, the method comprising determining the level of IgM specific for two or more different M. tuberculosis lipids in a biological sample obtained from the individual, wherein a level of M. tuberculosis lipid-specific IgM in the biological sample that is lower than a pre-treatment level in a control sample of the same type as the biological sample indicates that the individual is responding positively to the treatment.

2. The method of claim 1 , wherein the two or more M. tuberculosis lipids are selected from cardiolipin, phosphatidylinositol, phosphatidylethanolamine, phosphatidylcholine, and sphingolipid.

3. The method of claim 1 , wherein the biological sample is blood, a blood fraction, cerebrospinal fluid, pleural fluid, bronchoalveolar lavage fluid, or saliva.

4. The method of claim 1 , wherein the two or more different M. tuberculosis lipids are selected from phosphatidylinositol, phosphatidylethanolamine, phosphatidylcholine, and sphingolipid.

5. An assay device for use in detecting IgM specific for a Mycobacterium tuberculosis lipid in a liquid biological sample obtained from an individual, the device comprising a matrix defining an axial flow path, the matrix comprising:
i) a zone one comprising a sample receiving zone at an upstream end of the flow path configured to receive the fluid sample;
ii) a zone three comprising one or more test zones positioned within the flow path and downstream from zone one, each of said one or more test zones comprising a Mycobacterium tuberculosis lipid immobilized in each of said test zones, wherein each of said immobilized lipids is capable of binding a M. tuberculosis lipid-specific IgM present in said liquid sample to form an immobilized IgM-lipid complex;
iii) a zone two comprising a label zone comprising a detectably labeled antibody specific for human IgM, wherein the label zone is positioned within the flow path configured to receive the sample from zone one and is downstream of zone one and upstream of zone three, wherein the detectably labeled antibody is capable of binding an IgM present in an immobilized IgM-lipid complex to form a labeled immobilized IgM-lipid complex, and wherein the detectably labeled antibody is capable of binding to the human IgM present in the liquid sample to form a labeled antibody/human IgM complex that is configured to flow to the one or more test zones of zone three; and
iv) a zone four comprising one or more control zones positioned within the flow path and downstream from zone one, each of said one or more control zones comprising a positive control comprising immobilized human IgM in a known amount, or a negative control comprising no M. tuberculosis lipids, wherein zone four is configured to receive the sample from zone one,
wherein the M. tuberculosis lipid-specific IgM is detected when the detectably labeled antibody specific for human IgM is bound to the immobilized IgM-lipid complex.

6. A kit for determining the level of IgM specific for two or more different Mycobacterium tuberculosis lipids in a biological sample obtained from an individual, the kit comprising:
a) an anti-IgM antibody comprising a detectable label; and
b) a panel of two or more different Mycobacterium tuberculosis lipids.

7. The kit of claim 6 , further comprising purified human IgM for use in generating a standard curve.

8. The kit of claim 6 , further comprising a panel of positive control IgM antibodies specific for the lipids in the panel.

9. The kit of claim 6 , further comprising an insoluble support having the panel of lipids attached thereto.

10. The kit of claim 6 , wherein the panel of lipids comprises cardiolipin, phosphatidylinositol, phosphatidylethanolamine, phosphatidylcholine, and sphingolipid.

11. A method for detecting the presence of Mycobacterium tuberculosis in an individual, the method comprising detecting IgM specific for two or more different Mycobacterium tuberculosis lipids in a liquid biological sample obtained from the individual, wherein the presence in the biological sample of IgM specific for two or more different Mycobacterium tuberculosis lipids indicates the presence in the individual of Mycobacterium tuberculosis , and wherein the step of detecting the IgM comprises:
a) contacting two or more different, immobilized Mycobacterium tuberculosis lipids with the liquid biological sample, wherein any IgM specific for the immobilized Mycobacterium tuberculosis lipids binds the immobilized Mycobacterium tuberculosis lipids, forming an immobilized Mycobacterium tuberculosis lipid-IgM complex; and
b) contacting the immobilized Mycobacterium tuberculosis lipid-IgM complex with detectably labeled anti-IgM antibody, wherein the anti-IgM antibody forms a complex with the immobilized Mycobacterium tuberculosis lipid-IgM; and
c) detecting the anti-IgM antibody present in the anti-IgM-immobilized Mycobacterium tuberculosis lipid-IgM complex.

12. The method of claim 11 , wherein the two or more different, immobilized M. tuberculosis lipids are selected from cardiolipin, phosphatidylinositol, phosphatidylethanolamine, phosphatidylcholine, and sphingolipid.

13. The method of claim 11 , wherein the biological sample is blood, a blood fraction, cerebrospinal fluid, pleural fluid, bronchoalveolar lavage fluid, or saliva.

14. The method of claim 11 , wherein the anti-IgM antibody is coupled to a magnetic bead, an enzyme that generates a fluorescent product, an enzyme that generates a colored product, a fluorescent polypeptide, or a chromogenic polypeptide.

15. The method of claim 14 , wherein the anti-IgM antibody is coupled to a magnetic bead.

16. The method of claim 15 , wherein coupling of the anti-IgM antibody to the magnetic bead comprises binding of an anti-IgM antibody covalently linked to a biotin to the magnetic bead coated with a streptavidin to form a stable anti-IgM magnetic bead complex.

17. The method of claim 15 , wherein coupling of the anti-IgM antibody to the magnetic bead comprises binding of an anti-IgM antibody to the magnetic bead, wherein the magnetic bead is cross-linked to a protein G or a protein A.

18. The method of claim 11 , wherein said individual is co-infected with human immunodeficiency virus.

19. The method of claim 11 , wherein said individual has non-cavitary lung disease.

20. The method of claim 11 , wherein the labeled antibody is mobilizable in the presence of the liquid sample.

21. The method of claim 11 , wherein the labeled anti-IgM antibody comprises a label selected from the group consisting of:
a) a chemiluminescent agent;
b) a particulate label;
c) a colorimetric agent;
d) an energy transfer agent;
e) an enzyme;
f) a fluorescent agent; and
g) a radioisotope.

22. The method of claim 21 , wherein the particulate label is a colloidal gold.

Assignees

• Riley Lee W
• Mathies Richard A
• Goodridge Amador
• Kim Jungkyu
• Snyder Robert Eugene
• Univ California

Inventors

• Riley Lee W
• Mathies Richard A
• Goodridge Amador
• Kim Jungkyu
• Snyder Robert Eugene

Classifications

• G01N33/5695 Primary

  Mycobacteria · CPC title

• G01N33/6854

  Immunoglobulins · CPC title

• G01N2800/52

  Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis · CPC title

• G01N2469/20

  Detection of antibodies in sample from host which are directed against antigens from microorganisms · CPC title