Detection of secreted aspartyl proteases from Candida species

• US 20070134743A1
• Filed: 12/14/2005
• Published: 06/14/2007
• Est. Priority Date: 12/14/2005
• Status: Active Grant

• Current Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Original Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Inventors: Phillips, Erica, McGrath, Kevin, DiGiammarino, Enrico
• Granted Patent: US 7,745,158 B2
• Time in Patent Office: Days
• US Class Current: 435/7.230
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387
• CPC Class Codes: G01N 33/56961 Plant cells or fungi

Abstract

Methods and devices for the detection of proteins secreted by the hyphal growth form of Candida species are disclosed. The disclosed devices may constitute a method for the diagnosis of acute or chronic infections, including candidiasis, caused by microorganisms of the species Candida, such as C. albicans, for example. The devices of the present invention incorporate antibodies specific to secreted aspartyl protease proteins whose expression is upregulated upon the conversion of the Candida species from the commensal to the pathogenic form. The antibodies may be used in assays to allow the diagnosis of candidal infections and disease conditions. Either monoclonal antibodies or polyclonal antibodies may be used, and in the case of the monoclonals, the specific epitopes of the SAP protein may be detected as well as the SAP protein itself.

Claims

1. A diagnostic test kit for detecting a secreted aspartyl protease protein within a test sample, the diagnostic test kit comprising:

an assay device comprising a fluidic medium, the fluidic medium defining a detection zone within which is immobilized a receptive material, wherein the detection zone is capable of generating a detection signal that corresponds to the presence or absence of a secreted aspartyl protease protein; and

a detection probe conjugated with a binding member;

wherein the receptive material, the binding member, or both contain a first antibody that is capable of specifically binding to the secreted aspartyl protease protein.

2. The diagnostic test kit of claim 1, wherein the receptive material contains the first antibody.

3. The diagnostic test kit of claim 2, wherein the binding member contains a second antibody that specifically binds to the secreted aspartyl protease protein.

4. The diagnostic test kit of claim 3, wherein the first antibody is a monoclonal antibody that specifically binds to a first epitope of the secreted aspartyl protease protein and the second antibody is a monoclonal antibody that specifically binds to a second epitope of the secreted aspartyl protease protein.

5. The diagnostic test kit of claim 1, wherein the binding member contains the first antibody.

6. The diagnostic test kit of claim 1, wherein the first antibody is a monoclonal antibody.

7. The diagnostic test kit of claim 1, wherein the first antibody is a polyclonal antibody.

8. The diagnostic test kit of claim 1, wherein the fluidic medium is a porous membrane.

9. The diagnostic test kit of claim 1, wherein the first antibody has been developed against a recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

10. The diagnostic test kit of claim 9, wherein the first antibody has been developed by a hybridoma cell that has been generated through fusion of a human tumor cell with a cell from an animal immunized with the recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

11. The diagnostic test kit of claim 1, wherein the diagnostic test kit is capable of detecting infection by Candida albicans.

12. A diagnostic test kit for detecting a secreted aspartyl protease protein within a test sample, the diagnostic test kit comprising:

an assay device comprising a porous membrane, the porous membrane defining a detection zone within which is immobilized a first antibody, wherein the detection zone is capable of generating a detection signal that corresponds to the presence or absence of the secreted aspartyl protease protein; and

a detection probe conjugated with a second antibody;

wherein the first antibody and the second antibody are capable of specifically binding to the secreted aspartyl protease protein.

13. The diagnostic test kit of claim 12, wherein the first antibody is a monoclonal antibody that specifically binds to a first epitope of the secreted aspartyl protease protein and the second antibody is a monoclonal antibody that specifically binds to a second epitope of the secreted aspartyl protease protein.

14. The diagnostic test kit of claim 12, wherein the first antibody, the second antibody, or both are monoclonal antibodies.

15. The diagnostic test kit of claim 12, wherein the first antibody, the second antibody, or both are polyclonal antibodies.

16. The diagnostic test kit of claim 12, wherein the first antibody, the second antibody, or both are developed against a recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

17. The diagnostic test kit of claim 16, wherein the first antibody, the second antibody, or both are developed by a hybridoma cell that has been generated through fusion of a human tumor cell with a cell from an animal immunized with the recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

18. A method for detecting the presence of a secreted aspartyl protease protein within a test sample, said method comprising:

i) providing an assay device that comprises a fluidic medium, the fluidic medium defining a detection zone within which is immobilized a receptive material, the detection zone being in fluid communication with detection probes conjugated with a binding member, wherein the receptive material, the binding member, or both contain a first antibody that is capable of specifically binding to the secreted aspartyl protease protein;

ii) contacting the assay device with the test sample; and

iii) generating a detection signal within the detection zone that corresponds to the presence or absence of the secreted aspartyl protease protein.

19. The method of claim 18, wherein the receptive material contains the first antibody.

20. The method of claim 19, wherein the binding member contains a second antibody that specifically binds to the secreted aspartyl protease protein.

21. The method of claim 18, wherein the binding member contains the first antibody.

22. The method of claim 18, wherein the first antibody is a monoclonal antibody.

23. The method of claim 18, wherein the first antibody is a polyclonal antibody.

24. The method of claim 18, wherein the first antibody is developed against a recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

25. The method of claim 24, wherein the first antibody is formed by a hybridoma cell that is generated through fusion of a human tumor cell with a cell from an animal immunized with the recombinant secreted aspartyl protease protein or an immunogenic fragment thereof.

26. The method of claim 18, further comprising measuring the intensity of the detection signal, wherein the amount of the secreted aspartyl protease in the test sample is proportional to the intensity of the detection signal.

27. The method of claim 18, further comprising obtaining the test sample from a host, wherein the detection signal indicates the presence of a pathogen in the host, wherein the pathogen expresses secreted aspartyl protease proteins.

28. The method of claim 27, wherein the pathogen is Candida albicans.

29. The method of claim 18, wherein the test sample is obtained from vaginal fluid.

30. The method of claim 18, wherein the test sample is obtained form saliva.

31. The method of claim 18, wherein the test sample is obtained from blood.