Treponema Pallidum Triplet Antigen

Patent metadata

Field	Value
Publication number	US-2016115206-A1
Application number	US-201514980200-A
Country	US
Kind code	A1
Filing date	Dec 28, 2015
Priority date	Jan 13, 2011
Publication date	Apr 28, 2016
Grant date	—

Abstract

A Treponema pallidum triplet antigen construct is disclosed which includes three Treponema pallidum antigens (TP15, TP17, and TP47), as well as a ten amino acid leader sequence (tag 261) and human copper zinc superoxide dismutase (hSOD). This construct is optimized for in vitro diagnosis of syphilis infection. Plasmids containing DNA encoding the triplet antigen, host cells, production methods, detection methods, and kits are also disclosed.

First claim

1 . A recombinant plasmid encoding a Treponema pallidum triplet antigen, wherein the plasmid comprises nucleic acid encoding an amino acid sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, and SEQ ID NO:12.

2 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:7.

3 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:8.

4 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:9.

5 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:10.

6 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:11.

7 . The recombinant plasmid of claim 1 wherein the amino acid sequence is SEQ ID NO:12.

8 . The recombinant plasmid of claim 1 , wherein the nucleic acid has a nucleotide sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6.

9 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:1.

10 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:2.

11 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:3.

12 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:4.

13 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:5.

14 . The recombinant plasmid of claim 1 wherein the nucleic acid has a nucleotide sequence as shown in SEQ ID NO:6.

15 . A host cell comprising the recombinant plasmid of claim 1 .

16 . The host cell of claim 15 wherein the host cell is Escherichia coli.

17 . A method of producing a Treponema pallidum triplet antigen which comprises culturing the host cell of claim 15 under conditions suitable for expression of the Treponema pallidum triplet antigen, and recovering the Treponema pallidum triplet antigen from the cell culture.

18 . A recombinant plasmid encoding a Treponema pallidum triplet antigen, the recombinant plasmid selected from the group consisting of:
the plasmid designated p261nS-TP17-15-47 and deposited with the American Type Culture Collection as ATCC Accession No. PTA-11590; the plasmid designated p261nS-TP47-17-15 and deposited with the American Type Culture Collection as ATCC Accession No. PTA-11589; the plasmid designated p261nS-TP17-47-15; the plasmid designated p261nS-TP47-15-17; the plasmid designated p261nS-TP15-17-47; and the plasmid designated p261nS-TP15-47-17.

19 . The recombinant plasmid of claim 18 designated p261nS-TP17-15-47 and deposited with the American Type Culture Collection as ATCC Accession No. PTA-11590.

20 . The recombinant plasmid of claim 18 designated p261nS-TP47-17-15 and deposited with the American Type Culture Collection as ATCC Accession No. PTA-11589.

21 . The recombinant plasmid of claim 18 designated p261nS-TP17-47-15.

22 . The recombinant plasmid of claim 18 designated p261nS-TP47-15-17.

23 . The recombinant plasmid of claim 18 designated p261nS-TP15-17-47.

24 . The recombinant plasmid of claim 18 designated p261nS-TP15-47-17.

25 . A host cell comprising the recombinant plasmid of claim 18 .

26 . The host cell of claim 25 wherein the host cell is Escherichia coli.

27 . A method of producing a Treponema pallidum triplet antigen which comprising culturing the host cell of claim 25 under conditions suitable for expression of the Treponema pallidum triplet antigen, and recovering the Treponema pallidum triplet antigen from the cell culture.

28 . A Treponema pallidum triplet antigen having an amino acid sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, and SEQ ID NO:12.

29 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:7.

30 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:8.

31 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:9.

32 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:10.

33 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:11.

34 . The Treponema pallidum triplet antigen of claim 28 having an amino acid sequence as shown in SEQ ID NO:12.

35 . The Treponema pallidum triplet antigen of claim 28 labeled with a detectable marker.

36 . The Treponema pallidum triplet antigen of claim 35 wherein the detectable marker is horseradish peroxidase.

37 . A method of detecting the presence of Treponema pallidum antibodies in a sample, the method comprising:
contacting the sample with the Treponema pallidum triplet antigen of claim 28 , wherein Treponema pallidum antibodies present in the sample bind to the Treponema pallidum triplet antigen forming an antibody/antigen complex; and detecting the antibody/antigen complex, thereby detecting the presence of Treponema pallidum antibodies.

38 . The method of claim 37 wherein the Treponema pallidum triplet antigen is immobilized on a solid support.

39 . The method of claim 37 wherein detecting the antibody/antigen complex comprises:
contacting the antibody/antigen complex with a second Treponema pallidum triplet antigen of claim 28 , the second Treponema pallidum triplet antigen labeled with a detectable marker, wherein the second Treponema pallidum triplet antigen binds to the antibody present in the antibody/antigen complex forming an antigen/antibody/labeled antigen complex; and
detecting the antigen/antibody/labeled antigen complex, thereby detecting the presence of Treponema pallidum antibodies.

40 . The method of claim 37 wherein detecting the antibody/antigen complex comprises:
contacting the antibody/antigen complex with labeled anti-human antibody, wherein the labeled anti-human antibody binds to the antibody present in the antibody/antigen complex forming an antigen/antibody/labeled anti-antibody complex; and
detecting the antigen/antibody/labeled anti-antibody complex, thereby detecting the presence of Treponema pallidum antibodies.

41 . A component of a kit for detection of Treponema pallidum antibodies, the component comprising the Treponema pallidum triplet antigen of claim 28 .

42 . A kit for detection of Treponema pallidum antibodies, the kit comprising:
the Treponema pallidum triplet antigen of claim 28 ; and a second Treponema pallidum triplet antigen of claim 28 , the second Treponema pallidum triplet antigen labeled with a detectable marker.

43 . The kit of claim 42 further comprising positive or negative controls.

44 . The kit of claim 42

Assignees

• Ortho Clinical Diagnostics Inc

Inventors

• Zheng Jian
• Yura Renee
• Yang Jianping

Classifications

• C07K2319/00

  Fusion polypeptide · CPC title

• G01N2800/26

  Infectious diseases, e.g. generalised sepsis · CPC title

• G01N33/571

  for venereal disease, e.g. syphilis, gonorrhoea {(herpes G01N33/56994; chlamydia G01N33/56927)} · CPC title

• C07K14/20 Primary

  from Spirochaetales (O), e.g. Treponema, Leptospira · CPC title

• G01N33/56911

  Bacteria · CPC title