High-affinity
Provided are high affinity Mycobacterium tuberculosis capsule-specific antibodies and fragments thereof, as well as methods of use and devices employing such antibodies and/or fragments.
1. An assay device for selectively detecting Mycobacterium tuberculosis (Mtb) bacteria or Mtb arabinomannan (AM) in a biological sample, wherein the assay device is a lateral flow assay device or an Enzyme Linked Immunosorbent Assay (ELISA), the assay device comprising:
(a) a first portion comprising a first plurality of anti-Mtb AM antibodies, or Mtb AM-binding fragments thereof, wherein each anti-Mtb AM antibody, or Mtb AM-binding fragment thereof, comprises a heavy chain variable region and a light chain variable region, wherein each of the heavy chain and the light chain variable regions comprises a CDR1, CDR2, and CDR3; and wherein:
(i) the sequence of CDR1 of the heavy chain comprises SEQ ID NO: 1; the sequence of CDR2 of the heavy chain comprises SEQ ID NO:2; the sequence of CDR3 of the heavy chain comprises SEQ ID NO:3; the sequence of CDR1 of the light chain comprises SEQ ID NO:4; the sequence of CDR2 of the light chain comprises SEQ ID NO:5; and the sequence of CDR3 of the light chain comprises SEQ ID NO:6; or
(ii) the sequence of CDR1 of the heavy chain comprises SEQ ID NO: 1; the sequence of CDR2 of the heavy chain comprises SEQ ID NO:2; the sequence of CDR3 of the heavy chain comprises SEQ ID NO:45; the sequence of CDR1 of the light chain comprises SEQ ID NO:4; the sequence of CDR2 of the light chain comprises SEQ ID NO:5; and the sequence of CDR3 of the light chain comprises SEQ ID NO:6,
wherein the anti-Mtb antibodies, or Mtb AM-binding fragments thereof, are each attached to their own reporting entity; and
(b) a second portion comprising a second plurality of anti-Mtb AM antibodies, or Mtb AM-binding fragments thereof.
2. The assay device of claim 1 , wherein:
(a) the heavy chain variable region comprises SEQ ID NO:13 and the light chain variable region comprises SEQ ID NO:15;
(b) the heavy chain variable region comprises SEQ ID NO:14 and the light chain variable region comprises SEQ ID NO:15;
(c) the heavy chain variable region comprises SEQ ID NO:19 and the light chain variable region comprises SEQ ID NO:15; or
(d) the heavy chain variable region comprises SEQ ID NO:20 and the light chain variable region comprises SEQ ID NO:15.
3. The assay device of claim 1 , wherein the assay device is a lateral flow assay device.
4. The assay device of claim 3 , wherein the reporting entity comprises a gold nanoparticle or an enzyme.
5. The assay device of claim 4 , wherein the enzyme is horseradish peroxidase (HRP) or alkaline phosphatase (AP).
6. The assay device of claim 3 , further comprising a fluid sample pad prior in sequential order to the first and second portions.
7. The assay device of claim 3 , further comprising a control portion subsequent in sequential order to the first and second portions.
8. The assay device of claim 7 , further comprising a fluid-absorbent wicking pad subsequent in sequential order to the first and second portions.
9. The assay device of claim 7 , wherein the control portion comprises a third plurality of antibodies, immobilized on a solid support, and which third plurality of antibodies are capable of binding the first plurality of anti-Mtb AM antibodies, or Mtb AM-binding fragments thereof.
10. The assay device of claim 1 , wherein the second plurality of anti-Mtb AM antibodies, or Mtb AM-binding fragments thereof, is affixed to a solid support.
11. The assay device of claim 10 , wherein the solid support comprises nitrocellulose.
12. A method of detecting Mtb bacteria or Mtb AM in a biological sample comprising:
(a) contacting the assay device of claim 10 the sample; and
(b) detecting binding of Mtb bacteria or Mtb AM to the first and the second plurality of anti-Mtb AM antibodies.
13. The method of claim 12 , wherein the sample is urine or blood.
14. The method of claim 12 , wherein:
(a) the heavy chain variable region comprises SEQ ID NO:13 and the light chain variable region comprises SEQ ID NO:15;
(b) the heavy chain variable region comprises SEQ ID NO:14 and the light chain variable region comprises SEQ ID NO:15;
(c) the heavy chain variable region comprises SEQ ID NO:19 and the light chain variable region comprises SEQ ID NO:15; or
(d) the heavy chain variable region comprises SEQ ID NO:20 and the light chain variable region comprises SEQ ID NO:15.
15. The method of claim 12 , wherein the assay device is a lateral flow assay device.
16. The method of claim 15 , the assay device further comprises a control portion subsequent in sequential order to the first and second portions.
17. The method of claim 16 , wherein the control portion comprises a third plurality of antibodies, immobilized on a solid support, and which third plurality of antibodies are capable of binding the first plurality of anti-Mtb AM antibodies, or Mtb AM-binding fragments.
18. The method of claim 12 , wherein the reporting entity comprises a gold nanoparticle or an enzyme.
19. The method of claim 18 , wherein the enzyme is HRP or AP.