Cell-free biofragment compositions and related systems, devices, and methods

Patent metadata

Field	Value
Publication number	US-11105802-B2
Application number	US-201314650851-A
Country	US
Kind code	B2
Filing date	Dec 10, 2013
Priority date	Dec 10, 2012
Publication date	Aug 31, 2021
Grant date	Aug 31, 2021

Abstract

The present disclosure relates to biofragment compositions that comprise bioparticle fragments and at least one heterologous antigen-binding molecule. In some embodiments, the biofragment is typically derived from a larger, intact bioparticle that express the at least one heterologous antigen-binding molecule at the surface, and the biofragment has increased solubility to facilitate assays for antigen detection. The disclosure also relates the related methods of using and making the biofragment compositions, as well as systems and devices implementing the biofragment compositions. In some embodiments, the related methods, systems and devices do not require additional detection reagents, such as animal derived detection antibodies.

First claim

The invention claimed is:

1. A composition for selectively binding an antigen of interest in a sample, the composition comprising a cell wall fragment immobilized to a conductive or semi-conductive electrode surface by an intervening tether construct,
wherein the cell wall fragment displays at least one heterologous antigen-binding molecule and is a fragment of a yeast cell wall, a bacterium cell wall, or a plant cell wall;
wherein the cell wall fragment is less than 200 nm at its greatest dimension; and

wherein the at least one heterologous antigen-binding molecule is an antibody, an antigen-binding antibody fragment, or a T-cell receptor (TCR) that selectively binds the antigen of interest.

2. The composition of claim 1 , wherein the yeast is from the genus Saccharomyces or Pichia.

3. The composition of claim 1 , wherein the antibody or antigen-binding antibody fragment is a single-chain antibody, a bispecific antibody, a Fab fragment, or a F(ab) 2 fragment.

4. The composition of claim 3 , wherein the single-chain antibody is a single-chain variable fragment (scFv), single-chain Fab fragment (scFab), a single variable domain on a heavy-chain antibody fragment (V H H fragment), a variable domain of new antigen receptor fragment (V NAR fragment), or single domain antibody.

5. The composition of claim 1 , wherein the conductive or semi-conductive electrode surface is substantially free of cell wall fragments not displaying the at least one heterologous antigen-binding molecule.

6. The composition of claim 1 , wherein the cell wall fragment is produced by disruption of the cell wall.

7. The composition of claim 6 , wherein the at least one heterologous antigen-binding molecule is attached to the cell wall surface prior to disruption of the surface by inducing expression and translocation of the molecule to, or assembly of the molecule on, the interior or exterior cell wall surface.

8. A method of detecting the presence of an antigen of interest in a biological sample, comprising:
1) acting a biological sample with the composition of claim 1 under conditions sufficient to permit the binding of the heterologous antigen-binding molecule with the antigen of interest to form a complex; and
2) detecting formation of the complex.

9. The method of claim 8 , further comprising contacting the complex with a detection reagent that binds to the antigen of interest.

10. The method of claim 9 , wherein the detection reagent comprises a detectably-labeled reporter reagent, and the method further comprises separating the unbound detectably labeled-reporter from the composition.

11. The method of claim 8 , further comprising contacting the immobilized antigen of interest with an electroactive molecule and measuring electron transfer resistance at the electrode surface, wherein binding of the antigen of interest to the composition is detected by a change in the electron transfer resistance as compared to the electron transfer resistance when the antigen of interest is not present.

12. The method of claim 9 , wherein the biological sample is selected from the group consisting of blood, urine, sputum, mucus, saliva, cerebral spinal fluid, tissues, stool, nutrient sources, or processed derivatives thereof.

13. The method of claim 11 , wherein the electroactive molecule is a redox probe.

14. The method of claim 13 , wherein the redox probe is [Fe(CN) 6 ] 3−/4− .

15. An antigen detection system, comprising:
the composition of claim 1 ,
an electroactive molecule, and
a device to monitor electric current, electric potential, and/or electric impedance.

16. The system of claim 15 , wherein the electrode surface is on a screen-printed gold electrode.

17. The system of claim 15 , wherein the electroactive molecule is a redox probe.

18. The system of claim 15 , wherein the intervening tether construct is an epitope-tag binding molecule that binds to an epitope tag present in the heterologous antigen-binding molecule of the composition.

19. The system of claim 17 , wherein the redox probe is [Fe(CN) 6 ] 3−/4− .

Assignees

• Univ Queensland
• Seattle Childrens Hospital

Inventors

• Grewal Yadveer
• Cangelosi Gerard A
• Shiddiky Muhammad J A
• Trau Matt

Classifications

• G01N33/5432 Primary

  Liposomes or microcapsules · CPC title

• G01N33/5438

  Electrodes · CPC title

• G01N33/56961

  Plant cells or fungi · CPC title

• G01N33/56983

  Viruses · CPC title

• G01N33/56911

  Bacteria · CPC title