AGENTS AND METHODS FOR SPECTROMETRIC ANALYSIS
Disclosed herein are agents, methods, and kits for determining the presence or concentration of a target, or multiple targets, in a sample, in a uniplexed or multiplexed fashion. In general, the methods enable the analysis of small molecules produced or consumed in liquid-phase that may be analyzed using gas or vapor phase detection methods.
1 . A method of determining the presence of a target in a test sample: comprising:
(a) providing a test sample and a target present in the test sample;
(b) providing a capture agent capable of selectively binding to the target, wherein the capture agent is adhered to a solid support;
(c) providing a binder capable of selectively binding to the target, wherein the binder is coupled to a catalyst;
(d) contacting the test sample with the capture agent and the binder in an assay solution, wherein a captured target complex containing the capture agent, the target, and the binder is formed when the capture agent and the binder combine with the target present in the test sample;
(e) washing the solid support to separate captured target complex from non-complexed assay components;
(f) combining a substrate reactive with the catalyst to the separated captured target complex in a solution, wherein the catalyst converts the substrate to a catalysis product; and
(g) performing analysis of the solution of step (f) for an analyte selected from the substrate or the catalysis product; wherein the analyte is selected from 3,3′,5,5′-tetramethylbenzidine (TMB), hydrogen peroxide, nicotinamide, 8-hydroxyquinoline, pyridoxal, and pyridoxamine.
2 . The method of claim 1 , further comprising a step of ionizing components of the solution of the step (f) before the step (g).
3 . The method of claim 1 , wherein the substrate is 3,3′,5,5′-tetramethylbenzidine (TMB) and the catalyst is peroxidase.
4 . The method of claim 1 , wherein the substrate is 3-cyanopyridine and the catalyst is nitrile hydratase.
5 . The method of claim 1 , wherein the substrate is 8-hydroxyquinoline glucuronide and the catalyst is glucuronidase.
6 . The method of claim 1 , wherein the substrate is 8-hydroxyquinoline glucopyranoside and the catalyst is glucosidase.
7 . The method of claim 1 , wherein the substrate is 8-hydroxyquinoline β-D-galactopyranoside and the catalyst is galactosidase.
8 . The method of claim 1 , wherein the substrate is glucose and the catalyst is glucose oxidase.
9 . The method of claim 1 , wherein the substrate is hydrogen peroxide and the catalyst is catalase.
10 . The method of claim 1 , wherein the substrate is orthonitrophenylgalactoside and the catalyst is galactosidase.
11 . The method of claim 1 , wherein the substrate is orthonitrophenylglucopyranoside and the catalyst is glucosidase.
12 . The method of claim 1 , wherein the substrate is paranitrophenol phosphate and the catalyst is phosphatase.
13 . The method of claim 1 , wherein the substrate is phenylphosphate and the catalyst is phosphatase.
14 . The method of claim 1 , wherein the substrate is pyridoxal phosphate and the catalyst is phosphatase.
15 . The method of claim 1 , wherein the substrate is pyridoxamine phosphate and the catalyst is phosphatase.
16 . The method of claim 1 , wherein the capture agent is selected from an antibody, an aptamer, an affibody, and a ligand.
17 . The method of claim 1 , wherein the binder is selected from an antibody, an aptamer, an affibody, and a ligand.
18 . The method of claim 1 , wherein the solid support is selected from superparamagnetic particles, membranes, and polymer substrates.
19 . The method of claim 1 , wherein the solid support is selected from synthetic or modified naturally occurring polymers, comprising nitrocellulose, cellulose acetate, poly(vinyl chloride), dextran, polyacrylate, polyethylene, polyethersulfone, polypropylene, poly(4-methylbutene), polystyrene, polymethacrylate, poly(ethylene terephthalate), nylon, or poly(vinyl butyrate).
20 . The method of claim 1 , wherein the solid support comprises a plate, a well, a strip, a rod, or a particle.
21 . The method of claim 1 , wherein the solid support is a membrane and step (e) results from lateral flow of the non-complexed assay components across the membrane.
22 . The method of claim 1 , wherein the step (g) includes determining absence or presence of the analyte in the test sample.
23 . The method of claim 22 , wherein the absence or presence of the analyte in the test sample is correlated with the absence or presence of the target in the test sample.
24 . The method of claim 1 , wherein step (g) further includes quantifying an amount of the analyte in the test sample.
25 . The method of claim 24 , wherein the amount of the analyte detected in the test sample is correlated with quantity of the target in the test sample.
26 . The method of claim 1 , wherein the target is selected from prokaryotic cells, eukaryotic cells, bacteria, viruses, proteins, polypeptides, toxins, liposomes, particles, ligands, amino acids, nucleic acids, hormones, pharmaceuticals, toxic industrial chemicals, toxic industrial materials, or combinations thereof.
27 . The method of claim 2 , wherein the analyte is ionized using chemical, electrical, or photo ionization and step (g) is performed using a method selected from ion mobility spectrometry (IMS), ion mobility trap spectrometry (ITMS), mass spectrometry (MS), high-field asymmetric waveform ion mobility spectrometry (FAIMS), differential mobility spectrometry (DMS), and gas chromatography (GC).