High-throughput label-free enzymatic bioassays using DESI-MS

Patent metadata

Field	Value
Publication number	US-12130255-B2
Application number	US-202218084827-A
Country	US
Kind code	B2
Filing date	Dec 20, 2022
Priority date	May 11, 2020
Publication date	Oct 29, 2024
Grant date	Oct 29, 2024

Abstract

The invention generally relates to high-throughput label-free enzymatic bioassays using desorption electrospray ionization-mass spectrometry (DESI-MS).

First claim

What is claimed is:

1. A system for analyzing one or more enzymatic reactions, the system comprising:
a fluid handling system that includes a pinning device configured to interact with a microwell plate and prepare a plurality of discrete spots on a substrate, wherein each of the discrete spots comprises a substrate and a product of one or more enzymatic reactions, wherein both of the substrate and the product of the one or more enzymatic reactions are label-free;
an ionization source configured to direct sequentially a discharge from the ionization source onto each of the plurality of discrete spots to sequentially desorb the substrate and/or product from each of the discrete spots and sequentially generate ions of the substrate and/or product from each of the discrete spots;
a mass spectrometer positioned to receive the generated ions of the substrate and/or product from each of the discrete spots and analyze sequentially the ions of the substrate and/or product from each of the discrete spots in the mass spectrometer to thereby analyzing the one or more enzymatic reactions; and
a single control system that comprises one or more processors that control and coordinate the fluid handling system, the ionization source, and the mass spectrometer, such that the system is a fully integrated singular system and wherein one of the one or more processors is for determining whether the test compound inhibits the enzymatic reaction based on monitoring of the progress of the enzymatic reaction.

2. The system of claim 1 , wherein the ionization source is a desorption electrospray ionization probe (DESI) and the discharge is a DESI spray.

3. The system of claim 1 , wherein a subset of each of the plurality of the discrete spots further comprises a same test compound.

4. The system of claim 1 , wherein the control system further determines how completely the test compound inhibits the enzymatic reaction based on the monitoring of the progress of the enzymatic reaction.

5. The system of claim 4 , wherein the enzymatic reaction is associated with a physiological condition and determining how completely the test compound inhibits the enzymatic reaction determines whether the test compound should be considered for development into a drug to treat the condition.

6. A system for analyzing one or more enzymatic reactions, the system comprising:
a fluid handling system that includes a pinning device configured to interact with a microwell plate and prepare a plurality of discrete spots on a substrate, wherein each of the discrete spots comprises a substrate and a product of one or more enzymatic reactions, wherein both of the substrate and the product of the one or more enzymatic reactions are label-free;
an ionization source configured to direct sequentially a discharge from the ionization source onto each of the plurality of discrete spots to sequentially desorb the substrate and/or product from each of the discrete spots and sequentially generate ions of the substrate and/or product from each of the discrete spots;
a mass spectrometer positioned to receive the generated ions of the substrate and/or product from each of the discrete spots and analyze sequentially the ions of the substrate and/or product from each of the discrete spots in the mass spectrometer to thereby analyzing the one or more enzymatic reactions; and
a single control system that comprises one or more processors that control and coordinate the fluid handling system, the ionization source, and the mass spectrometer, such that the system is a fully integrated singular system wherein a subset of each of the plurality of the discrete spots further comprises an inhibitor of an enzyme of the enzymatic reaction and a test compound and wherein one of the one or more processors determines whether the test compound can counteract the inhibitor and re-start the enzymatic reaction.

7. The system of claim 6 , wherein one of the one or more processors determines how completely the test compound counteracts the inhibitor and how completely the enzymatic reaction re-started based on the monitoring of the progress of the enzymatic reaction.

8. The system of claim 7 , wherein one of the one or more processors determines how completely the test compound counteracts the inhibitor and how completely the enzymatic reaction is re-started determines whether the test compound should be considered for development into a drug that counteracts the inhibitor.

9. The system of claim 1 , wherein each of the plurality of discrete spots is from a different time point in an enzymatic reaction.

Assignees

• Purdue Research Foundation

Inventors

• Cooks Robert Graham
• Morato Nicolás M
• Holden Dylan T

Classifications

• H01J49/0409

  Sample holders or containers (containers for retaining a material to be analyzed, B01L3/50, for DNA, C12Q1/6834, for biological materials, G01N33/543) · CPC title

• G01N27/626

  using heat to ionise a gas · CPC title

• G01N27/64

  using wave or particle radiation to ionise a gas, e.g. in an ionisation chamber · CPC title

• G01N2500/02

  Screening involving studying the effect of compounds C on the interaction between interacting molecules A and B (e.g. A = enzyme and B = substrate for A, or A = receptor and B = ligand for the receptor) · CPC title

• G01N33/6851

  Methods of protein analysis involving laser desorption ionisation mass spectrometry · CPC title