Desktop EPR Imaging for 3D Oxygen Mapping in Multi-Well Plates
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Solution Overview
Problem
Current methods for assessing viable cells, such as MTT, PicoGreen, luciferase, or clonogenic assays, are inadequate for providing spatiotemporal information, especially when cells are seeded in biomaterials or the extracellular matrix, and there is a need for three-dimensional EPR imaging of in vitro cell environmental parameters like oxygen partial pressure, pH, and viscosity using a relatively low magnetic field.
Innovation Solution
An electron paramagnetic resonance imaging (EPRI) system and method using a resistive magnet, magnetic field gradient coils, RF signal source, and computer system to generate coherent RF pulses, focus RF power, and acquire images of multiple-well cell plates, employing a spin probe to measure oxygen-dependent relaxation times and map environmental parameters.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional assay methods (MTT, PicoGreen, luciferase, clonogenic assays) are used to assess viable cells, then cell viability can be measured, but spatiotemporal information is not provided and the methods fail when cells are seeded in biomaterials or extracellular matrix
Solution Approach 1:
The patent replaces conventional mechanical/chemical assay methods with electron paramagnetic resonance (EPR) imaging technology. EPR imaging provides three-dimensional spatial information and temporal dynamics without requiring cell extraction or disruption, enabling measurement of cell viability and environmental parameters (oxygen, pH, viscosity) directly within biomaterials and extracellular matrices where cells are seeded.
Solution Approach 2:
The patent utilizes changes in EPR parameters (signal intensity, relaxation times) of spin probes in response to environmental conditions (oxygen concentration, pH, viscosity) to infer cell viability and metabolic activity. By monitoring these parameter changes over time and space, the system provides spatiotemporal information about cell behavior in three-dimensional cultures within biomaterials.
2Measurement precision
If high magnetic field strength is used for EPR imaging, then imaging quality improves, but device complexity and cost increase
Solution Approach 1:
The patent optimizes the magnetic field strength parameter for EPR imaging of spin probes, using a relatively low magnetic field strength (e.g., 0.3-1.0 Tesla) that is sufficient to achieve adequate imaging quality for detecting spin probe signals from cells in biomaterials. This reduced field strength simplifies the magnet system, reduces power consumption, and lowers overall device complexity while maintaining the ability to provide three-dimensional spatial information.
3Productivity
If fast imaging is implemented for rapid experiments on well plates, then productivity increases, but measurement precision may be compromised
Solution Approach 1:
The patent employs periodic pulsed EPR imaging sequences that rapidly acquire data from multiple wells in a cell plate over time. The pulsed nature of the imaging allows for fast repetition rates, enabling monitoring of environmental parameters (oxygen, pH, viscosity) and cell viability across multiple wells in succession, providing both high productivity and sufficient measurement precision through time-resolved data collection.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Enables fast and efficient three-dimensional imaging of cell environmental parameters like oxygen partial pressure, pH, and viscosity in a benchtop instrument, suitable for cell viability assessment and tumor progression monitoring, with rapid experiments on small objects like well plates.
Implementation Method 1
a resistive magnet driven by a power supply to generate a static magnetic field
Implementation Method 2
three magnetic field gradient coils configured to provide three magnetic fields in orthogonal alignment
Implementation Method 3
a radio frequency (RF) signal source, an RF power amplifier, and a pulse programmer configured to generate a substantially coherent polyphase sequence of RF pulses and excite a spin probe of a sample in the plate without heating the sample
Implementation Method 4
a resonator configured to focus RF power on the sample
Implementation Method 5
acquire, quantify and map pO2 data associated with a spin probe having oxygen-dependent T2* and/or spin-spin relaxation time and/or spin-lattice relaxation time
Data Source
AI summary
A small-size electron paramagnetic resonance imaging (EPRI) instrument includes an environment-controlled resonator configured to excite and detect electron spins at a resonance frequency. The instrument enables oxygen imaging of cell-seeded wells of standard 96-wells of multi-well plate manipulated by a mechanical stage. One example is configured for long multiple-well strips. In combination with an oxygen-reporting molecular, one example provides partial oxygen pressure (pO2) maps of cells in wells that can be used for cell viability measurements and analysis of drug efficacy when used with drugs and cells.