Holographic Microscope Cell State Analysis
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Solution Overview
Problem
Current methods for determining the undifferentiated state of pluripotent stem cells, such as iPS and ES cells, are invasive, time-consuming, and prone to human error, making them unsuitable for industrial-scale production.
Innovation Solution
A cell analysis method and system using a holographic microscope to extract cell areas, calculate background and intracellular phase values, and determine cell states based on phase differences, enabling non-invasive and automated differentiation between undifferentiated and differentiated cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Illumination intensity
If phase-contrast microscope is used for observing cells, then cell transparency is compensated, but focusing is required for each photographing which increases measurement time
Solution Approach 1:
The patent creates a holographic copy of the cell that contains both amplitude and phase information. By recording the interference pattern between object light (from the cell) and reference light, the system generates a digital hologram that can be reconstructed into focused images without mechanical focusing adjustments, eliminating the time-consuming focusing step while preserving cell visibility.
Solution Approach 2:
The patent performs focusing in advance during the hologram recording process. The holographic microscope captures the three-dimensional optical information of the cell in a single exposure, and the focused reconstructed image is obtained during data processing rather than during physical adjustment, thereby eliminating repeated focusing operations.
2Measurement precision
If undifferentiation marker staining is performed to determine cell state, then accurate determination is achieved, but cells die which prevents use in regenerative medicine
Solution Approach 1:
The patent replaces the chemical staining method with a physical optical measurement method. By measuring the phase difference of light passing through the cell using holographic microscopy, the system determines cell state (undifferentiated vs. differentiated) based on optical properties rather than chemical markers, thereby avoiding cell death while maintaining determination accuracy.
Solution Approach 2:
The patent changes the measurement parameter from chemical marker presence to optical phase difference. Undifferentiated cells and differentiated cells exhibit different phase characteristics due to differences in cell morphology, density, and internal structure. By measuring phase difference instead of chemical markers, the system achieves accurate cell state determination without invasive staining.
3Reliability
If morphological observation by phase-contrast microscope is used to determine cell state, then cell viability is maintained, but human judgment is required which introduces error and reduces efficiency
Solution Approach 1:
The patent enables the system to automatically determine cell state without human intervention. The holographic microscope captures phase information, and the built-in processing system automatically calculates phase differences and classifies cells as undifferentiated or differentiated based on predetermined thresholds, making the determination process self-service and eliminating dependency on observer skill.
Solution Approach 2:
The patent implements an automated feedback system where the measured phase difference is compared against reference values or thresholds to automatically classify cell state. This feedback mechanism enables consistent, repeatable determinations without human judgment variability, improving both efficiency and reliability for industrial-scale cell culture monitoring.
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
This approach allows for accurate and efficient determination of cell states, reducing human error and facilitating quality control and productivity in pluripotent stem cell culture.
Implementation Method 1
acquires an interference pattern (hologram) formed on the detection surface of an image sensor or the like by object light which is light from a light source reflected or transmitted on the surface of an object and reference light directly reaching from the same light source
Implementation Method 2
a phase-contrast microscope has been widely used for observing a cell
Data Source
AI summary
A cell area extraction unit (241) extracts a cell area in a phase image that is created based on a hologram obtained by in-line holographic microscope (IHM). A background value acquisition unit (242) obtains a background value from phase values at a plurality of positions outside the cell area. An intracellular phase value acquisition unit (243) averages a plurality of phase values on a sampling line set at a position close to the periphery of a cell, while avoiding a central portion in which the phase value may be lowered in the cell area, to obtain an intracellular phase value. A phase change amount calculation unit (244) obtains the difference between the intracellular phase value and the background value. A phase change amount determination unit (245) compares the value of the difference with thresholds in two levels to determine whether the cell is in an undifferentiated state or an undifferentiation deviant state. It is thereby possible to automatically make a correct determination while removing the influence of a theoretical measurement error by IHM.