Viability Cell Counting via Differential Light Absorption
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Solution Overview
Problem
Current methods for distinguishing between live and dead cells in biological tissues or fluids are unreliable and difficult to automate, as they often require additional steps or suffer from miscounting due to the lack of contrast between dead cells, especially when they are clumped or irregularly shaped.
Innovation Solution
Using a vital stain and capturing two images at different wavelength bands, one showing minimal absorption and the other showing higher absorption, allows for accurate counting of both live and dead cells by exploiting the differential absorption of light by the stain, enabling distinction between live and dead cells through contrasting cell outlines.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If vital stains are used to eliminate dead cells from total cell count, then dead cells can be distinguished from live cells, but cell outlines become indiscernible and multiple cells can be miscounted
Solution Approach 1:
The patent transitions from single-wavelength imaging to multi-wavelength imaging, adding a spectral dimension to the observation. By capturing images at multiple wavelengths and processing them to create composite images with enhanced contrast, the system resolves the contradiction between stain absorption and outline visibility.
Solution Approach 2:
The patent uses color enhancement and pseudo-color mapping in the composite image generation process. Different wavelength bands are assigned to different color channels, and the combination creates color contrasts that make both stained cells and cell outlines simultaneously visible and distinguishable.
2Productivity
If automated instruments are used for cell counting, then productivity increases, but they cannot readily incorporate contrast agent treatment steps
Solution Approach 1:
The patent extracts the contrast enhancement function from physical chemical treatment steps and implements it through optical processing and image computation. This allows automated instruments to achieve contrast enhancement without adding complex chemical treatment modules.
Solution Approach 2:
The patent replaces mechanical/chemical contrast enhancement mechanisms with optical and computational methods. Instead of using physical contrast agents that require additional handling steps, the system uses multi-wavelength optical imaging and digital image processing to achieve the same effect.
3Area of stationary object
If a large field of view is used in cell counters, then more cells can be observed, but resolution decreases making it difficult to differentiate live from dead cells
Solution Approach 1:
The patent adds spectral dimension through multi-wavelength imaging to compensate for the loss of spatial resolution. By using wavelength as an additional discrimination parameter, the system can differentiate cell types even when spatial details are less distinct in the larger field of view.
Solution Approach 2:
The patent uses color coding in composite images to enhance cell differentiation. Different wavelength responses are mapped to different colors, allowing cells to be distinguished by their color characteristics rather than relying solely on spatial resolution and grayscale intensity variations.
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 method provides accurate and reliable cell counts by differentiating between live and dead cells, even when they are clumped or irregularly shaped, improving the precision of cell counting in biological samples.
Implementation Method 1
treating the population with a vital stain, illuminating the population or an aliquot of the population, and taking as few as two images of light from the illumination that penetrates the population. The images are of light from different wavelength bands that are absorbed by the vital stain to different degrees.
Data Source
AI summary
Cell counts that distinguish between live and dead cells while providing an accurate count of the total of live and dead cells are obtained by the use of a vital stain in conjunction with illumination of the cell population and the taking of light images at different wavelengths, one which is not absorbed by the stain and one that is absorbed by the stain. Masking and inaccuracies in the counting of dead cells is thereby avoided.