H&E Stained Tissue Eosin Fluorescence Removal
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Solution Overview
Problem
Current methods for cancer diagnosis face challenges in analyzing multiple biological targets from a single sample, particularly when samples are limited, as existing techniques like H&E staining interfere with molecular analysis techniques such as IHC and FISH, and removal of H&E stains is not feasible for IF and FISH due to residual eosin fluorescence.
Innovation Solution
A method involving a borate salt-based photo-induced chemical bleaching (PICB) process that partially removes eosin from H&E stained samples, allowing for the same tissue section to be used for immunofluorescence and FISH by reducing residual eosin fluorescence through electron transfer and irradiation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If H&E staining is used for morphological assessment, then tissue morphology can be visualized, but residual eosin fluorescence interferes with subsequent immunofluorescence and FISH analysis
Solution Approach 1:
The patent applies extraction by removing the harmful eosin fluorescence component from the H&E stained tissue while preserving the beneficial hematoxylin staining and tissue morphology. This is achieved through selective photobleaching techniques that target and remove only the eosin fluorescence, allowing the same tissue section to be used for both morphological assessment and molecular analysis without interference from residual staining.
Solution Approach 2:
The patent utilizes parameter changes by altering the optical properties of the tissue section through controlled illumination and photobleaching processes. By changing the light exposure parameters and using specific wavelengths, the eosin fluorescence is selectively removed while maintaining the structural integrity and morphological features of the tissue, enabling multi-purpose analysis from a single section.
2Measurement precision
If multiple molecular targets are analyzed using IHC or IF, then molecular characterization is improved, but the number of detectable targets is limited by the fluorescence-based detection system
Solution Approach 1:
The patent applies segmentation by dividing the detection process into distinct temporal and spatial phases. Multiple molecular targets are detected in sequential steps using the same tissue section, with each target analyzed at different time points. This segmentation allows unlimited targets to be detected without the constraints of simultaneous fluorescence detection, as each target can be probed independently in sequence.
Solution Approach 2:
The patent uses preliminary action by performing H&E staining and morphological assessment first, then systematically removing the eosin fluorescence before proceeding to molecular analysis. This preliminary preparation creates an optimal state for subsequent multi-target molecular detection, ensuring that the tissue is ready for sensitive fluorescence-based assays without interference from residual H&E stains.
3Measurement precision
If additional biological samples are used for molecular analysis, then complete characterization of disease is achieved, but sample availability is reduced and relative characteristics cannot be determined
Solution Approach 1:
The patent applies universality by making a single tissue section serve multiple functions: it provides morphological assessment through H&E staining, then after eosin removal, enables molecular analysis through IHC, IF, and FISH. This multi-functional use of a single sample allows determination of relative characteristics such as presence, absence, concentration, and spatial distribution of multiple targets without requiring additional biological samples, thereby preserving sample availability for other analyses.
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 the detection of multiple targets in a single biological sample with minimal tissue disruption and interference, facilitating simultaneous morphological and molecular analysis, thereby improving diagnostic efficiency and reducing the need for additional tissue samples.
Implementation Method 1
contacting the sample with an electron transfer reagent, and irradiating the sample to remove the residual eosin fluorescence
Implementation Method 2
contacting the sample with an electron transfer reagent
Data Source
AI summary
Methods comprising the use probing multiple targets in a H&E stained biological sample are provided. The methods include the steps of providing a hematoxylin and eosin stained biological sample containing multiple targets, observing the sample, removing the hematoxylin and partially removing the eosin by washing the sample, contacting the sample with a borate salt, and irradiating the sample to remove the residual eosin fluorescence. The method further includes the optionally performing the additional steps of binding at least one probe to one or more targets to the sample, observing a signal from the probe and contacting the sample with a bleaching agent. The process of binding, observing and bleaching may be iteratively repeated.