Expanded Sample Bulk Labeling for Cellular Ultrastructure Imaging
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Solution Overview
Problem
Fluorescence microscopy is limited in revealing cellular ultrastructures due to insufficient optical resolution and the inability to label the entire sample at high density, while electron microscopy requires extensive data acquisition time for three-dimensional imaging.
Innovation Solution
A method involving physical expansion of biological samples by a factor of two or more, combined with bulk labeling using reagents such as fluorescent dyes or metallic particles, to enhance contrast and resolve fine structures without specific labeling or electron microscopy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If fluorescence microscopy is used to image proteins, then high contrast and high precision labeling are achieved, but optical resolution is insufficient to reveal fine structures and labeling density cannot be sufficiently high
Solution Approach 1:
The patent applies physical expansion to transform the sample from its native scale to an enlarged scale (e.g., 4-fold or 10-fold expansion). This dimensional transformation allows fine structures that were previously below the optical resolution limit to be magnified into the resolvable range, effectively adding a scaling dimension to overcome the resolution barrier of conventional microscopy
Solution Approach 2:
The patent changes the physical parameters of the sample by expanding its volume and altering its refractive index through hydrogel embedding. This parameter change enables bulk labeling at high density throughout the expanded sample volume, allowing uniform contrast enhancement without the limitations of conventional sparse fluorescent labeling
2Manufacturing precision
If electron microscopy is used to obtain three-dimensional images of samples, then fine structures can be resolved, but days to weeks of continuous data acquisition are required
Solution Approach 1:
The patent replaces the electron beam-based imaging mechanism with light-based fluorescence microscopy. By physically expanding the sample and applying bulk labeling, the method achieves EM-like structural resolution using optical microscopy, which operates much faster and does not require the lengthy data acquisition and processing times characteristic of electron microscopy
Solution Approach 2:
The patent transforms the sample's physical state by embedding it in expandable hydrogel and physically expanding it, which enlarges fine structures to a scale resolvable by light microscopy. This parameter change in sample dimensions allows conventional optical microscopes to achieve resolution previously only attainable through electron microscopy, dramatically improving imaging productivity
3Manufacturing precision
If the sample is physically expanded by embedding in expandable polymer, then fine structures are enlarged to resolvable scale, but the sample requires chemical fixation and hydrogel embedding
Solution Approach 1:
The expandable hydrogel polymer serves multiple functions simultaneously: it chemically fixes the sample through crosslinking, provides the expansion matrix for physical enlargement, maintains sample structure during processing, and enables bulk labeling. This multi-functionality consolidates several separate preparation steps into a single integrated system, reducing overall procedural complexity despite the added expansion capability
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 high-resolution imaging of cellular ultrastructures with improved labeling density, allowing for rapid generation of three-dimensional images without the need for electron microscopy.
Implementation Method 1
physically expanding the sample by at least a factor of two in at least one dimension by embedding the sample in an expandable polymer
Implementation Method 2
bulk labeling a plurality of amino acids of the sample with at least one reagent to introduce contrast
Data Source
Figure 1a~1c
Figure 1d~1g
Figure 1h
AI summary
Methods and systems for physical expansion and imaging of biological samples are described herein. In one aspect of the disclosure, a method for preparing a biological sample for the purpose of generating images of its ultrastructure with an imaging instrument includes a) physically expanding the sample by at least a factor of two in at least one dimension; and b) bulk labeling a plurality of components of the sample with at least one reagent to introduce contrast.