Side Illumination Inverted Microscope Geometry for Flat Sample Imaging
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Solution Overview
Problem
Selective plane illumination microscopy (SPIM) faces challenges with instrument design and sample geometry due to the perpendicular arrangement of objective lenses, requiring specific sample preparation, large sample containers, and limited field of view for flat samples, as well as lack of isolation and increased reagents needed, especially when dealing with hazardous samples.
Innovation Solution
A device comprising a side illumination unit, a two-window sample chamber with magnetic attachment, and refractive index matching, allowing for distortion-free imaging of flat samples and enabling high-throughput 3D imaging without the need for dipping into a sample container, using a microscope with a side illumination unit that generates a light sheet and a two-window sample chamber with optically transparent windows perpendicular to each other, and refractive index matching to minimize optical aberrations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If two objective lenses are arranged perpendicular to each other for SPIM, then axial sectioning capability is achieved, but instrument design complexity and sample preparation requirements increase
Solution Approach 1:
The patent inverts the conventional SPIM geometry by placing the detection objective on the bottom (through the transparent substrate) and the illumination objective on top, rather than the typical side-illumination configuration. This inversion allows the use of standard flat samples on transparent substrates while maintaining axial sectioning capability, thereby reducing instrument design complexity and sample preparation requirements
2Ease of operation
If a large sample container is used to accommodate both lenses at 45° angle, then both lenses can be positioned, but immersion volume increases causing sample disturbance and increased reagent usage
Solution Approach 1:
The patent segments the illumination and detection paths into separate vertical positions rather than requiring a single large container. The illumination objective is positioned above the sample while the detection objective is positioned below, allowing each lens to have its own optimized optical path without requiring a large shared immersion volume, thereby reducing reagent usage and sample disturbance
3Ease of operation
If observation plane is at an angle with respect to sample container, then lenses can be positioned, but field of view for flat samples is limited
Solution Approach 1:
By inverting the detection geometry to observe from the bottom through the transparent substrate, the patent aligns the observation plane parallel to the sample surface rather than at an angle. This allows the full field of view of the detector to be utilized for flat samples such as monolayers, thereby maximizing the observable area
4Ease of manufacture
If conventional sample mounts are used with SPIM, then sample preparation is simplified, but specific sample preparation requirements of SPIM are violated
Solution Approach 1:
The patent makes the system universal by accepting conventional sample mounts with transparent substrates (coverslips, glass slides, well plates) that are already in widespread use. The inverted bottom-observation geometry allows these standard mounts to be used without modification while maintaining SPIM imaging quality, thereby achieving both ease of sample preparation and measurement precision
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 efficient imaging of thick and flat samples with high numerical aperture lenses, maximizing field of view and allowing for high-throughput 3D imaging of multiple samples while maintaining sample isolation and reducing reagent usage, even with hazardous samples.
Implementation Method 1
a side illumination unit, which includes components to generate a light sheet illuminating the sample
Implementation Method 2
The refractive index matching comprises raising a sample in the two window sample chamber using an optically transparent material with a refractive index identical to the sample immersion fluid
Implementation Method 3
The two window sample chamber further comprises a magnetic attachment to ensure easy to handle, stable, and reproducible mounting of a sample
Data Source
AI summary
A sample imaging device includes a side illumination unit, a two window sample chamber, and refractive index matching. An optically transparent sample holder is in the sample well as is sample immersion fluid. The refractive index matching includes matching of the refractive index of material of a sample to be imaged.