Auto-focusing Multi-well Plate Scanning via Plane Computation
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing auto-focusing methods for multiwell plates with non-planar surfaces, particularly those with U-shaped bottoms, face difficulties in focusing due to similar refraction indices of oil immersion lenses and solutions, leading to time-consuming image analysis and challenges with laser-based autofocusing methods.
Innovation Solution
An auto-focusing method and device that uses a first objective lens to identify in-focus positions in a subset of wells, computes a plane where multiple wells will be in focus with a second objective lens of equal or lower magnification, allowing for scanning without additional focusing operations, and aligns the multiwell plate using stored alignment data across scanning devices.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If laser-based autofocusing methods are used on multiwell plates with non-planar surfaces, then focusing can be achieved, but the method fails when using oil immersion lenses due to similar refraction indices canceling refraction points
Solution Approach 1:
The patent introduces an intermediary substance (water or oil with known refractive index) between the objective lens and the multiwell plate. This intermediary serves as a mediator that creates a controllable refraction interface, allowing the laser-based autofocusing method to work with oil immersion lenses by preventing the cancellation of refraction points that occurs when the lens and solution have similar refraction indices.
Solution Approach 2:
The patent changes the refractive index parameter of the medium between the lens and sample by introducing water or oil with specific refractive indices. This parameter change creates a sufficient refractive index difference to generate detectable refraction points and scanning peaks, enabling the autofocusing method to function properly with oil immersion lenses.
2Reliability
If existing autofocusing methods are used on multiwell plates with non-planar surfaces, then focusing can be achieved, but image analysis becomes time-consuming
Solution Approach 1:
The patent replaces complex image analysis methods with a simpler laser-based scanning approach. Instead of capturing and analyzing full images to determine focus, the system uses laser scanning to detect refraction points and identify scanning peaks, which provides a more direct and time-efficient method for determining focus position on non-planar surfaces.
Solution Approach 2:
The system uses the optical properties of the multiwell plate and sample itself to generate the focusing signal. The refraction of laser light at the interface between the intermediary substance and the plate creates natural scanning peaks that directly indicate focus position, eliminating the need for external image analysis processes.
3Measurement precision
If a first objective lens with high magnification is used to identify in-focus positions, then precise focusing can be achieved, but scanning speed decreases
Solution Approach 1:
The patent divides the process into two segments: first using a high magnification objective lens to identify in-focus positions with high precision, then using the determined plane information to guide scanning with appropriate speed. This segmentation allows each stage to be optimized independently for its specific function.
Solution Approach 2:
The patent performs preliminary focusing using the high magnification objective lens to establish the in-focus plane before conducting the actual scanning operation. This preliminary action ensures that subsequent scanning occurs at the correct focus position, maintaining measurement precision while allowing for optimized scanning speed.
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 auto-focusing on multiwell plates with non-planar surfaces, separating focusing from scanning processes, and allows for higher scanning and imaging speeds, even with oil immersion lenses, by determining in-focus positions and translating them across different objective lenses based on optical characteristics.
Implementation Method 1
focusing the objective using laser based autofocusing methods as described in U.S. Pat. No. 7,109,459 is difficult or impossible, as the similar refraction index of the lens and solution in the wells cancels one of the refraction points
Implementation Method 2
using a first objective lens having a first magnification to identify, in each of at least three wells of a selected subset of the plurality of wells an in-focus position of each said well with respect to the first objective lens
Data Source
AI summary
An auto-focusing method for determining an in-focus position of a plurality of wells in at least a portion of a multi-well plate, the method including using a first objective lens having a first magnification to identify, in each of at least three wells of a selected subset of the plurality of wells, an in-focus position of each well with respect to the first objective lens, on the basis of at least three the in-focus positions, computing a plane along which the at least three wells will be in focus with respect to at least one objective lens having a second magnification that is not greater than the first magnification, and using the at least one objective lens to scan, along the plane, at least some of the plurality of wells in the portion of the plate.