Automated Fluorescence Imaging System with Quenching Unit
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Solution Overview
Problem
Current immunofluorescence analysis techniques are labor-intensive and limited in the number of images that can be obtained from a biological sample due to manual handling steps, restricting the multiplexing potential and image resolution.
Innovation Solution
An automated system that sequentially applies fluorescent reagents to a biological sample using a robotically controlled pipetting system, combines fluorescence imaging with a quenching unit emitting different wavelengths of light to eliminate fluorescence signals, allowing for efficient detection and quenching of fluorescence signals in orthogonal dimensions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If manual handling steps are used for sequential staining and imaging, then the process is flexible and easy to set up, but the number of images obtainable from a single sample is limited and labor-intensive
Solution Approach 1:
The system enables self-service automation where the automated instrument performs sequential staining, imaging, and quenching operations without manual intervention. The robotic arm automatically transfers reagents, the imaging system captures images, and the quenching system eliminates fluorescence signals, allowing the system to serve itself through programmed routines.
Solution Approach 2:
Manual mechanical handling operations are replaced with an automated robotic system. The robotic arm with pipettes substitutes human hands for reagent transfer, the automated imaging system replaces manual microscope operation, and the quenching system automates the chemical treatment process, collectively replacing the entire manual mechanical handling workflow.
2Adaptability or versatility
If multiple fluorescent reagents are applied sequentially to increase multiplexing, then more antigens can be detected, but the time required between steps increases and sample manipulation becomes more complex
Solution Approach 1:
The system maintains continuous useful action by immediately proceeding from one staining step to the next without idle time. The robotic arm continuously transfers reagents, the imaging system continuously captures images, and the quenching system continuously processes samples, eliminating idle time between operations and maintaining uninterrupted workflow.
Solution Approach 2:
Multiple fluorescent reagents are prepared in advance in separate wells of a microtiter plate before the staining process begins. The robotic arm is pre-programmed with the sequence of reagent applications, and all necessary components are positioned beforehand, allowing the system to execute the multiplexing protocol without delays for preparation or setup during the actual staining process.
3Measurement precision
If image resolution is increased to improve quality of information, then better detection precision is achieved, but the time required for imaging and the complexity of handling increases
Solution Approach 1:
The imaging process uses periodic action by capturing images at specific intervals during the staining protocol rather than continuously. The system images at key moments (after each staining step) and uses quenching between images to eliminate background fluorescence, allowing high-resolution imaging at discrete time points without requiring continuous high-resolution capture throughout the entire process.
4Adaptability or versatility
If more staining cycles are performed on a single sample, then multiplexing increases, but the sample undergoes more manipulation which may affect sample integrity
Solution Approach 1:
The quenching solution acts as an intermediary between staining cycles. It temporarily suspends the fluorescent signal and stabilizes the sample between imaging events, preventing carryover effects and maintaining sample integrity during multiple staining cycles. The quenching step mediates the transition between different staining conditions without compromising the biological sample.
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-throughput analysis of biological samples with improved image resolution and multiplexing potential by automating the staining, imaging, and quenching processes, significantly increasing the number of images that can be obtained from a single sample.
Implementation Method 1
a fluorescence system that comprises an excitation unit and a detection unit for fluorescence signals obtained from the fluorescent dye
Implementation Method 2
the fluorescence may be quenched by a second optical system disposed laterally adjacent to the fluorescence imaging system, which irradiates the sample with sufficient light to disable or destroy the fluorescing moiety
Data Source
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AI summary
Systems and methods are described for analyzing a plurality of biological samples with a plurality of fluorescent reagents in an automated fashion. The system may include two optical systems, a fluorescence imaging system and an optical quenching system. These two systems may be placed laterally adjacent to one another, and generally beneath one of the wells of sample-containing microtiter plate. The biological sample contained therein may be stained with a series of fluorescent reagents, with the fluorescence quenched between stainings. By precise positioning of the sample with respect to the imaging system, the sample may be imaged with a plurality of serially applied reagents.