Flow Cytometer Drop Delay Calibration via Waste Stream Fluorescence
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Solution Overview
Problem
Flow cytometers face challenges in accurately determining the drop delay period, which is critical for proper droplet charging and sorting, often resulting in cells of interest ending up in the waste stream or incorrect deflected streams due to incorrect drop delay times.
Innovation Solution
A method and system for automatically selecting a drop delay period by operating a flow cytometer with calibration particles, deflecting and collecting droplets, interrogating fluorescent emissions, and sensing them with a detector to determine the optimal drop delay period with minimal calibration particles in the waste stream, using a waste stream receiver, optical source, and analyzer to select the appropriate delay.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If manual drop delay calibration is used, then operator flexibility is maintained, but measurement precision and reliability deteriorate due to human error
Solution Approach 1:
The system performs self-calibration by automatically determining the optimal drop delay period through fluorescent detection of calibration particles in the waste stream, eliminating the need for manual operator intervention and subjective judgment while maintaining measurement precision
Solution Approach 2:
The system uses fluorescent detection feedback from calibration particles in the waste stream to automatically adjust and determine the optimal drop delay period, creating a closed-loop calibration process that improves measurement precision through iterative optimization
2Reliability
If incorrect drop delay period is used, then sorting speed is maintained, but reliability deteriorates as cells of interest end up in waste stream or incorrect deflected streams
Solution Approach 1:
The system performs preliminary automated calibration using fluorescent detection of calibration particles before actual sorting begins, establishing the correct drop delay period in advance to ensure sorting accuracy without time loss during operation
Solution Approach 2:
The system replaces manual mechanical calibration methods with automated optical detection and electronic control, using fluorescent sensing to determine drop delay period, thereby improving reliability while minimizing calibration time through non-contact, high-speed detection
3Measurement precision
If fluorescent detection in waste stream is implemented, then measurement precision of drop delay period improves, but device complexity increases due to additional optical components
Solution Approach 1:
The fluorescent detection system serves multiple functions: it detects calibration particles in the waste stream for drop delay calibration, and can simultaneously monitor sorting efficiency, making the optical components multi-functional and justifying the added complexity through enhanced measurement precision and additional analytical 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
This approach allows for accurate and automated calibration of the drop delay period, minimizing the number of calibration particles in the waste stream and ensuring proper droplet sorting, reducing errors and increasing the efficiency of the flow cytometry process.
Implementation Method 1
interrogating the waste stream to cause fluorescent emissions of the calibration particles in the waste stream for each drop delay period
Implementation Method 2
sensing the fluorescent emissions from the calibration particles with a detector for each drop delay period
Data Source
AI summary
Disclosed is an automated method and apparatus for automatically setting a drop delay period by detecting calibration particles in a waste stream. The drop delay is incremented over a series of drop delays and the number of calibration particles in the waste stream is detected for each drop delay. The drop delay is selected which has the least number of calibration particles in the waste stream.