Flow Cytometer Drop Delay Measurement Using Spatially Separated Lasers
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Solution Overview
Problem
Variability in particle size and drift in flow cytometer components lead to inaccurate predictions of droplets containing particles of interest, resulting in imprecise cell sorting and contamination in flow cytometry.
Innovation Solution
Irradiate a sample with two or more spatially separated lasers, detect light from each particle using multiple photodetector channels, calculate particle velocity, and determine drop delay based on the detected light to adjust droplet charging timing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a single laser is used for particle interrogation, then the device complexity is low, but the measurement precision of drop delay is insufficient due to particle velocity variability
Solution Approach 1:
The single laser interrogation system is segmented into multiple spatially separated lasers (first laser, second laser, and third laser) positioned at different locations along the flow stream. Each laser independently interrogates particles at different positions, enabling velocity calculation through time-of-flight measurement between interrogation points. This segmentation transforms a single measurement point into multiple measurement points, improving drop delay measurement precision while maintaining manageable system complexity through modular laser arrangement.
2Reliability
If particle velocity is not measured, then the system operation is simple, but the predictability of droplet charging timing is poor due to velocity variability
Solution Approach 1:
The system performs preliminary velocity measurement of particles using multiple lasers before the droplet charging decision is made. By interrogating particles at multiple positions upstream, the system calculates particle velocity in advance, which is then used to predict the exact timing when the particle-containing droplet will reach the charging point. This preliminary action ensures reliable droplet charging timing prediction while maintaining operational simplicity through automated velocity-based timing adjustment.
3Manufacturing precision
If fixed drop delay timing is used, then the device operation is simple, but sorting precision deteriorates due to drift in flow cytometer components
Solution Approach 1:
The system transitions from fixed drop delay timing to dynamic, particle-specific timing control. By continuously measuring particle velocity through multiple laser interrogation points and calculating individual drop delay values based on each particle's velocity, the system dynamically adjusts charging timing to compensate for drift in flow cytometer components. This dynamic approach maintains high sorting precision while managing complexity through real-time velocity-based timing calculation and adjustment.
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 precise and consistent particle sorting by dynamically determining drop delay for each particle, improving sorting accuracy and reducing contamination.
Implementation Method 1
As particles of interest (e.g., cells) move through the interrogation point, light from the irradiation source (e.g., laser) is scattered.
Data Source
AI summary
Aspects of the present disclosure include methods for determining drop delay of a particle in a flow stream (e.g., in a particle analyzer). Methods according to certain embodiments include irradiating the particle in the flow stream with two or more spatially separated lasers, detecting light from the particle in a first photodetector channel and a second photodetector channel calculating a velocity of the particle in the flow stream based on the detected light in the first photodetector channel and the second photodetector channel and determining the drop delay of the particle based on the calculated velocity. Systems (e.g., particle analyzers) having a light source with two or more spatially separated lasers and a light detection system for practicing the subject methods are also described. Integrated circuits and non-transitory computer readable storage medium having instructions stored thereon for determining drop delay according to the subject methods are also provided.