Flow Cytometry Viability Quantification for Lactic Acid Bacteria
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Solution Overview
Problem
Current methods for quantifying viable lactic acid bacteria cells are labor-intensive and difficult to automate, leading to inefficiencies in high-throughput screening and inconsistent results due to sensitivity to assay conditions.
Innovation Solution
A flow cytometry-based method that stabilizes the cellular membrane potential of lactic acid bacteria cells by mixing them with a dye and a diluted complex growth medium, allowing for reliable and fast quantification of viable cells through optical detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If the plate count technique is used to determine viable cells, then the measurement is considered reliable, but the process is labor-intensive and difficult to automate
Solution Approach 1:
The patent replaces the manual mechanical process of plate counting with an automated flow cytometry system that uses optical detection and electronic data processing to quantify viable cells, thereby eliminating the labor-intensive nature of the traditional method while maintaining measurement reliability
Solution Approach 2:
The patent changes the detection parameter from visual colony counting to automated optical property measurement of dye-associated cells, enabling high-throughput automated analysis while preserving the reliability of viability assessment
2Reliability
If the plate count technique is used to determine viable cells, then the measurement is considered reliable, but the productivity is low
Solution Approach 1:
The patent substitutes the slow manual plate counting process with automated flow cytometry that can analyze thousands of cells per second, dramatically increasing productivity while maintaining the reliability of viability measurements through objective optical detection
Solution Approach 2:
The patent enables continuous automated analysis of cell samples through flow cytometry, eliminating the intermittent nature of manual plate counting and allowing for high-throughput continuous measurement of viable cells
3Loss of time
If cells are kept for extended periods for analysis, then more time is available for measurement, but the membrane potential decreases or is lost
Solution Approach 1:
The patent applies preliminary staining with the membrane potential indicator dye before analysis, allowing cells to be prepared in advance and stored for extended periods while maintaining stable membrane potential signals, thus enabling flexible scheduling without compromising measurement reliability
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 method enables resilient and high-throughput viability testing, maintaining cell membrane potential for extended periods, thus providing consistent results across varying conditions and improving the efficiency of lactic acid bacteria cell quantification.
Implementation Method 1
The dye stains bacteria with green fluorescence, which presumably reflects the distribution of dye toward lipids in the membrane and hydrophobic constellations of the cell
Implementation Method 2
The electrical potential (inside negative) changes the distribution of DiOC via an inward electroforesis
Implementation Method 3
the forward angle light scatter (FSC), the side-angle light scatter (SSC), and the fluorescence of individual cells at selected wavelengths are measured
Data Source
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AI summary
The present invention pertains to a method of quantifying the number of viable lactic acid bacteria (LAB) cells in a sample.