Microbial Enumeration via SYBR Gold Fluorometry
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Solution Overview
Problem
Current methods for counting microbes in environmental samples are slow, require expensive equipment, and struggle with low microbial concentrations, making them unsuitable for rapid and cost-effective monitoring in fields like biological warfare defense, water quality assessment, and microbial ecology.
Innovation Solution
The use of SYBR Gold nucleic acid stain and low protein binding filters in conjunction with fluorometry to rapidly estimate microbial numbers in samples, including environmental and laboratory samples, allowing for high-throughput enumeration of microbial cells without culturing, and the inclusion of DNase I to reduce background noise.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional epifluorescent microscopy is used to count microbes, then measurement precision can be achieved, but the method is slow and requires expensive equipment
Solution Approach 1:
The patent replaces the mechanical microscopy system with a fluorometric measurement system. Instead of using epifluorescent microscopy which requires expensive equipment and manual counting, the invention uses SYBR Gold nucleic acid stain combined with fluorometry to automatically detect and quantify microbial cells, thereby improving productivity while maintaining measurement precision
Solution Approach 2:
The patent changes the detection parameter from optical microscopy to fluorometric detection. By using SYBR Gold stain that binds to nucleic acids and emits fluorescence, the system can detect microbial cells through fluorescence intensity measurements, enabling high-throughput automated counting while preserving accuracy
2Reliability
If traditional methods are used to detect microbes, then detection capability is maintained, but the methods struggle with low microbial concentrations
Solution Approach 1:
The patent enhances detection sensitivity by changing from non-fluorescent or weakly fluorescent stains to SYBR Gold, which provides strong fluorescence signal. This parameter change in staining intensity allows reliable detection and precise quantification of microbial cells even at low concentrations in environmental samples
Solution Approach 2:
The patent uses SYBR Gold nucleic acid stain as an intermediary that amplifies the detection signal. The stain binds to microbial nucleic acids and produces a strong fluorescence signal that can be detected by fluorometers, enabling reliable detection of low-concentration microbial populations that would be difficult to detect with traditional methods
3Productivity
If high-throughput methods are implemented, then productivity increases, but device complexity increases
Solution Approach 1:
The patent replaces complex mechanical counting systems with a simpler fluorometric detection system. By using automated fluorometry instead of manual microscopy, the system achieves high-throughput enumeration while actually reducing operational complexity, as the fluorometric method requires simpler sample preparation and automated data processing
Solution Approach 2:
The patent implements a self-service system where the fluorometer automatically measures fluorescence intensity and the system自行 calculates microbial cell concentrations. This automation eliminates the need for manual counting and complex operational procedures, achieving high throughput while keeping the system easy to operate
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 precise and rapid enumeration of microbial cells at low concentrations, offering higher precision than traditional epifluorescent microscopy and being adaptable for automatic estimation in various samples, including water and soil environments, with potential for field-based applications.
Implementation Method 1
adding SYBR Gold nucleic acid stain, or equivalent; measuring the total fluorescence, thereby enumerating (counting) microbial cells in the sample
Implementation Method 2
filtering the liquid or water sample with at least one low protein binding filter
Data Source
AI summary
The invention provides compositions (e.g., kits) and methods for determine the number of bacteria and other microbes in samples having low concentrations of microbes, for use, e.g., in biological warfare defense, microbe detection and agricultural and environmental sciences.