Scanning Molecule Counting Speed Adjustment for Probe Discrimination
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Solution Overview
Problem
In optical analysis techniques like the Scanning Molecule Counting Method, unbound light-emitting probes can be erroneously detected as bound probes due to stochastic transitions between non-light-emitting and light-emitting states, reducing precision and requiring unnecessary removal processes.
Innovation Solution
Adjusting the moving speed of the light detection region to ensure the encompassing time for unbound probes exceeds their average lifetime in the light-emitting state, allowing differentiation between unbound and bound probe signals, thereby reducing erroneous detection and eliminating the need for unbound probe removal.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If the light detection region moves quickly through the sample solution, then the measurement time is reduced and productivity is improved, but unbound probes in light-emitting states may be erroneously detected as bound probes, reducing measurement precision
Solution Approach 1:
The patent changes the temporal parameter of the detection process by adjusting the moving speed of the light detection region. By optimizing the scanning speed to match the lifetime characteristics of fluorescent states, the system can distinguish between transient unbound probe emissions and sustained bound probe emissions, thereby maintaining high measurement speed while improving detection accuracy through parameter optimization
Solution Approach 2:
The patent introduces dynamic control of the light detection region's movement through the sample solution. The scanning speed is dynamically adjusted based on the fluorescent lifetime characteristics of the probes, allowing the system to adapt the measurement process to the temporal behavior of the fluorescent signals, thus resolving the contradiction between speed and precision
2Measurement precision
If unbound probes are removed from the sample solution before measurement, then measurement precision is improved, but the device complexity and operation complexity increase due to additional removal processes
Solution Approach 1:
The patent extracts the probe separation function from the physical removal process and transfers it to the temporal discrimination mechanism. By using the fluorescent lifetime difference between bound and unbound probes as a discrimination criterion, the system eliminates the need for physical separation steps while maintaining the ability to distinguish specific binding events from non-specific background signals
Solution Approach 2:
The patent replaces the mechanical/physical probe removal system with an optical-temporal discrimination system. Instead of using physical methods to separate unbound probes, the system uses the temporal characteristics of fluorescent emission to distinguish bound from unbound probes, thereby eliminating complex removal apparatus and simplifying the overall measurement process
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 significantly reduces the likelihood of erroneous detection of unbound probes, enhancing the accuracy of light-emitting particle detection and eliminating the need for unbound probe removal, which is particularly advantageous for rare or expensive samples.
Implementation Method 1
the light emitted from a light-emitting particle is individually detected... the light emitted from the light-emitting particle may be fluorescence
Implementation Method 2
detection and/or measurement of faint light at the single photon or single fluorescent molecule level have become possible by using an optical system of a confocal microscope and a super high sensitive light detection technique capable of the photon counting
Implementation Method 3
in which probe there occurs a stochastic transition between a non-light-emitting state that no light is emitted in the wavelength band of the detected light and a light-emitting state that light is emitted in the wavelength band of the detected light
Data Source
AI summary
In the scanning molecule counting method of measuring light intensity from a light detection region while moving the position of the light detection region of a confocal or multiphoton microscope in a sample solution containing light-emitting particles, generating time series light intensity data and detecting each of signals of the light-emitting particles individually in the data, wherein the light-emitting particles are formed by binding to a particle to be observed a light-emitting probe which emits light through binding to the particle to be observed and in which a stochastic transition between a non-light-emitting state and a light-emitting state occurs in the unbound state, the moving speed of the position of the light detection region is adjusted to make the time during which the unbound probe is encompassed by the moving light detection region longer than an average lifetime during which the probe is in the light-emitting state.