Pure Dye Instrument Normalization for Genomic Analysis
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Solution Overview
Problem
Current genomic analysis faces challenges in comparing results from multiple instruments due to physical variations in components like light sources and optical elements, leading to inconsistent results from identical biological samples.
Innovation Solution
A method and system for normalizing spectra across multiple instruments using a reference instrument and a test instrument, with pure dye plates and filter pairs, where fluorescent spectra are corrected and normalized to minimize variations, involving the use of correction factors and iterative adjustments to achieve consistent multicomponent data.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiple instruments are used for genomic analysis, then productivity increases, but measurement precision deteriorates due to physical variations in components
Solution Approach 1:
The patent introduces pure dye samples as intermediary reference materials that mediate between different instruments. These pure dyes serve as a common reference standard that all instruments can measure, enabling cross-instrument calibration and normalization without requiring direct comparison of all instrument pairs.
Solution Approach 2:
The patent applies parameter changes by measuring fluorescence intensity parameters of pure dyes across different instruments and using these measurements to calculate normalization factors. The physical parameters of the instruments are effectively adjusted through mathematical transformation based on the pure dye reference measurements.
2Reliability
If traditional normalization methods using dye mixtures are used, then instrument variation is reduced, but manufacturing precision deteriorates due to difficulty in controlling dye mixture ratios
Solution Approach 1:
The patent extracts the normalization function from complex dye mixtures and isolates it to pure individual dye samples. By removing the mixture component, the method eliminates the manufacturing complexity and ratio control issues associated with preparing multiple dye mixtures, while maintaining the normalization capability.
Solution Approach 2:
The patent applies local quality by using different pure dye samples for different normalization purposes. Each pure dye serves as a specific reference for calibrating particular wavelength ranges or filter sets, allowing targeted normalization without requiring perfect mixtures of multiple dyes.
3Manufacturing precision
If pure dye normalization is implemented, then manufacturing precision improves, but device complexity increases due to additional normalization steps
Solution Approach 1:
The patent applies preliminary action by pre-measuring the fluorescence spectra of pure dyes under various filter combinations and storing these as reference data. This preliminary characterization allows the normalization factors to be calculated and applied automatically during routine operations, reducing the apparent complexity during actual use.
Solution Approach 2:
The patent creates a digital copy of the pure dye reference spectra and stores it as a lookup table or reference matrix. This copied reference data can be repeatedly used for normalization without requiring physical replication of the pure dye samples, simplifying the process while maintaining precision.
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
The normalization process significantly reduces instrument-to-instrument variability, achieving consistent results within ±8% deviation, improving the reliability and accuracy of experimental comparisons across different instruments.
Implementation Method 1
Each well contains a plurality of dyes where each dye comprises a fluorescent component. Fluorescent spectra are obtained from each instrument for each dye across multiple filter combinations
Data Source
AI summary
The present teachings relate to a method and system for normalizing spectra across multiple instruments. In an embodiment of the present invention, the method comprises at least one reference instrument and a test instrument. Each instrument comprises at least one excitation filter and at least one emission filter arranged in pairs. Each instrument further comprises a pure dye plate comprising a plurality of wells. Each well contains a plurality of dyes where each dye comprises a fluorescent component. Fluorescent spectra are obtained from each instrument for each dye across multiple filter combinations to contribute to a pure dye matrix Mref for the reference instrument and pure dye matrix M for the test instrument. The pure dye spectra can then be multiplied by correction factors for each filter pair to result in corrected pure dye spectra, then normalized and the multicomponenting data can be extracted.