Insulin Mass Spectrometry Quantitation Without Immunopurification
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing methods for quantifying insulin, particularly in diabetic and pre-diabetic patient samples, face challenges in accuracy and efficiency, especially when immunological techniques and conventional mass spectrometric methods fail to provide precise measurements without immunopurification steps.
Innovation Solution
The method employs tandem mass spectrometry and high resolution/high accuracy mass spectrometry to determine insulin levels by enriching samples through solid phase extraction and high performance liquid chromatography, generating insulin ions under specific conditions, and analyzing these ions without prior immunopurification, using techniques such as electrospray ionization and collision-induced dissociation to identify specific mass-to-charge ratios.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Difficulty of detecting and measuring
If immunological techniques are used for insulin quantitation, then measurement capability is improved, but measurement precision deteriorates due to cross-reactivity and lack of specificity
Solution Approach 1:
The patent replaces immunological detection methods with mass spectrometry-based detection. Instead of using antibodies that suffer from cross-reactivity issues, the invention uses mass-to-charge ratio measurement to specifically identify and quantify insulin molecules, thereby eliminating cross-reactivity problems while maintaining detection capability
Solution Approach 2:
The patent changes the detection parameter from antibody binding affinity to mass-to-charge ratio. By measuring the precise mass of insulin ions (e.g., [M+6H]6+ at m/z 968.95) and their fragmentation patterns, the method achieves high specificity without the cross-reactivity issues inherent in immunological methods
2Ease of operation
If conventional mass spectrometric methods are used without immunopurification, then ease of operation is improved, but measurement precision deteriorates due to insufficient enrichment
Solution Approach 1:
The patent performs preliminary enrichment of insulin using solid phase extraction and HPLC before mass spectrometry analysis. This preliminary action concentrates insulin from complex biological matrices, ensuring sufficient signal intensity and measurement precision while maintaining operational simplicity by automating the enrichment steps
Solution Approach 2:
The patent introduces solid phase extraction cartridges and HPLC columns as intermediary steps between sample collection and mass spectrometry analysis. These intermediaries selectively retain and concentrate insulin molecules, enabling precise measurement without requiring complex immunopurification procedures
3Measurement precision
If immunopurification steps are added to mass spectrometry methods, then measurement precision is improved, but device complexity and loss of time worsen
Solution Approach 1:
The patent replaces complex immunopurification systems with simpler solid phase extraction and HPLC systems coupled directly to mass spectrometry. This substitution maintains measurement precision through selective enrichment while significantly reducing device complexity and procedural steps
Solution Approach 2:
The patent extracts and enriches insulin from complex biological samples using solid phase extraction and HPLC, separating insulin from interfering substances. This extraction approach achieves the necessary purification for precise measurement without requiring the complex multi-step immunopurification process
4Measurement precision
If immunopurification steps are added to mass spectrometry methods, then measurement precision is improved, but productivity deteriorates
Solution Approach 1:
The patent performs preliminary insulin enrichment using automated solid phase extraction and HPLC methods that can process multiple samples in sequence. This preliminary enrichment ensures sufficient insulin concentration for precise measurement while maintaining high throughput, achieving both precision and productivity
Solution Approach 2:
The patent implements continuous flow HPLC coupled online to mass spectrometry, eliminating the need for manual sample handling between enrichment and analysis steps. This continuous operation maintains measurement precision while significantly improving productivity by enabling uninterrupted sample processing
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 allows for precise quantitation of insulin in biological samples, achieving high sensitivity and accuracy, enabling reliable diagnostic and treatment monitoring at insulin levels ranging from 10 ρIU/mL to 500 μIU/mL without the need for immunopurification, thus overcoming limitations of previous methods.
Implementation Method 1
subjecting a sample to solid phase extraction
Implementation Method 2
high performance liquid chromatography to obtain a fraction enriched in insulin
Implementation Method 3
subjecting the enriched insulin to an ionization source under conditions suitable to generate one or more insulin ions
Implementation Method 4
determining the amount of one or more insulin ions by tandem mass spectrometry
Implementation Method 5
one or more fragment ions from an insulin precursor ion with m/z of 1162.5±0.5 and one or more fragment ions from an insulin precursor ion with m/z of 968.9±0.5
Data Source
AI summary
Methods are described for determining the amount of insulin in a sample. More specifically, mass spectrometric methods are described for detecting and quantifying insulin in a biological sample utilizing purification methods coupled with tandem mass spectrometric or high resolution/high accuracy mass spectrometric techniques.