Capillary Electrophoresis Internal Standard Charge Adjustment
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Solution Overview
Problem
In capillary electrophoresis, variations in migration time due to electrophoresis conditions make it difficult to accurately identify substances, especially when the earliest detected peak is mistakenly recognized as the internal standard substance, leading to errors in estimating the size of RNA or DNA.
Innovation Solution
A capillary electrophoresis method where an internal standard substance with a positive or negative net charge of 2 or more is used, which migrates faster than the analysis target, allowing the earliest detected peak to be identified as the internal standard substance, and subsequent peaks are corrected based on this, using fluorescent substances like 5(6)-carboxyfluorescein, 2', 7'-bis (2-carboxyethyl)-5(and-6)-carboxyfluorescein, and 5-carboxy-(and-6)-fluorescein-sulfonic acid in an electrolyte solution with a pH of 5.0 to 10.0.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If an internal standard substance is used for migration time correction, then measurement precision is improved, but identification accuracy deteriorates when the internal standard peak is mistakenly recognized
Solution Approach 1:
The invention changes the charge parameter of the internal standard substance to +2 or more, which fundamentally alters its migration behavior. This parameter change ensures the internal standard migrates faster than oligo-DNA samples, resolving the identification conflict where the internal standard peak was mistakenly recognized as the earliest sample peak.
2Productivity
If conventional internal standard substances are used, then migration time correction can be performed, but peak identification errors occur due to detection order variations
Solution Approach 1:
The invention performs preliminary action by pre-determining the migration order through charge selection. By choosing an internal standard with charge of +2 or more, the migration order is predetermined (internal standard first, then samples), eliminating the need for post-acquisition peak identification and preventing identification errors.
3Ease of operation
If the earliest detected peak is automatically recognized as the internal standard, then operation simplicity is improved, but identification reliability deteriorates when oligo-DNA migrates faster
Solution Approach 1:
The invention changes the charge parameter of the internal standard to +2 or more, which ensures it always migrates faster than oligo-DNA samples under normal electrophoresis conditions. This parameter change makes the automatic recognition rule (earliest peak = internal standard) reliable, resolving the contradiction between operational simplicity and identification accuracy.
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 accurate identification of peaks by ensuring the internal standard substance-derived peak is recognized correctly, thereby improving the estimation of RNA or DNA sizes by reducing errors caused by premature detection of oligo-DNA peaks.
Implementation Method 1
Capillary electrophoresis is a technique in which a certain voltage is applied across opposite ends of a capillary having an inner diameter of about 100 μm to electrophorese substances in the capillary filled with an electrolytic solution
Implementation Method 2
the internal standard substance consists of a fluorescent substance
Data Source
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AI summary
Disclosed is a capillary electrophoresis method for simultaneously electrophoresing an unknown sample and an internal standard material, determining an earliest peak as a peak derived from the internal standard material, and identifying the unknown sample on the basis of the internal standard substance-derived peak. The internal standard substance consists of a fluorescent substance or a salt thereof which has a positive or negative net charge of 2 or more in an electrolyte solution used for capillary electrophoresis. An earliest one of detected peaks is determined as a peak derived from the internal standard substance, and the remaining peaks are identified on the basis of the internal standard substance-derived peak.