Blood Analysis Dilution Accuracy via Dual Standard Buffer
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Solution Overview
Problem
Conventional methods for analyzing trace amounts of blood components using internal or external standard substances face challenges such as enzyme inhibition, adsorption issues, and denaturation during storage and transportation, leading to inaccurate dilution ratios and measurement errors.
Innovation Solution
A method involving the use of both internal and external standard substances, where lithium or glycerol-3-phosphate is used as an internal standard and sodium as an external standard, in a buffer with specific osmolality and composition to accurately determine the dilution factor and quantify biological components in blood samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If glycerol-3-phosphate is used as an internal standard substance in the diluted solution of plasma, then the dilution ratio can be calculated, but the internal standard substance is hydrolyzed by alkaline phosphatase to produce glycerol, leading to inaccurate dilution ratio when stored for long time or at high temperature
Solution Approach 1:
The patent extracts and removes the problematic internal standard substance (glycerol-3-phosphate) that undergoes enzymatic hydrolysis by alkaline phosphatase. By eliminating this substance from the measurement system, the patent avoids the accuracy degradation that occurs during storage, while maintaining the ability to calculate dilution ratios using alternative stable substances.
Solution Approach 2:
The patent employs a disposable whole blood collection device that integrates the collection tube with a pre-added buffer solution. This disposable system eliminates the need for separate storage and handling of reagents, reducing the time and conditions under which enzymatic reactions could occur, thereby maintaining measurement accuracy without requiring long-term storage stability of labile substances.
2Object-generated harmful factors
If EDTA and phosphoric acid are added to inhibit enzymes, then enzyme inhibition is achieved, but the activity of aspartate aminotransferase or alanine aminotransferase decreases
Solution Approach 1:
The patent converts the harmful effect of endogenous alkaline phosphatase (which hydrolyzes the internal standard) into a beneficial situation by using a different internal standard substance that is not susceptible to hydrolysis. This approach eliminates the need to add enzyme inhibitors like EDTA and phosphoric acid, thereby avoiding their harmful side effect of reducing aminotransferase activity and enabling accurate enzyme activity measurement.
Solution Approach 2:
The patent introduces a new internal standard substance that acts as an intermediary - one that does not interact with alkaline phosphatase but still allows dilution ratio calculation. This intermediary substance mediates between the need for dilution measurement and the presence of enzymatic activity, avoiding the conflict that previously required enzyme inhibitors.
3Measurement precision
If excess pyridoxal phosphate is added to maintain enzyme activity, then enzyme activity is preserved, but the complexity of the buffer composition increases
Solution Approach 1:
The patent extracts and removes the need for excess pyridoxal phosphate addition by eliminating the requirement for enzyme inhibition. By using a stable internal standard substance that does not undergo hydrolysis, the patent eliminates the entire chain of events that led to the need for enzyme inhibitors and subsequently excess enzyme activators, simplifying the buffer composition while maintaining measurement precision.
4Adaptability or versatility
If a trace amount of blood is diluted with a buffer containing an internal standard substance, then quantification can be performed at any time and place, but the internal standard substance may be adsorbed to a plastic container during long-term storage
Solution Approach 1:
The patent employs a disposable whole blood collection device that is used immediately upon collection. This short-living, single-use system eliminates long-term storage of the internal standard substance in plastic containers, thereby preventing adsorption issues while maintaining the flexibility of point-of-care testing.
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 and reliable analysis of blood components, reducing measurement errors and maintaining stability during storage and transportation, enabling accurate quantification of biological components and enzyme activity.
Implementation Method 1
a biological sample such as blood is diluted with a buffer having a specific composition, and a plasma sample component or enzyme activity is analyzed from the mixed solution in the diluted sample
Implementation Method 2
glycerol-3-phosphate is hydrolyzed by alkaline phosphatase, which is an enzyme present in vivo, to produce glycerol
Implementation Method 3
lithium or glycerol-3-phosphate is used as an internal standard and sodium as an external standard, in a buffer with specific osmolality and composition to accurately determine the dilution factor and quantify biological components
Data Source
AI summary
There is provided a method of analyzing a biological sample component that allows easy and accurate quantification and counting of any of a plasma component and a blood cell component in a trace and unknown amount of a whole blood sample collected from a finger, for example. The method of the present invention is a method of analyzing a biological sample component in a trace amount of blood, comprising analyzing a diluent buffer into which the blood has been mixed and an internal standard substance and/or an external standard substance contained in the diluent buffer, calculating a dilution ratio, and analyzing a biological component in a plasma or serum component in the blood.