Analyte Detection in Hemolyzed Blood Using pH-Controlled RAM Chromatography
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Solution Overview
Problem
Current methods for detecting analytes in whole blood samples, especially those that are hemolyzed, face interference from high concentrations of hemoglobin, which complicates the detection process and requires extensive sample processing, making online detection challenging.
Innovation Solution
The method involves using a buffer with a pH above 8.0 when applying a hemolyzed whole blood sample to a restricted access chromatography material (RAM) to prevent hemoglobin from binding and interfering with the analyte detection, allowing for the direct online detection of analytes like immunosuppressive drugs and folate.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If hemolyzed whole blood sample is applied to RAM column for online detection, then detection speed and efficiency are improved, but hemoglobin interference increases making detection inaccurate
Solution Approach 1:
The patent changes the pH parameter of the mobile phase to above 8.0 (e.g., pH 8.5-9.5 using ammonium bicarbonate or ammonium carbonate buffer). This parameter change modifies the charge state of hemoglobin and RAM surface, preventing hemoglobin adsorption onto the RAM column while maintaining analyte binding capability, thus resolving the contradiction between fast online detection and accurate measurement
Solution Approach 2:
The patent introduces a specific buffer system (ammonium bicarbonate or ammonium carbonate at pH > 8.0) as an intermediary between the hemolyzed blood sample and the RAM column. This buffer mediates the interaction by controlling the electrostatic environment to prevent hemoglobin adsorption, allowing the analyte to be detected accurately without hemoglobin interference
2Measurement precision
If extensive sample processing is performed to remove hemoglobin, then detection accuracy is improved, but processing time and complexity increase
Solution Approach 1:
The patent extracts or removes the harmful factor (hemoglobin) not through complex processing but by changing the mobile phase pH to prevent hemoglobin adsorption in the first place. This eliminates the need for extensive processing steps while maintaining detection accuracy, effectively separating the analyte from hemoglobin interference through pH-controlled selective binding
Solution Approach 2:
The patent performs preliminary action by adjusting the mobile phase pH to >8.0 before sample injection. This preliminary pH adjustment prevents hemoglobin adsorption proactively, eliminating the need for subsequent time-consuming removal steps and enabling direct online detection without extensive 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 effectively reduces hemoglobin interference, enabling efficient and precise online detection of analytes in whole blood samples without the need for extensive sample processing, enhancing clinical diagnostic capabilities.
Implementation Method 1
applying a sample of hemolyzed whole blood known or suspected to contain an analyte of interest to a column comprising a restricted access chromatography material (RAM) thereby binding the analyte
Implementation Method 2
at least in the first step a buffer with a pH above 8.0 is used... ensures that hemoglobin does not interfere with the detection of the analyte of interest
Data Source
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AI summary
The present invention relates to a method of detecting an analyte in a hemolyzed whole blood sample the method comprising the steps of applying a sample of hemolyzed whole blood known or suspected to contain an analyte of interest to a column comprising a restricted access chromatography material (RAM) thereby binding the analyte, eluting the analyte from the RAM and detecting the analyte, wherein at least in the first step a buffer with a pH above 8.0 is used. Preferably the hemolyzed whole blood sample is obtained by differential hemolysis. The novel method ensures that hemoglobin does not interfere with the detection of the analyte of interest, especially in any subsequent analyses. The method can be easily used in the online detection of many analytes, e.g. from a hemolyzed whole blood sample, like in the detection of an antibiotic, of folate or of immunosuppressive drugs, like tacrolimus or sirolimus.