Bisulphite Treatment for Direct Nucleic Acid Detection
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Solution Overview
Problem
Current molecular detection assays require time-consuming and costly sample pre-treatment steps, including cell lysis and nucleic acid purification, which increase processing time and equipment costs, and are not suitable for direct detection of infectious diseases or genetic traits without specialized kits.
Innovation Solution
A molecular detection assay that treats biological samples with a bisulphite agent to disrupt cells and modify nucleic acids, allowing for direct detection of target nucleic acids without the need for pre-treatment, using sodium bisulphite at specific concentrations and pH levels, followed by removal and resuspension in a high-pH elution buffer for PCR amplification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If sample pre-treatment (cell lysis and nucleic acid purification) is performed using commercial kits, then detection sensitivity and reliability are improved, but processing time and equipment costs increase
Solution Approach 1:
The patent combines cell lysis and nucleic acid purification steps into a single integrated bisulphite treatment step. The bisulphite reagent simultaneously disrupts cellular structures and modifies nucleic acids, eliminating the need for separate lysis and purification operations while maintaining detection sensitivity.
Solution Approach 2:
The patent extracts and eliminates the complex commercial kit components (proteases, organic solvents, multiple purification columns) by using a simplified bisulphite-based system that achieves both lysis and purification functions in one reagent system, reducing processing steps and time.
2Reliability
If sample pre-treatment with commercial kits is used, then nucleic acid purification quality is improved, but equipment costs and operational complexity increase
Solution Approach 1:
The patent removes the need for complex commercial kit components including protease enzymes, organic solvents, multiple centrifugation steps, and specialized purification columns. The bisulphite treatment system achieves purification quality without requiring these complex equipment and reagent components.
Solution Approach 2:
The patent changes the chemical parameters of the treatment system by using bisulphite reagent at specific pH and concentration ranges to achieve both cell lysis and nucleic acid purification in a single step, replacing the multi-parameter optimization required by commercial kits.
3Productivity
If conventional sample treatment is used, then cell lysis efficiency is improved for easy-to-lyse samples, but detection capability for difficult-to-lyse samples (parasites, RNA viruses) deteriorates
Solution Approach 1:
The patent optimizes the bisulphite treatment parameters (pH 4.5-5.5, temperature 75-95°C, time 10-30 minutes) to achieve effective cell disruption across diverse sample types including parasites and RNA viruses, while simultaneously maintaining nucleic acid integrity for detection.
Solution Approach 2:
The bisulphite treatment system serves multiple functions simultaneously: it acts as a cell lysis agent, nucleic acid purification reagent, and nucleic acid modification agent. This multi-functional approach enables effective processing of diverse difficult-to-lyse samples without requiring sample-type-specific protocols.
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 simplifies the detection process, reducing sample processing time, eliminating the need for expensive purification kits, and enabling simultaneous detection of RNA and DNA viruses in the same sample, while maintaining sensitivity and efficiency for difficult-to-lyse samples like parasites and RNA viruses, such as hepatitis C virus, and is applicable to various sample types including faecal and serum samples.
Implementation Method 1
treating a biological sample directly with a bisulphite agent under conditions that allow cell disruption and nucleic acid treatment
Implementation Method 2
under conditions that allow cell disruption and nucleic acid treatment
Data Source
AI summary
A molecular detection assay including treating a biological sample directly with a bisulphite agent under conditions that allow cell disruption and nucleic acid treatment; removing the bisulphite agent from the treated sample; and detecting a target nucleic acid in the treated sample.