Selective Nucleic Acid Separation via Aqueous Lysis and Filtration
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Solution Overview
Problem
Current methods for enriching nucleic acids from rare cells are hindered by low purity and high contamination from non-disease-associated nucleic acids, leading to insensitive and prone-to-false-results analysis, and fixation processes further complicate RNA integrity and recovery.
Innovation Solution
A method involving an aqueous medium that selectively releases nucleic acids from non-rare cells while preserving those from rare cells, followed by filtration to separate and concentrate intact nucleic acids from rare cells, using a porous matrix to retain rare cells and allow non-rare cells and released nucleic acids to pass through.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If cell filtration is performed without fixation to preserve nucleic acid integrity, then RNA quality is improved, but recovery of diseased cells decreases to less than 40%
Solution Approach 1:
The patent applies preliminary action by performing fixation after filtration rather than before. The method first separates cells through filtration while they are intact and viable, then applies fixation to the collected rare cells. This sequence preserves nucleic acid integrity during the critical separation phase while still achieving high recovery rates through subsequent fixation of the concentrated rare cell population.
2Productivity
If fixation is applied to increase recovery of diseased cells to greater than 90%, then productivity is improved, but nucleic acids become heavily fragmented and chemically modified
Solution Approach 1:
The patent reverses the conventional sequence by performing filtration before fixation. Rare cells are first isolated through filtration while maintaining their natural state, ensuring nucleic acids remain intact. Only after the rare cells are collected and concentrated does the method apply fixation, thereby achieving high recovery rates without compromising nucleic acid quality for downstream applications.
3Quantity of substance
If nucleic acids are concentrated from samples with few rare cells, then the amount of nucleic acids is increased, but purity decreases due to contamination from non-disease-associated nucleic acids
Solution Approach 1:
The patent applies segmentation by separating the enrichment process into two distinct stages: first, physical separation of rare cells from the bulk sample through filtration; second, concentration of nucleic acids from the isolated rare cells. This segmentation ensures that nucleic acids are concentrated only from the purified rare cell population, excluding non-disease-associated nucleic acids from non-rare cells, thereby achieving both high concentration and high purity.
Solution Approach 2:
The method performs preliminary cell separation before nucleic acid concentration. By first filtering to isolate rare cells and remove the majority of non-rare cells, the subsequent nucleic acid extraction and concentration steps operate on a pre-purified cell population. This preliminary action prevents contamination from non-disease-associated nucleic acids while still achieving sufficient nucleic acid concentration for detection.
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 enhances the recovery and purity of disease-associated nucleic acids, enabling sensitive and accurate determination of rare cell nucleic acids, even at low concentrations, and maintains RNA integrity for downstream applications.
Implementation Method 1
The combination is held for a period of time and at a temperature for selectively releasing nucleic acids from the non-rare cells but not from the rare cells
Implementation Method 2
The sample is subjected to filtration to separate rare cells from non-rare cells
Data Source
AI summary
Nucleic acids in non-rare cells in a sample containing non-rare cells and rare cells are selectively released from the non-rare cells. The sample is combined with an aqueous medium, and the combination is held for a period of time and at a temperature for selectively releasing nucleic acids from the non-rare cells but not from the rare cells. The sample is subjected to filtration to separate rare cells from non-rare cells. Rare cells with intact nucleic acids are separated from non-rare cells and nucleic acids from the non-rare cells. Nucleic acids from the rare cells are subjected to one or more identification techniques either with or without extraction of nucleic acids from the rare cells.