Particle-Based CE-SELEX Aptamer Screening
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Solution Overview
Problem
Current nucleic acid aptamer screening methods, such as CE-SELEX, face challenges in designing experimental conditions and are limited by the size of target molecules, leading to low yield rates and difficulty in detecting specific complexes, especially for low molecular compounds.
Innovation Solution
A method combining particles with capillary electrophoresis (CE-SELEX) is developed, where a target molecule is immobilized on a solid phase support, allowing for easier collection and amplification of nucleic acid aptamer candidates using PCR, without requiring expensive fluorescence detectors, and enabling the selection of aptamers for various target molecules, including low molecular weight compounds.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional CE-SELEX method is used, then nucleic acid aptamer can be screened, but the yield rate is low (30% or less) and the design of collection region is difficult
Solution Approach 1:
The patent introduces a solid phase support as an intermediary carrier to immobilize target molecules. This mediator simplifies the collection region design by providing a fixed position for target molecules, eliminating the need for complex electrophoretic mobility-based collection region determination. The solid phase support acts as a stable platform that enables straightforward washing and elution operations.
Solution Approach 2:
The patent replaces the complex electrophoretic separation-based collection method with a simple mechanical washing and elution process. Instead of relying on electrophoretic mobility differences to define collection regions, the invention uses physical immobilization of target molecules on solid phase supports, allowing direct washing away of unbound sequences and simple elution of bound aptamers.
2Difficulty of detecting and measuring
If conventional CE-SELEX method is used, then aptamer screening is possible, but expensive fluorescence detectors are required for detecting target-aptamer complexes
Solution Approach 1:
The patent replaces the complex fluorescence detection system with a simple mechanical washing and elution process. Detection is achieved through direct observation of aptamer binding to immobilized targets during washing steps, eliminating the need for expensive fluorescence detectors while maintaining sufficient detection capability for screening purposes.
Solution Approach 2:
The patent uses inexpensive solid phase supports (such as magnetic beads or microtiter plate wells) that can be easily disposed of or regenerated, replacing expensive and complex fluorescence detection equipment. The solid phase supports serve as single-use or limited-use carriers that eliminate the need for costly detection instruments.
3Adaptability or versatility
If conventional CE-SELEX method is used, then aptamer screening can be performed, but it is limited to target molecules of certain size due to electrophoretic mobility requirements
Solution Approach 1:
The patent creates a universal platform using solid phase supports that can immobilize various types of target molecules (proteins, low molecular weight compounds, cells) regardless of their size. The same basic washing and elution protocol can be applied to different target types, making the method universally applicable and eliminating size-based limitations of electrophoretic methods.
Solution Approach 2:
The patent changes the fundamental parameter of target molecule presentation from free-floating (requiring electrophoretic mobility) to immobilized state on solid phase supports. This parameter change allows target molecules of any size to be effectively used, as immobilization eliminates the need for electrophoretic separation and enables direct binding and washing operations.
4Reliability
If conventional CE-SELEX method is used, then aptamer screening is possible, but sequences with low binding ability may be included in the collection region due to dissociation during electrophoresis
Solution Approach 1:
The patent performs preliminary immobilization of target molecules on solid phase supports before introducing the aptamer library. This preliminary action ensures that only sequences with sufficient binding ability can withstand the washing steps, as weakly binding sequences will be removed during washing. The immobilization is performed in advance, allowing rigorous washing to eliminate non-specific binders before elution of high-affinity aptamers.
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 simplifies the design of collection regions, improves aptamer yield rates, and allows for the selection of aptamers with higher binding affinity, reducing the number of required rounds and eliminating non-specific sequences, thereby enhancing the efficiency of nucleic acid aptamer screening.
Implementation Method 1
collecting the nucleic acid aptamer candidate binding with the target molecule by a capillary electrophoresis
Implementation Method 2
amplifying the nucleic acid aptamer candidate by PCR
Data Source
Figure 1A~1B
Figure 2A~2L
Figure 3A~3J
AI summary
The present invention relates to a method for screening a nucleic acid aptamer comprising: (a) contacting a target molecule immobilized on a solid phase support with a nucleic acid aptamer candidate; (b) collecting the nucleic acid aptamer candidate binding with the target molecule by a capillary electrophoresis; and (c) amplifying the nucleic acid aptamer candidate by PCR.