Lyophilized Amplification Microspheres for Flexible Template Loading
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Solution Overview
Problem
Existing nucleic acid amplification methods require aqueous reconstitution, limiting template addition flexibility, affecting reaction sensitivity and specificity, and are inflexible in reagent use and storage/transport conditions.
Innovation Solution
A microsphere preparation is developed by lyophilizing amplification reagents, allowing direct use with templates in a stable supersaturated state, enhancing sensitivity and specificity without aqueous reconstitution.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If aqueous reconstitution is used to prepare amplification reagents, then the reagents can be stored and transported in a stable form, but the flexibility of template addition is limited and the adjustable range of template amount is narrow
Solution Approach 1:
The patent changes the physical state parameter of amplification reagents from aqueous solution to lyophilized microsphere form. This parameter change enables the reagents to maintain stability in a dry state while allowing flexible template addition when reconstituted, as the lyophilized form can accommodate varying template amounts without the constraints of pre-prepared aqueous solutions.
Solution Approach 2:
The patent applies preliminary action by pre-freezing and lyophilizing the amplification reagents into microsphere form before use. This preliminary preparation stabilizes the reagents for storage and transport, while the microsphere format allows users to add templates flexibly at the time of use, combining advance stabilization with operational flexibility.
2Adaptability or versatility
If templates are added without considering concentration changes of effective components, then template addition flexibility increases, but the specificity of the reaction decreases
Solution Approach 1:
The patent implements feedback control by providing clear guidance on the relationship between template addition amount and effective component concentration. The system monitors and controls the concentration changes of effective components based on template addition, ensuring that specificity is maintained while allowing flexible template addition within defined parameters.
3Stability of the object's composition
If in-situ lyophilizing is used to prepare amplification reagents, then storage and transportation conditions are improved, but the flexibility of reagent use and compatible models are limited
Solution Approach 1:
The patent segments the amplification reagents into discrete microsphere units through lyophilization. Each microsphere contains the necessary reagents in a stabilized dry form, improving storage and transportation stability. The segmented microsphere format also enhances flexibility by allowing users to add different templates and adjust reaction conditions for various compatible models.
4Ease of operation
If amplification reagents are stored in aqueous form, then they can be used immediately, but the requirements on storage and transportation conditions are stringent
Solution Approach 1:
The patent utilizes phase transition by converting amplification reagents from aqueous solution phase to solid lyophilized microsphere phase for storage and transportation. This phase change stabilizes the reagents, reducing stringency of storage and transportation conditions. When use is required, the microspheres are reconstituted back to aqueous form, maintaining immediate usability.
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
The microsphere preparation improves nucleic acid amplification sensitivity and specificity by stabilizing primer concentrations, enabling flexible template addition and maintaining reaction accuracy.
Implementation Method 1
a microsphere preparation for nucleic acid amplification, and the microsphere preparation comprises reaction microspheres obtained by lyophilizing the mixed reagents required in an amplification reaction
Data Source
AI summary
The present disclosure relates to the technical field of molecular diagnosis, and particularly, to a microsphere formulation for nucleic acid amplification, an amplification method, and use in joint detection. The disclosed microsphere formulation can be stored for a long duration at 2-8° C. When the microsphere preparation is used, no solvent is additionally added, and the microsphere formulation is directly mixed with a sample to be detected. On the premise that the concentration of an original system is not changed, the template content in the system can be remarkably improved, so that the detection sensitivity is improved. By using the microsphere formulation in recombinase polymerase amplification (RPA), recombinase-aided amplification (RAA), or a double or multiple detection formed by combining a second reaction on the basis of RPA or RAA, the sensitivity can be remarkably improved, and meanwhile, the amplification specificity is ensured.