Single Molecule Loading via Binding Site Mediators
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Solution Overview
Problem
Current single molecule analysis techniques face challenges in loading reaction/observation regions with multiple analyte molecules, leading to inefficient data collection and high reagent consumption due to random distribution methods, which result in low loading densities and high data discard rates.
Innovation Solution
The development of methods and compositions that control the loading of analyte molecules into size-confined regions by creating single binding sites, using sizing moieties, and controlling the size and charge of analytes, allowing for non-random distribution and higher loading densities in arrays like Zero Mode Waveguides (ZMWs).
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If random distribution methods are used to load analyte molecules into reaction/observation regions, then the loading process is simple, but the loading density is low and data discard rate is high
Solution Approach 1:
The patent introduces binding sites as intermediary structures that mediate between the analyte molecules and the reaction/observation regions. These binding sites are delivered via particles that facilitate controlled loading, acting as intermediaries that enable non-random distribution while maintaining process simplicity.
Solution Approach 2:
The patent applies preliminary action by pre-delivering binding sites to the array before adding analyte molecules. This preliminary placement of binding sites enables subsequent controlled loading of analytes, achieving high loading density without requiring complex real-time control during the loading process.
2Ease of manufacture
If random distribution methods are used to load analyte molecules, then reagent consumption is high, but the loading method is simple
Solution Approach 1:
Binding sites serve as intermediaries that enable precise targeting of analyte molecules to specific reaction/observation regions. This intermediary mechanism ensures that reagents are delivered only where needed, minimizing waste and reducing overall reagent consumption compared to random distribution methods.
Solution Approach 2:
The patent changes the parameter of distribution randomness by introducing controlled positioning mechanisms. By transforming the loading process from random to controlled distribution, the system achieves higher efficiency and reduced reagent consumption while maintaining operational simplicity.
3Productivity
If multiple analyte molecules are loaded into a single reaction/observation region, then data collection efficiency decreases, but the region utilization increases
Solution Approach 1:
The patent applies segmentation by dividing the array into distinct reaction/observation regions, each with its own binding site. This segmentation ensures that analyte molecules are distributed to separate regions rather than accumulating in the same region, thereby maintaining data collection efficiency while achieving proper region utilization.
Solution Approach 2:
Binding sites act as intermediaries that ensure one-to-one correspondence between reaction/observation regions and analyte molecules. This intermediary mechanism prevents multiple analytes from occupying the same region, maintaining data collection efficiency while achieving optimal region utilization through controlled distribution.
Data Source
AI summary
Methods, compositions and arrays for non-random loading of single analyte molecules into array structures are provided. For example, methods are presented for providing a surface comprising the plurality of array regions by exposing the surface to a solution comprising polymerase enzymes where each polymerase enzyme is bound to a binding structure having several functional moieties. The functional moieties of the binding structure react with the binding elements on the array regions such that the functional moieties on the binding structure react with other available binding sites in an array region, preventing other polymerase-binding structures from loading, and resulting in a single polymerase molecule bound to each of these regions.