Analyte Proximity Detection Through Barcode Ligation
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Solution Overview
Problem
Current methods for analyzing the proximity of analytes within a sample are limited in their ability to accurately detect and quantify the proximity of specific analytes, particularly in complex biological samples such as tissue samples.
Innovation Solution
The use of probes with specific binding sites and barcodes, followed by ligation and amplification to generate amplicons, allows for the detection of analyte proximity by identifying the complement of the barcode, enabling precise determination of the proximity of analytes like nucleic acids and polypeptides in samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods are used for analyzing analyte proximity, then the analysis can be performed on tissue samples, but the measurement precision and resolution of analyte proximity detection are limited
Solution Approach 1:
The detection method is segmented into distinct functional components: probes with specific binding sites for different analytes, barcode sequences for identification, ligation steps for proximity marking, and amplification steps for signal generation. This segmentation allows each component to be optimized independently while achieving high overall measurement precision
Solution Approach 2:
Barcodes serve as intermediary elements that bridge the gap between analyte proximity events and detectable signals. The barcodes are ligated to probes only when analytes are in proximity, then amplified and detected to provide precise proximity information without direct interaction between the analytes themselves
2Manufacturing precision
If probes with barcodes and ligation amplification are used, then high-resolution analyte proximity analysis is achieved, but the procedure complexity and processing time increase
Solution Approach 1:
Probes are pre-designed with specific binding sites and barcode sequences before the detection experiment. This preliminary preparation allows the actual detection procedure to proceed more efficiently, as the probes are ready to immediately bind to target analytes and perform ligation amplification without additional setup time
Solution Approach 2:
The ligation and amplification steps are designed to occur in continuous sequence without interruption. Once probes bind to proximal analytes, ligation immediately marks the proximity event, followed by continuous amplification to generate detectable signals, maximizing the efficiency of each time interval in the procedure
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 provides a high-resolution analysis of analyte proximity, suitable for fresh-frozen and formalin-fixed paraffin-embedded tissue samples, enhancing the detection of nucleic acid modifications and protein interactions.
Implementation Method 1
a first probe and a second probe, wherein the first probe comprises: (i) a first binding site configured to couple to a first analyte at a first portion of the first analyte; (ii) a second binding site configured to couple to the first analyte at a second portion of the first analyte
Implementation Method 2
c) ligating the first end and the second end to form a circular oligonucleotide
Implementation Method 3
d) amplifying the circular oligonucleotide to generate an amplicon, wherein the amplicon comprises a complement of the barcode
Data Source
AI summary
Provided herein are methods and systems for identifying a proximity of analytes in a sample. The method may comprise contacting one or more analytes with one or more probes. The proximity of the one or more analytes to each other may cause a ligation event between the one or more probes. An amplification reaction may be performed comprising one or more copies of a barcode or derivative thereof. The one or more copies of the barcode or derivative thereof may be detected to identify a proximity between the one or more analytes.