Multiplex Proximity Probes With Shared Hybridization Background Control

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Solution Overview

Problem

Current multiplex proximity assays suffer from high background signal due to random interactions of unbound proximity probes, necessitating a separate negative control to determine true positive signals, which complicates the assay process and introduces variability.

Innovation Solution

Implementing proximity probe pairs that share hybridization sites to form background signal uniformly, allowing true and false positive signals to be distinguished based on paired barcode sequences, eliminating the need for a separate negative control and reducing assay complexity.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If a separate negative control is used to determine background signal levels, then measurement precision is improved, but device complexity and time consumption increase

Engineering Contradiction:

Improvebackground signal determination accuracyVSAvoidassay process complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The invention merges the background control function into the main assay by having all probe pairs share common hybridisation sites. This allows background signal to be generated uniformly across all probes simultaneously, eliminating the need for a separate negative control reaction while maintaining accurate background determination.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The hybridisation sites are designed to be universal across all probe pairs, serving dual purposes: enabling specific analyte detection while simultaneously generating uniform background signal. This multi-functionality integrates background control into the core assay mechanism rather than requiring a separate control system.

Inventive Principle:

Principle #6Universality (Multi-functionality)

2Ease of operation

If probe pairs share hybridisation sites to generate uniform background signal, then assay simplicity is improved, but false positive detection difficulty increases

Engineering Contradiction:

Improveassay process simplicityVSAvoidfalse positive distinction difficulty

Core Design Contradiction:

Ease of operationVSDifficulty of detecting and measuring

Solution Approach 1:

While hybridisation sites are shared universally, the barcode sequences associated with each probe pair remain unique and locally specific. This local differentiation in barcode identification allows clear distinction between true positives (matching barcode pairs) and false positives (non-matching pairs) even though all probes share the same hybridisation mechanism.

Inventive Principle:

Principle #3Local quality

3Measurement precision

If separate negative control is performed, then background signal levels can be determined accurately, but productivity decreases due to additional assay steps

Engineering Contradiction:

Improvebackground signal determinationVSAvoidassay throughput

Core Design Contradiction:

Measurement precisionVSProductivity

Solution Approach 1:

The background control function is merged into the single main assay reaction by using shared hybridisation sites across all probe pairs. This integration eliminates the need for separate control reactions, thereby maintaining accurate background determination while improving assay throughput and productivity.

Inventive Principle:

Principle #5Merging (Combining)

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 provides a more straightforward and accurate method to control false positives by determining background signal levels, enhancing the assay's accuracy and simplifying the process.

Implementation Method 1

nucleic acid moieties linked to the analyte-binding domains of a probe pair hybridise to one another when the probes are in close proximity

Methodology Applied

Scientific EffectHybridisation:

Data Source

PatentUS12534751B1Products for protein analyte detection assays using antibodies

Publication Date: 2026.01.27 OLINK PROTEOMICS AB

AI summary

Products for detecting a plurality of protein analytes comprise a plurality of proximity probe pairs comprising first and second proximity probes having an antibody or antibody fragment specific for the same protein analyte and a nucleic acid domain, which probes can simultaneously bind to the analyte. Each pair is specific for a different analyte. Each nucleic acid domain comprises an ID sequence and at least a first hybridisation sequence. In each probe pair, ID sequences correspond to a particular analyte, and the probes comprise paired hybridisation sequences. For each probe pair, a splint oligonucleotide comprises hybridisation sequences complementary to each of the paired hybridisation sequences. When probes bind to their protein analyte, the respective paired hybridisation sequences can hybridise to the splint oligonucleotide. At least one pair of hybridisation sequences is shared by at least two pairs of proximity probes. A plurality of sample index oligonucleotides is also included.