Nucleic acid linked immune-sandwich assay (NULISA)
Inventors
Luo, Yuling • Feng, Wei • Grzybowski, Adrian • YIN, Yiyuan • Chen, Shiping
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-12704508-B2
Publication Date
2026-08-11
Expiration Date
Abstract
Disclosed herein are highly sensitive immunoassays that utilize a capture/release mechanism to reduce non-specific binding and achieve detection with attomolar-level sensitivity. Kits that can be used for carrying out these highly sensitive immunoassays are also disclosed herein.
Core Innovation
The disclosed assay detects one or multiple analytes using paired binding moieties that bind an analyte to form an immunocomplex. Paired first and second binding moieties each comprise a binder, a target label, and a presenting group. Proximity between first and second target labels on the binding moieties supports generation of a reporter that is detected to identify the target analyte.
The assay architecture captures the immunocomplex on a first solid surface, followed by release and recapture on a second solid surface via presenting and receiving groups. Target and sample identity barcodes facilitate multiplex detection, sample indexing, pooling, and digital quantification, while paired binder identifiers reject cross-reactive signals. Reporter readout is carried out using qPCR, digital PCR, or next-generation sequencing, supported by nucleic acid internal references for normalization.
Claims Coverage
The provided claims identify one independent claim directed to an assay method for detecting one or multiple analytes in a sample, comprising four inventive features.
Paired binding moieties and immunocomplex formation
First and second binding moieties comprise binders that bind an analyte to form an immunocomplex, with each binding moiety comprising a binder, a target label, and a presenting group.
Immunocomplex capture and transfer between solid surfaces
The immunocomplex is captured on a first solid surface, released, and captured on a second solid surface via presenting groups and receiving groups coupled to the surfaces. [procedural detail omitted for safety]
Proximity-based reporter generation and analyte detection
A reporter is generated from the immunocomplex based on proximity between the first and second target labels, and detection of the reporter detects the analyte.
Multiplexed target identification and parallel detection
The claims specify analyte-specific identity barcodes, nucleic acid tags paired with nucleic acid capture probes, and parallel detection of at least twenty different analytes in the same sample.
The independent claim covers paired analyte-binding moieties, capture and transfer of the immunocomplex between solid surfaces, and proximity-based reporter generation, with dependent claims adding identity barcodes, capture probes, and multiplex parallel detection.
Stated Advantages
The assay is described as having low-background and high-plex capabilities, with repeated capture and release intended to reduce nonspecific background.
Examples report reduced background and improved assay sensitivity when immunocomplexes are released before reporter generation.
The disclosure states attomolar limits of detection, including attomolar detection of EGFR.
Target and sample identifiers support multiplexed analyte and sample identification, pooling, and digital quantification.
Paired binder identifiers can help reject cross-reactive signals.
Partial capture proportionally reduces signals from abundant analytes to manage detection bandwidth.
Examples report variable coefficient-of-variation improvements from internal-reference normalization in a four-analyte assay.
Documented Applications
Detection of one or multiple different analytes in a sample, including parallel detection of at least twenty analytes in the same sample.
Multiplex detection and sample pooling using target and sample identity encoding, with reporter readout by PCR, qPCR, digital PCR, or next-generation sequencing.
Detection of specific analytes and panels including EGFR, IL-4, IL-6, IL-10, TNFα, and P24, including comparison of P24 quantification by qPCR and sequencing.
Interested in licensing this patent?