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bioRxiv · 10.64898/2026.01.23.701152

DISCO-LAMP: A Novel discontinuous LAMP assay for isothermal antigen detection

Thomas, B. M.·Simeon, R. A.·Yan, K. L.·Chonira, V.·Chen, W. T.·Webb, E. L.·Mutchler, C.·Fernandez, A.·Han, J.·Chen, Z.
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Abstract

Proximity ligation assay (PLA), in which the ligation of two DNA probes is greatly accelerated by the associating target molecules, has emerged as a highly sensitive technique for protein detection. The detection of the ligated DNA typically relies on PCR, which requires temperature cycling. In this study, we report on a novel discontinuous (DISCO)-LAMP assay that enables the wash-free detection of PLA products via loop-mediated isothermal amplification (LAMP). Due to the exponential amplification nature of LAMP, a careful balance between efficient amplification of the ligated full-length DNA and minimal background amplification from the individual constituent probes is essential but often challenging to achieve. After extensive template/primer design and assay optimization, DISCO-LAMP assay achieved a detection limit of 1 fM for the ligated DNA probe while maintaining undetectable background amplification at 1 nM of each individual probe. DISCO-LAMP detected Shiga toxin 2 (Stx2) with a limit of detection (LoD) of 100 fM when functionalized with Stx2-binders, as well as both Wuhan-1 and Omicron spike protein when functionalized with DS16, a newly engineered DARPin targeting a conserved epitope on the SARS-CoV-2 Spike protein. We believe DISCO-LAMP represents a versatile and efficient LAMP-based PLA technology that is readily adaptable for sensing diverse targets.

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Thomas, B. M., Simeon, R. A., Yan, K. L., Chonira, V., Chen, W. T., Webb, E. L., Mutchler, C., Fernandez, A., Han, J., Chen, Z.. 2026-01-28. DISCO-LAMP: A Novel discontinuous LAMP assay for isothermal antigen detection. https://doi.org/10.64898/2026.01.23.701152

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