Chromato-kinetic Fingerprinting Enables Multiomic Digital Counting of Single Disease Biomarker Molecules

Overview

Journal bioRxiv

Date 2025 Feb 20

PMID 39975368

Authors

Pavel Banerjee

Sujay Ray

Liuhan Dai

Erin Sandford

Tanmay Chatterjee

Shankar Mandal

Javed Siddiqui

Muneesh Tewari

Nils G Walter

Affiliations

Soon will be listed here.

Abstract

Early and personalized intervention in complex diseases requires robust molecular diagnostics, yet the simultaneous detection of diverse biomarkers-microRNAs (miRNAs), mutant DNAs, and proteins-remains challenging due to low abundance and preprocessing incompatibilities. We present Biomarker Single-molecule Chromato-kinetic multi-Omics Profiling and Enumeration (Bio-SCOPE), a next-generation, triple-modality, multiplexed detection platform that integrates both chromatic and kinetic fingerprinting for molecular profiling through digital encoding. Bio-SCOPE achieves femtomolar sensitivity, single-base mismatch specificity, and minimal matrix interference, enabling precise, parallel quantification of up to six biomarkers in a single sample with single-molecule resolution. We demonstrate its versatility in accurately detecting low-abundance miRNA signatures from human tissues, identifying upregulated miRNAs in the plasma of prostate cancer patients, and measuring elevated interleukin-6 (IL-6) and hsa-miR-21 levels in cytokine release syndrome patients. By seamlessly integrating multiomic biomarker panels on a unified, high-precision platform, Bio-SCOPE provides a transformative tool for molecular diagnostics and precision medicine.

References

Aitken C, Marshall R, Puglisi J . An oxygen scavenging system for improvement of dye stability in single-molecule fluorescence experiments. Biophys J. 2007; 94(5):1826-35. PMC: 2242739. DOI: 10.1529/biophysj.107.117689. View

Lu C, Tej S, Luo S, Haudenschild C, Meyers B, Green P . Elucidation of the small RNA component of the transcriptome. Science. 2005; 309(5740):1567-9. DOI: 10.1126/science.1114112. View

Pritchard C, Cheng H, Tewari M . MicroRNA profiling: approaches and considerations. Nat Rev Genet. 2012; 13(5):358-69. PMC: 4517822. DOI: 10.1038/nrg3198. View

Johnson-Buck A, Su X, Giraldez M, Zhao M, Tewari M, Walter N . Kinetic fingerprinting to identify and count single nucleic acids. Nat Biotechnol. 2015; 33(7):730-2. PMC: 4559481. DOI: 10.1038/nbt.3246. View

Geekiyanage H, Rayatpisheh S, Wohlschlegel J, Brown Jr R, Ambros V . Extracellular microRNAs in human circulation are associated with miRISC complexes that are accessible to anti-AGO2 antibody and can bind target mimic oligonucleotides. Proc Natl Acad Sci U S A. 2020; 117(39):24213-24223. PMC: 7533700. DOI: 10.1073/pnas.2008323117. View

Liu L, Li T, Zhang S, Song P, Guo B, Zhao Y . Simultaneous Quantification of Multiple Cancer Biomarkers in Blood Samples through DNA-Assisted Nanopore Sensing. Angew Chem Int Ed Engl. 2018; 57(37):11882-11887. DOI: 10.1002/anie.201803324. View

Schnitzbauer J, Strauss M, Schlichthaerle T, Schueder F, Jungmann R . Super-resolution microscopy with DNA-PAINT. Nat Protoc. 2017; 12(6):1198-1228. DOI: 10.1038/nprot.2017.024. View

Shah S, Dubey A, Reif J . Programming Temporal DNA Barcodes for Single-Molecule Fingerprinting. Nano Lett. 2019; 19(4):2668-2673. DOI: 10.1021/acs.nanolett.9b00590. View

Chakraborty S, Sharma G, Karmakar S, Banerjee S . Multi-OMICS approaches in cancer biology: New era in cancer therapy. Biochim Biophys Acta Mol Basis Dis. 2024; 1870(5):167120. DOI: 10.1016/j.bbadis.2024.167120. View

10.

Lin C, Jungmann R, Leifer A, Li C, Levner D, Church G . Submicrometre geometrically encoded fluorescent barcodes self-assembled from DNA. Nat Chem. 2012; 4(10):832-9. PMC: 3523708. DOI: 10.1038/nchem.1451. View

11.

Hye A, Lynham S, Thambisetty M, Causevic M, Campbell J, Byers H . Proteome-based plasma biomarkers for Alzheimer's disease. Brain. 2006; 129(Pt 11):3042-50. DOI: 10.1093/brain/awl279. View

12.

Wade O, Woehrstein J, Nickels P, Strauss S, Stehr F, Stein J . 124-Color Super-resolution Imaging by Engineering DNA-PAINT Blinking Kinetics. Nano Lett. 2019; 19(4):2641-2646. PMC: 6463241. DOI: 10.1021/acs.nanolett.9b00508. View

13.

Mestdagh P, Hartmann N, Baeriswyl L, Andreasen D, Bernard N, Chen C . Evaluation of quantitative miRNA expression platforms in the microRNA quality control (miRQC) study. Nat Methods. 2014; 11(8):809-15. DOI: 10.1038/nmeth.3014. View

14.

Lopez-Santillan M, Larrabeiti-Etxebarria A, Arzuaga-Mendez J, Lopez-Lopez E, Garcia-Orad A . Circulating miRNAs as biomarkers in diffuse large B-cell lymphoma: a systematic review. Oncotarget. 2018; 9(32):22850-22861. PMC: 5978269. DOI: 10.18632/oncotarget.25230. View

15.

Alles J, Fehlmann T, Fischer U, Backes C, Galata V, Minet M . An estimate of the total number of true human miRNAs. Nucleic Acids Res. 2019; 47(7):3353-3364. PMC: 6468295. DOI: 10.1093/nar/gkz097. View

16.

Jin Z, Geissler D, Qiu X, Wegner K, Hildebrandt N . A Rapid, Amplification-Free, and Sensitive Diagnostic Assay for Single-Step Multiplexed Fluorescence Detection of MicroRNA. Angew Chem Int Ed Engl. 2015; 54(34):10024-9. DOI: 10.1002/anie.201504887. View

17.

Cohen J, Li L, Wang Y, Thoburn C, Afsari B, Danilova L . Detection and localization of surgically resectable cancers with a multi-analyte blood test. Science. 2018; 359(6378):926-930. PMC: 6080308. DOI: 10.1126/science.aar3247. View

18.

Arroyo J, Chevillet J, Kroh E, Ruf I, Pritchard C, Gibson D . Argonaute2 complexes carry a population of circulating microRNAs independent of vesicles in human plasma. Proc Natl Acad Sci U S A. 2011; 108(12):5003-8. PMC: 3064324. DOI: 10.1073/pnas.1019055108. View

19.

Koch C, Reilly-ODonnell B, Gutierrez R, Lucarelli C, Ng F, Gorelik J . Nanopore sequencing of DNA-barcoded probes for highly multiplexed detection of microRNA, proteins and small biomarkers. Nat Nanotechnol. 2023; 18(12):1483-1491. PMC: 10716039. DOI: 10.1038/s41565-023-01479-z. View

20.

Zhang D, Chen S, Yin P . Optimizing the specificity of nucleic acid hybridization. Nat Chem. 2012; 4(3):208-14. PMC: 4238961. DOI: 10.1038/nchem.1246. View