title: Self-assembling protein platform for direct quantification of circulating microRNAs in serum with total internal reflection fluorescence microscopy.
abstract: MicroRNA (miRNA) has recently emerged as a new and important class of cellular regulators. Strong evidences showed that aberrant expression of miRNA is associated with a broad spectrum of human diseases, such as cancer, diabetes, cardiovascular and psychological disorders. However, the short length and low abundance of miRNA place great challenges for conventional techniques in the miRNA quantification and expression profiling. Here, we report a direct, specific and highly sensitive yet simple detection assay for miRNA without sample amplification. A self-assembled protein nanofibril acted as an online pre-concentrating sensor to detect the target miRNA. Locked nucleic acid (LNA) of complimentary sequence was served as the probe to capture the target miRNA analyte. The quantification was achieved by the fluorescence intensity measured with total internal reflection fluorescence microscopy. A detection limit of 1 pM was achieved with trace amount of sample consumption. This assay showed efficient single-base mismatch discrimination. The applicability of quantifying circulating mir-196a in both normal and cancer patient's serums was also demonstrated.
authors.affiliation: Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong, PR China.
authors.forename: See-Lok
authors.initials: SL
authors.lastname: Ho
authors.affiliation: Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong, PR China.
authors.forename: Ho-Man
authors.initials: HM
authors.lastname: Chan
authors.affiliation: Department of Biology, Hong Kong Baptist University, Kowloon Tong, Hong Kong, PR China.
authors.forename: Ricky Ngok-Shun
authors.initials: RN
authors.lastname: Wong
authors.affiliation: Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong, PR China. Electronic address: hwli@hkbu.edu.hk.
authors.forename: Hung-Wing
authors.initials: HW
authors.lastname: Li
mesh_terms: D006801:Humans; D035683:MicroRNAs / Q000097:blood*; D008856:Microscopy, Fluorescence / Q000379:methods*; D049329:Nanostructures / Q000737:chemistry; D012680:Sensitivity and Specificity
chemical_list: D035683:MicroRNAs