title: Rapid and sensitive detection of multiple microRNAs in cell lysate by low-fouling surface plasmon resonance biosensor.
abstract: We report an ultra-low fouling surface plasmon resonance imaging (SPRi) biosensor for the rapid simultaneous detection of multiple miRNAs in erythrocyte lysate (EL) at subpicomolar levels without need of RNA extraction. The SPRi chips were coated with ultra-low fouling functionalizable poly(carboxybetaine acrylamide) (pCBAA) brushes having optimized thicknesses and directly functionalized with amino-modified oligonucleotide probes. We have characterized the effect of the brush thickness on the probe loading capacity: a loading capacity of ~9.8×10(12) probes/cm(2) was achieved for pCBAA having a thickness of ~40 nm. The probe-functionalized sensor also exhibited a high resistance to fouling from ~90% EL samples (<2 ng/cm(2)). A two-step detection assay was employed for multiplexed miRNA detection in EL. Specifically, the assay consisted of (i) a sandwich-type hybridization of the probe-functionalized pCBAA with target miRNA in EL (bound to biotinylated oligonucleotides) and (ii) the capture of streptavidin-functionalized gold nanoparticles to the aforementioned biotinylated probes. We have demonstrated that this approach enables the detection of miRNAs in EL at concentrations as low as 0.5 pM. Finally, we have confirmed the detection of four endogenous miRNAs representing a set of potential miRNA biomarkers of myelodysplastic syndrome (MDS) in clinical EL samples (miR-16, miR-181, miR-34a, and miR-125b). The results revealed significantly higher levels of miR-16 in all the clinical EL samples compared to the other measured miRNAs.
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Hana
authors.initials: H
authors.lastname: Vaisocherová
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Hana
authors.initials: H
authors.lastname: Šípová
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Ivana
authors.initials: I
authors.lastname: Víšová
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Markéta
authors.initials: M
authors.lastname: Bocková
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Tomáš
authors.initials: T
authors.lastname: Špringer
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Maria Laura
authors.initials: ML
authors.lastname: Ermini
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic.
authors.forename: Xue
authors.initials: X
authors.lastname: Song
authors.affiliation: Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Zdeněk
authors.initials: Z
authors.lastname: Krejčík
authors.affiliation: Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Leona
authors.initials: L
authors.lastname: Chrastinová
authors.affiliation: Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Ondřej
authors.initials: O
authors.lastname: Pastva
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic; Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Kristýna
authors.initials: K
authors.lastname: Pimková
authors.affiliation: Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Michaela
authors.initials: M
authors.lastname: Dostálová Merkerová
authors.affiliation: Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague, Czech Republic.
authors.forename: Jan E
authors.initials: JE
authors.lastname: Dyr
authors.affiliation: Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská 57, 18251 Prague, Czech Republic. Electronic address: homola@ufe.cz.
authors.forename: Jiří
authors.initials: J
authors.lastname: Homola
mesh_terms: D000178:Acrylamides / Q000737:chemistry*; D015374:Biosensing Techniques / Q000295:instrumentation*; D002458:Cell Fractionation; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis; D045424:Complex Mixtures / Q000032:analysis; D004867:Equipment Design; D019544:Equipment Failure Analysis; D035683:MicroRNAs / Q000032:analysis* / Q000737:chemistry* / Q000235:genetics; D011108:Polymers / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000295:instrumentation*
chemical_list: D000178:Acrylamides; D020099:Coated Materials, Biocompatible; D045424:Complex Mixtures; D035683:MicroRNAs; D011108:Polymers; C533866:poly(carboxybetaine acrylamide)
keywords: DNA array; Erythrocyte lysate; Low-fouling surface chemistry; Polymer brushes; Surface plasmon resonance imaging; microRNA