title: Real-time ligation chain reaction for DNA quantification and identification on the FO-SPR.
abstract: Different assays have been developed in the past years to meet point-of-care diagnostic tests requirements for fast and sensitive quantification and identification of targets. In this paper, we developed the ligation chain reaction (LCR) assay on the Fiber Optic Surface Plasmon Resonance (FO-SPR) platform, which enabled simultaneous quantification and cycle-to-cycle identification of DNA during amplification. The newly developed assay incorporated FO-SPR DNA melting assay, previously developed by our group. This required establishment of several assay parameters, including buffer ionic strength and thermal ramping speed as these parameters both influence the ligation enzyme performance and the hybridization yield of the gold nanoparticles (Au NPs) on the FO-SPR sensor. Quantification and identification of DNA targets was achieved over a wide concentration range with a calibration curve spanning 7 orders of magnitude and LOD of 13.75 fM. Moreover, the FO-SPR LCR assay could discriminate single nucleotide polymorphism (SNPs) without any post reaction analysis, featuring thus all the essential requirements of POC tests.
authors.affiliation: BIOSYST-MeBioS, KU Leuven - University of Leuven, Willem de Croylaan 42, B-3001 Leuven, Belgium. Electronic address: knez.karel@gmail.com.
authors.forename: Karel
authors.initials: K
authors.lastname: Knez
authors.affiliation: BIOSYST-MeBioS, KU Leuven - University of Leuven, Willem de Croylaan 42, B-3001 Leuven, Belgium. Electronic address: Dragana.Spasic@biw.kuleuven.be.
authors.forename: Dragana
authors.initials: D
authors.lastname: Spasic
authors.affiliation: BIOSYST-MeBioS, KU Leuven - University of Leuven, Willem de Croylaan 42, B-3001 Leuven, Belgium. Electronic address: Filip.Delport@biw.kuleuven.be.
authors.forename: Filip
authors.initials: F
authors.lastname: Delport
authors.affiliation: BIOSYST-MeBioS, KU Leuven - University of Leuven, Willem de Croylaan 42, B-3001 Leuven, Belgium. Electronic address: jeroen.lammertyn@biw.kuleuven.be.
authors.forename: Jeroen
authors.initials: J
authors.lastname: Lammertyn
mesh_terms: D015374:Biosensing Techniques; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D005336:Fiber Optic Technology; D006046:Gold / Q000737:chemistry; D006801:Humans; D022521:Ligase Chain Reaction; D053768:Metal Nanoparticles / Q000737:chemistry; D021141:Nucleic Acid Amplification Techniques; D009691:Nucleic Acid Denaturation / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D020349:Surface Plasmon Resonance
chemical_list: D006046:Gold; D004247:DNA
keywords: FO-SPR melting assay; Fiber Optic Surface Plasmon Resonance (FO-SPR); Gold nanoparticles (Au NPs); Ligation chain reaction (LCR)