Enzymatically Amplified Surface Plasmon Resonance Imaging Detection of DNA by Exonuclease III Digestion of DNA Microarrays

• Published in: Analytical chemistry (Washington) 2005-08, Vol.77 (16), p.5096-5100
• Main Authors: Lee, Hye Jin, Li, Yuan, Wark, Alastair W, Corn, Robert M
• Format: Article
• Language: English
• Subjects: Analytical biochemistry: general aspects, technics, instrumentation; Analytical chemistry; Analytical, structural and metabolic biochemistry; Biological and medical sciences; Chemistry; Deoxyribonucleic acid; DNA; DNA - analysis; DNA - metabolism; Enzymes; Exact sciences and technology; Exodeoxyribonucleases - metabolism; Fundamental and applied biological sciences. Psychology; General, instrumentation; Hybridization; Oligonucleotide Array Sequence Analysis - methods; Scientific imaging; Surface Plasmon Resonance - methods
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac050815w

This paper describes a novel approach utilizing the enzyme exonuclease III in conjunction with 3‘-terminated DNA microarrays for the amplified detection of single-stranded DNA (ssDNA) with surface plasmon resonance (SPR) imaging. When ExoIII and target DNA are simultaneously introduced to a 3‘-terminated ssDNA microarray, hybridization adsorption of the target ssDNA leads to the direction-dependent ExoIII hydrolysis of probe ssDNA strands and the release of the intact target ssDNA back into the solution. Readsorption of the target ssDNA to another probe creates a repeated hydrolysis process that results over time in a significant negative change in SPR imaging signal. Experiments are presented that demonstrate the direction-dependent surface enzyme reaction of ExoIII with double-stranded DNA as well as this new enzymatically amplified SPR imaging process with a 16-mer target ssDNA detection limit of 10−100 pM. This is a 102−103 improvement on previously reported measurements of SPR imaging detection of ssDNA based solely on hybridization adsorption without enzymatic amplification.