Label-free detection of DNA single-base mismatches using a simple reflectance-based optical technique.
Rapid and quantitative detection of the binding of nucleic acids to surface-immobilized probes remains a challenge in many biomedical applications. We investigated the hybridization of a set of fully complementary and defected 12-base long DNA oligomers by using the Reflective Phantom Interface (RPI), a recently developed multiplexed label-free detection technique. Based on the simple measurement of reflected light intensity, this technology enables to quantify the hybridization directly as it occurs on the surface with a sensitivity of 10 pg mm(-2). We found a strong effect of single-base mismatches and of their location on hybridization kinetics and equilibrium binding. In line with previous studies, we found that DNA-DNA binding is weaker on a surface than in the bulk. Our data indicate that this effect is a consequence of weak nonspecific binding of the probes to the surface.
Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, via Fratelli Cervi 93, 20090 Segrate, Milano, Italy. giuliano.zanchetta@unimi.it.
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D020137:Base Pair Mismatch; D015374:Biosensing Techniques; D004247:DNA / Q000737:chemistry*; D015342:DNA Probes / Q000737:chemistry; D007700:Kinetics; D008027:Light; D009693:Nucleic Acid Hybridization
D015342:DNA Probes; D004247:DNA