A study is presented on the binding kinetics and mechanism of the adsorption of dsDNA on citrate-capped gold nanoparticles (AuNPs). Methods include fluorescence titration, isothermal calorimetry (ITC) titration, dynamic light scattering and gel electrophoresis. It is found that the fluorescence of probe DNA (labeled with Rhodamine Green and measured at excitation/emission peaks of 498/531 nm) is quenched by addition of AuNPs. The Stern-Volmer quenching constant (Ksv) is 1.67 × 109 L·mol at 308 K and drops with increasing temperature. The quenching mechanism is mainly static. The results of both fluorescence titrations and ITC show negative values for ΔH and ΔS values. This shows ion-induced dipole-dipole interaction to be the main attractive forces between dsDNA and AuNPs, while electrostatic interactions result in repulsion. The repulsive forces lead to a lower affinity between dsDNA and AuNPs (compared to single-strand DNA). It is also found that dsDNA can prevent the aggregation of AuNPs which is accompanied by a color change from red into blue. The visual detection limit with bare eyes for dsDNA is 36 pM. Based on these findings, a colorimetric method was developed to detect the proto-oncogene of serine/threonine-protein kinase B-Raf V600E point mutation in HT29, Ec109, A549, Huh-7 and SW480 cell lines. Graphical abstract Schematic of the salt-induced aggregation of uncapped gold nanoparticles (AuNPs) which leads to a color change from red to blue. If the AuNPs are coated with dsDNA, aggregation is suppressed.