Rivastigmine tartrate (RT) is a noncompetitive carbamate acetylcholinesterase inhibitor that is carbamy-lated, long-acting, and reversible. It's used to alleviate Alzheimer's disease symptoms that range from mild to moderate. Although various cholinesterase inhibitors have been researched for their DNA binding mode, the RT-dsDNA binding mechanism has not been investigated yet. As a result, both experimentally and theoretically, the binding mechanism and reversible interactions of RT with double-stranded fish sperm deoxyribonucleic acid (dsDNA) have been thoroughly investigated. Our overall purpose is to have a better understanding of the potential of cholinesterase inhibitor medications in order to find novel medications that target dsDNA and develop drugs with fewer adverse effects. We applied a variety of spectroscopic methods under physiological situations, including UV-vis and fluorescence, spectroscopy, thermal denaturation, electrochemical and viscosity measurements, as well as molecular docking investi-gations. In addition, the interactions between RT and dsDNA were investigated computationally with the assistance of molecular docking and molecular dynamics simulations. Data from both experimental and theoretical results showed that RT interacts with dsDNA through the groove binding mode.(c) 2022 Published by Elsevier B.V.