Nucleophilic oxime reactivators of organophosphate (OP) inhibited human acetylcholinesterase (hAChE) are reactivating antidotes against OP intoxication from nerve agent (e.g., VX, sarin) or OP pesticide (e.g., paraoxon) exposure. Within the past decade it has become increasingly clear that for effective and complete recovery from OP intoxication antidotal action is needed in both peripheral and central nervous system tissues. We are using structure-based design to create uncharged bis-oxime reactivators that possess superior in vitro properties for reactivation of various OP-conjugates of hAChE compared to the known charged pyridinium aldoxime-based therapies. Our strategies involve studying hAChE from the atomic to the molecular level by employing a combination of experimental and theoretical techniques, including X-ray and neutron crystallography, neutron vibrational spectroscopy and molecular simulations. Starting with RS-170B, MMB4 (both charged) and RS194B (uncharged) oximes we have obtained a clear view of their binding to native and VX- or paraoxon-inhibited hAChE. Neutron vibrational spectra in the 5-50 cm^-1 frequency regime indicated softening of the vibrational dynamics in the paraoxon-bound hAChE, pointing to significant softening of picosecond vibrational motions of the acyl pocket loop that must accommodate one of the pesticide’s ethoxy groups. We have used our crystal structures to design and test uncharged bis-oximes, demonstrating in vitro that several of them exceeded efficacy of RS194B in reactivation of sarin, cyclosarin, VX and paraoxon inhibited hAChE. To further the reactivator design, we are working on obtaining neutron structures of hAChE which will provide direct experimental observation of hydrogen atoms to determine protonation states of hAChE residues and bound reactivators. Our designed uncharged bis-oxime antidotes are a conceptually novel, effective scaffold of nucleophilic reactivators and a promising new resource for creation of adjustable, accelerated centrally active antidotes against OP intoxication.