Dielectrophoretic nanocolloid assay is a promising technique for sensitive molecular detection and identification, as target molecule hybridization onto the probe-functionalized nanocolloids can change their surface conductance and consequently their dielectrophoretic crossover frequencies. Thus, instead of relying on surface charge density increase after hybridization, as in many capacitive and field effect transistor impedance sensing techniques, the current assay utilizes the much larger surface conductance (and dielectrophoresis crossover frequency) changes to effect sensitive detection. Herein, we present a Poisson-Boltzmann theory for surfaces with finite-size molecular probes that include the surface probe conformation, their contribution to surface charge with a proper delineation of the slip and Stern planes. The theory shows that the most sensitive nanocolloid molecular sensor corresponds to a minimum in the dielectrophoretic crossover frequency with respect to the bulk concentration of the molecular probes (oligonucleotides in our case) during nanocolloid functionalization. This minimum yields the lowest number of functionalized probes that are also fully stretched because of surface probe-probe interaction. Our theory provides the surface-bulk oligonucleotide concentration isotherm and a folding number for the surface oligonucleotide conformation from the crossover frequency, the zeta potential, and the hydrodynamic radius data.