Total internal reflection (TIR)/fluorescence recovery after photobleaching (FRAP), which has been used to study adsorption and surface diffusion of proteins, was modified and applied to study DNA oligonucleotides at liquid/solid interfaces. Conventional TIR/spot FRAP and TIR/pattern FRAP techniques use a photomultiplier tube (PMT) to reveal the adsorption dynamics and surface diffusion rates of biomolecules, respectively. However, they do not provide spatial information on these interfacial processes. In this work, a cooled charge-coupled device camera is substituted for the PMT normally used. Studies of adsorption and surface diffusion of the well-characterized protein bovine serum albumin (BSA) validated the system's operation. Then, the desorption rate constant for a fluorescently tagged 21-mer DNA oligonucleotide (MW 7140 Da) was determined by spot FRAP. The desorption rate constants for strongly and weakly adsorbed oligonucleotides from (3-aminopropyl)triethoxy silane (APTES) glass were determined to...