ADSORPTION AND SURFACE-DIFFUSION OF DNA OLIGONUCLEOTIDES AT LIQUID SOLID INTERFACES/

Total internal reflection (TIR)/fluorescence recovery after photobleac hing (FRAP), whit-h has been used to study adsorption and surface diff usion of proteins, was modified and applied to study DNA oligonucleoti des at liquid/solid interfaces. Conventional TIR/spot FRAP and TIR/pat tern FRAP techniques use a photomultiplier tube (PMT) to reveal the ad sorption dynamics and surface diffusion rates of biomolecules, respect ively. However, they do not provide spatial information on these inter facial processes. in this work, a cooled charge-coupled device camera is substituted for the PMT normally used. Studies of adsorption and su rface diffusion of the well-characterized protein bovine serum albumin (BSA) validated the system's operation. Then, the desorption rate con stant for a fluorescently tagged 21-mer DNA oligonucleotide (MW 7140 D a) was determined by spot FRAP. The desorption rate constants for stro ngly and weakly adsorbed oligonucleotides from (3-aminopropyl)triethox y silane (APTES)glass were determined to be 0.02 and 0.19 s(-1), respe ctively. These are of the same order of magnitude as those for ESA (MW 67 000 Da) on APTES glass. The surface diffusion coefficients of olig onucleotide are approximately the same as those for BSA and are depend ent on the surface concentration of the molecules on APTES-coated glas s. Since the molecules differ by a factor of 10 in molecular weight, t hese results suggest that the shape of a adsorbate molecule and the st rength of adsorbate/substrate interactions play a strong role in inter facial adsorption and diffusion. The substitution of a methyl group in APTES For a hydrogen atom increased the desorption rate constants and surface diffusion coefficients significantly.