Surface immobilization of DNA aptamers for biosensing and protein interaction analysis

To utilize aptamers as molecular recognition agents in biosensors and biodiagnostics, it is important to develop strategies for reliable immobilization of aptamers so that they retain their biophysical characteristics and binding abilities. Here we report on quartz crystal microbalance (QCM) measurements and atomic force microscope (AFM)-based force spectroscopy studies to evaluate aptasensors fabricated by different modification strategies. Gold surfaces were modified with mixed self assembled monolayers (SAMs) of aptamer and oligoethylene glycol (OEG) thiols (HS-C11-(EG)nOH, n = 3 or 6) to impart resistance to nonspecific protein adsorption. By affinity analysis, we show that short OEG thiols have less impact on aptamer accessibility than longer chain thiols. Backfilling with OEG as a step subsequent to aptamer immobilization provides greater surface coverage than using aptamer and OEG thiol to form a mixed SAM in one-step. Immunoglobulin E and vascular endothelial growth factor (VEGF) were studied as target proteins in these experiments. Binding forces obtained by these strategies are similar, demonstrating that the biophysical properties of the aptamer on the sensors are independent from the immobilization strategy. The results present mixed SAMs with aptamers and co-adsorbents as a versatile strategy for aptamer sensor platforms including ultrasensitive biosensor design.