(12d) Peptoid-Based Coatings for Microarray Sandwich ELISA

Microarray sandwich ELISA is a protein detection technique capable of simultaneously screening 10 ? 50 proteins within a single sample, ideal for the diagnosis of cancers, where multiple protein biomarkers must be tested and small sample volumes are available. While the theoretical detection limit for microarrays is very low, researchers have not yet been able to attain such low sensitivities in practice. The main reason that ELISA microarrays have not yet been able to match theoretical predictions is that current experimental protocols are a combination of DNA microarray and 96-well plate ELISA protocols, and have not yet been optimized for microarray sandwich ELISA. Our research focuses on creating peptoid-based coatings for microarray sandwich ELISA that allow the functional attachment of antibodies. Desirable properties for the coating include: (i) a large surface area for antibody attachment, (ii) retention of antibody structure and function when immobilized, (iii) simple and inexpensive to produce with good slide to slide reproducibility, (iv) low non-specific protein binding, and (v) the ability to be stored for long periods of time. Peptoid 12-mers designed to have these properties have been synthesiszed and purified. Secondary structure is determined by CD and the formation of microstructures is characterized by SEM. Finally, the coatings are compared using microarray ELISA studies, specifically assessing spot size and morphology, slide noise, spot background, lower limit of detection, and reproducibility.