Organized cell adherent array (OC2A) for real-time multiplex detection of secreted molecules.
Résumé
The organized cell adherent array (OC2A) is an integrated tool useful for real-time and label-free detection of secreted molecules. The OC2A novel cell chip is composed of two substrates, one allowing localized cell culture and the other one constituting the sensitive layer detection. These two units are assembled thanks to a thickness controlled spacer (250µm) which forms a microfluidic lumen. In practice, the top layer of the chip is a polycarbonate substrate covered by a poly (ethylene oxide)-like (PEO-like) film deposited by plasma polymerization. This PEO-like film is cells repellent but allows the direct immobilization of protein through low volume spotting. Thus, fibronectin spots were deposited on this PEO-like film in order to create cell adhesive areas surrounded by non-adhesive background. On the sensitive layer part, various antibodies were immobilized in an organized manner on a surface plasmon resonance imaging (SPRi) chip useful for the multiplex label-free detection of proteins. The two units are then assembled and the cells loaded in the chip. After one hour of culture, non-adherent cells are removed by washing and the chip is ready to experiment. Evidences were obtained of the real-time detection of Prostate Specific Antigen (PSA) and β-2-microglobulin (B2M) secreted by LNCaP cells, androgen-sensitive human prostate adenocarcinoma cells, following induction by dihydrotestosterone (DHT). Different kinetics of the two secreted proteins were also demonstrated and precisely determined using the OC2A system.