Efficient probe immobilization on poly (dimethylsiloxane) for sensitive detection of proteins.
Chemical surface modification methods were investigated to activate surface of poly-(dimethylsiloxane) (PDMS) for probe immobilization in enzyme-linked immunosorbent assay (ELISA). The investigations started from dramatization of PDMS surface with (3-aminopropyl)-triethoxysilane (APTES). Amino groups generated by APTES were either derived into carboxyl group by Succinic acid anhydride (SAA), then captured the protein through the heterobifunctional cross-linker, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), or directly reacted with homobifunctional cross-linker, glutaraldehyde (GA). Rabbit IgG, Goat IgG, Human IgA and Rat IgG were covalently immobilized on PDMS by both methods, and examined through well-organized competitive inhibition ELISA. The results demonstrated that both GA and EDC based methods had good covalent immobilization capability, and the EDC method had higher efficiency than the GA method. The competitive ELISA with probe immobilization through EDC had a detection limit of pg/ml level. The method was proved to be applicable for immobilization of different proteins. The topography of the immobilized protein properties was studied by atomic force microscope (AFM), demonstrating that the immobilization by GA had protein conglomeration, resulting in poorer uniformity and lower immobilization efficiency than EDC method. The possible reason is protein inter-molecule crossing linkage by the homobifunctional group of GA. Due to its simplicity, low cost, and high immobilization efficiency, EDC based immobilization method could provide great potential for making ELISA protein chips based on PDMS.
Yu
Ling
L
School of Chemical and Biomedical Engineering, Nanyang Technological University, Nanyang Avenue, 639798, Singapore.
Li
C M
CM
Zhou
Qin
Q
D000818:Animals; D004129:Dimethylpolysiloxanes / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007118:Immunoassay; D018625:Microscopy, Atomic Force / Q000379:methods; D015335:Molecular Probes
D004129:Dimethylpolysiloxanes; D015335:Molecular Probes