Carbon electrode obtained via pyrolysis of plasma-deposited parylene-C for electrochemical immunoassays.

In this study, parylene-C films from plasma deposition as well as thermal deposition were pyrolyzed to prepare a carbon electrode for application in electrochemical immunoassays. Plasma deposition could prepare parylene-C in a faster deposition rate and more precise control over the thickness in comparison with the conventional thermal deposition. To analyze the influence of the deposition method, the crystal and electronic structures of the pyrolyzed parylene-C films obtained via both deposition methods were compared using Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and Raman spectroscopy. For application as a carbon electrode in immunoassays, the electrochemical properties of the pyrolyzed carbon films from two both deposition methods were analyzed, including the double layer capacitance (2.10 µF cm-2 for plasma deposition and 2.20 µF cm-2 for thermal deposition), the apparent electron transfer rate (approximately 1.1 × 10-3 cm s-1 for both methods), and the electrochemical window (approximately -1.0 ∼ 2.1 V for both methods). Finally, the applicability of the pyrolyzed carbon electrode from parylene-C was demonstrated for the diagnosis of human hepatitis-C using various amperometric methods, such as cyclic voltammetry, chronoamperometry, square-wave voltammetry and differential pulse voltammetry.

Band-type microelectrodes for amperometric immunoassays.

A band-type microelectrode was made using a parylene-N film as a passivation layer. A circular-type, mm-scale electrode with the same diameter as the band-type microelectrode was also made with an electrode area that was 5000 times larger than the band-type microelectrode. By comparing the amperometric signals of 3,5,3',5'-tetramethylbenzidine (TMB) samples at different optical density (OD) values, the band-type microelectrode was determined to be 9 times more sensitive than the circular-type electrode. The properties of the circular-type and the band-type electrodes (e.g., the shape of their cyclic voltammograms, the type of diffusion layer used, and the diffusion layer thickness per unit electrode area) were characterized according to their electrode area using the COMSOL Multiphysics software. From these simulations, the band-type electrode was estimated to have the conventional microelectrode properties, even when the electrode area was 100 times larger than a conventional circular-type electrode. These results show that both the geometry and the area of an electrode can influence the properties of the electrode. Finally, amperometric analysis based on a band-type electrode was applied to commercial ELISA kits to analyze human hepatitis B surface antigen (hHBsAg) and human immunodeficiency virus (HIV) antibodies.

Development of SPR biosensor for the detection of human hepatitis B virus using plasma-treated parylene-N film.

A plasma-treated parylene-N film was presented for the immobilization of proteins through physical adsorption. The changes in surface properties of the parylene-N film after plasma-treatment were analyzed using contact angle microscopy and AFM. To demonstrate the high protein-immobilization efficiency of the plasma-treated parylene-N film, the immobilization efficiencies of differently modified surfaces were compared using model proteins with different surface charges, such as streptavidin (pI=5, negatively charged at pH 7), horseradish peroxidase (pI=6.6, nearly neutral at pH 7), and avidin (pI=10, positively charged at pH 7). The application of the plasma-treated parylene-N film as an SPR biosensor was also tested by immobilizing model proteins. An SPR biosensor based on the plasma-treated parylene-N film was developed for the detection of the human hepatitis virus surface antigen (HBsAg), and the plasma-treated parylene-N film was estimated to improve the sensitivity of SPR biosensor as much as 1000-fold by enhancing immobilization of receptor antibodies.

One step immobilization of peptides and proteins by using modified parylene with formyl groups.

One-step immobilization method for peptides and proteins is developed by using modified parylene film with formyl groups which is suitable for microplate-based immunoassay and SPR biosensor application. The immobilization of peptides and proteins is achieved through the covalent bonding of the formyl group with the primary amine groups of peptides and proteins, which no additional activation step is required. In this work, the immobilization efficiency of parylene-H is estimated in comparison with parylene-A and physical adsorption, using biotinylated-cyclic citrullinated peptide (biotinylated-CCP), human chorionic gonadotropin (hCG) and horseradish peroxidase (HRP) as model proteins. The applicability of parylene-H film to SPR biosensor is demonstrated by estimating the detection range and sensitivity of SPR biosensor at various thicknesses. The immobilization efficiency of parylene-H film for SPR biosensor was compared with physical adsorption by using HRP as a model protein.