Patterning Functional Proteins with High Selectivity for Biosensor Applications

Abstract

In this article, two-dimensional hexamethyldisilazane (HMDS) micropatterns were generated on glass substrates using photolithographic techniques for the assembly of functional proteins. The non-HMDS patterned areas were backfilled with poly(ethylene glycol) (PEG) silane to reduce the nonspecific protein adsorption. The hydrophobic methyl-terminated HMDS monolayer was verified to be favorable for physical protein adsorption with bovine serum albumin (BSA). The PEG-silane derivatized surface significantly reduced the BSA nonspecific binding by 97% compared to the pristine glass substrate so that high patterning selectivity was achieved. A universal streptavidin template was generated using preadsorbed biotinylated BSA on HMDS surface to sequentially bind additional biotinylated antibodies. Using this patterning strategy, the biotinylated goat anti-mouse (biotin-GAM) antibodies can be specifically recognized by the fluorescently labeled mouse immunoglobulin G, which indicated that the immobilized biotin-GAM was still bioactive. Also, the immobilized alkaline phosphatase was demonstrated to retain its enzymatic functionality by the ability to convert its fluorogenic substrate fluorescein diphosphate into fluorescent products. This simple and effective protein patterning technique can also be extended to create nanoscale protein arrays. Additionally, its adaptability for the assembly of arbitrary proteins and antibodies provides great potentials for biosensor and biomicroelectromechanical systems (MEMS) applications.

Ormocomp-Modified Glass Increases Collagen Binding and Promotes the Adherence and Maturation of Human Embryonic Stem Cell-Derived Retinal Pigment Epithelial Cells

“…Treatment with organosilicon compound HMDS also increased the hydrophobicity to a value close to that of the Ormocomp-treated glass. This is not unexpected because HMDS is also known to bind covalently to the surface hydroxyl groups with the methyl groups increasing the surface hydrophobicity …”

“…This is not unexpected because HMDS is also known to bind covalently to the surface hydroxyl groups with the methyl groups increasing the surface hydrophobicity. 50 The binding of Ormocomp to the borosilicate glass surface was further confirmed by the XPS results, which indicated that the Ormocomp treatment altered the chemical state of the glass surface by increasing the number of molecules containing C−O and CO bonds. This is also consistent with previously reported ATR-FTIR results showing that Ormocomp contains polar O−H and CO bonds, which increases the polarity of the surface and may also facilitate cell adhesion.…”

Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids

“…To covalently couple methoxy-PEG-silane (2-[methoxy(polyethyleneoxy)propyl]-trimethoxysilane; J&K Scientific, Beijing, China) to the PWA chip surface, the bare chips were treated in an oxygen plasma for 30 s to clean the surfaces (Mercator Control Systems, Inc., Santa Ana, CA, USA). Then, they were placed in 1 g of methoxy-PEG-silane dissolved in 100 mL of anhydrous toluene with 1% triethylamine as a catalyst overnight at room temperature [68,69]. Before the application of the PWA chip, the physically adsorbed methoxy-PEG-silane moieties were removed by rinsing the chip in ethanol and DI water, sequentially.…”

BioMEMS –Advancing the Frontiers of Medicine

“…These active electronic microarrays provide electronic addressing of probes and increased DNA hybridization rate through the application of appropriate electric field [94]. Several protein chip formats have also been developed due to the advancement of micro fabrication technologies [95-101]. …”