Tethered Lipid Bilayers on Electrolessly Deposited Gold for Bioelectronic Applications

Abstract

This paper presents the formation of a novel biomimetic interface consisting of an electrolessly deposited gold film overlaid with a tethered bilayer lipid membrane (tBLM). Self-assembly of colloidal gold particles was used to create an electrolessly deposited gold film on a glass slide. The properties of the film were characterized using field-effect scanning electron microscopy, energy dispersive spectroscopy, and atomic force microscopy. Bilayer lipid membranes were then tethered to the gold film by first depositing an inner molecular leaflet using a mixture of 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-[3-(2-pyridyldithio)propionate], 1,2-di-O-phytanyl-snglycero-3-phosphoethanolamine (DPGP), and cystamine in ethanol onto a freshly prepared electrolessly deposited gold surface. The outer leaflet was then formed by the fusion of liposomes made from DPGP or 1,2-dioleoylsn-glycero-3-phosphocholine on the inner leaflet. To provide functionality, two membrane biomolecules were also incorporated into the tBLMs: the ionophore valinomycin and a segment of neuropathy target esterase containing the esterase domain. Electrochemical impedance spectroscopy, UV/visible spectroscopy, and fluorescence recovery after pattern photobleaching were used to characterize the resulting biomimetic interfaces and confirm the biomolecule activity of the membrane. Microcontact printing was used to form arrays of electrolessly deposited gold patterns on glass slides. Subsequent deposition of lipids yielded arrays of tBLMs. This approach can be extended to form functional biomimetic interfaces on a wide range of inexpensive materials, including plastics. Potential applications include high-throughput screening of drugs and chemicals that interact with cell membranes and for probing, and possibly controlling, interactions between living cells and synthetic membranes. In addition, the gold electrode provides the possibility of electrochemical applications, including biocatalysis, bio-fuel cells, and biosensors.

Fabrication of highly insulating tethered bilayer lipid membrane using yeast cell membrane fractions for measuring ion channel activity

“…Many membrane proteins retain their active conformation only when embedded in a BLM or have activities that involve direct interaction with a BLM. Improved experimental tools are needed that maintain membrane proteins in an environment that allows their activities to be simultaneously expressed and measured in vitro [1]. Such tools could be used in functional proteomics research to characterize novel proteins, in high-throughput drug screening to investigate receptor-drug interactions, and in development of novel biosensor systems that exploit specific interactions between proteins and analytes [2,3].…”

Neuropathy Target Esterase Biosensor

“…This result suggests the immobilization of approximately 50 ng/cm 2 of active NEST in tBLMs. To our knowledge, this is the first time NEST has been immobilized in an active conformation on a surface (Kohli et al, 2006).…”

“…We have tested both LBL assembly and tBLM methods to fabricate novel nanostructured NEST biosensors, as schemed in Figure 1 (a) & (b) (Kohli et al, 2006;Kohli et al, 2007;Srivastava, 2008). In this Chapter, we will focus on the former approach.…”

“…The molecular architecture of the biosensor interfaces are shown schematically in Figure 1 (a) & (b). Two molecular self-assembly approaches, (a) layer-by-layer (LBL) assembly (Decher, 1997) and (b) tethered lipid bilayer membranes (tBLM) (Kohli et al, 2006), were utilized in constructing nanostructured NEST biosensors. Gold electrodes cleaned in piranha solution were dipped in a 5 mM solution of thioctic acid in ethanol for 30 min.…”

“…For potential step voltammetry experiments, small aliquots of the resulting phenyl valerate micellar solution (5.286 mM nominal concentration) were added to the phosphate buffer to obtain the desired apparent concentrations. (Kohli et al, 2007;Srivastava, 2008), and (b) tethered lipid bilayer membranes (tBLM) adapted with permission from (Kohli et al, 2006). dried films were determined using 44 wavelengths between 414.0 and 736.1 nm.…”