Ion channel sensors for glutamic acid

Abstract

Coulometric biosensors using glutamate receptor (GluR) ion channel protein as a signal-amplifying sensory element that exploit the glutamate-triggered Na+ ion current through bilayer lipid membranes have been fabricated. The formation of stable planar bilayer lipid membranes was achieved by applying the folding method across a small circular aperture bored through a thin polyimide film. The multichannel type sensing membranes, formed across an aperture of ca. 120 microns diameter, contained more than 10 GluR proteins and showed L-glutamate-triggered response as a composite of individual single-channel currents. The single-channel type sensing membranes, formed across an aperture of ca. 20 microns diameter, contained a sufficiently small number of GluR proteins so that the response was observed as a series of single-channel pulse currents. Dependence of the integrated channel current on the glutamate concentration was examined. A sharp concentration dependence of up to ca. 1.5 x 10(-7) M and 3 x 10(-6) M for the multichannel and single-channel type sensors, respectively, was observed. A high selectivity for L-glutamate compared with D-glutamate for inducing the channel current was observed. A detection limit as low as ca. 3 x 10(-8) M was attained for the multichannel type sensor. This remarkable sensitivity is discussed in terms of the potential use of GluR ion channel protein for a new type of sensing system.

An Evaluation of Signal Amplification by the Ion Channel Sebsir Based on a Glutamate Receptor Ion Channel Protein

“…Whereas the detection limit of the sensors based on the cyclodextrin derivative could be regarded as in the order of 10`4 M [4,6,9], that of the present sensor based on the GIuR ion channel protein as a sensory element was found to be lower than 3 x 10"7 M [8]. Considering the quite large numbers (10121013) of the cyclodextrin molecules contained in the sensing membranes [4,6,9], the detection limit of the GIuR ion channel sensor that is lower than those of the artificial systems by a factor greater than 10 can be regarded as extremely low, because the present ion channel sensor contained only a single channel protein.…”

“…With this single-channel sensor, a detection limit lower than 10"l M was attained. Even lower detection limit (< 10"$ M) was attained with the multi-channel sensor containing ten or more channel proteins [8]. The remarkable detection limits of these ion channel sensors are evidently based on the intense signalamplifying ability of the G1uR ion channel protein, as supported by an estimation in our previous study that the average number of ions permeated through the channel during each of the channel-open period is ca.…”

“…Although this value contains some ambiguity due to the inaccuracy of the number of the channel proteins incorporated, the molecular response ratios of both single-and multichannel sensors were, as expected, in the same order of magnitude. As a result, the integrated activity of the channel proteins incorporated in the multi-channel sensor lead to a higher sensitivity (steeper slope of the calibration curve) and a lower detection limit as compared to the single-channel sensor [8].…”

“…We have recently reported a series of fundamental studies aimed at a new class of sensors that would be designated as "ion channel sensors" [2][3][4][5][6][7][8]. In the studies based on artificial systems, the "molecular response ratio" has been calculated as an indication of signal amplification [2][3][4]6].…”

Factors Affecting Background Permeabilities of Ordered Mono-, Bi- and Multilayers as Channel Mimetic Sensing Membranes

“…The apparatus used for the formation of BLMs and for the conductance measurements was described in our previous papers. 16,17 Monolayer preparation, measurements of it-A isotherms and film deposition…”