We report a rapid and simple method to detect endotoxin using a nanopore sensor based on the Coulter principle. An ion current momentary decreases when an endotoxin micelle translocates through the nanopore at a tip of the glass capillary with the voltage biased across the pore. Quantitative detection is available by counting pulsed currents appear within a limited period. First, we characterized the nanopores of fabricated glass capillaries using a scanning electron microscope. The size of the pore at the tip of the glass capillary was 260 nm. Next, endotoxin assays using a nanopipette were performed. An Ag/AgCl wire inserted into a glass capillary filled with phosphate buffer saline (PBS) containing endotoxin was biased at 1 V versus another Ag/AgCl wire inserted in a bulk solution containing endotoxin-free PBS. No pulsed current was observed during 4 min of monitoring when the endotoxin concentrations in the glass capillary were below 100 Endotoxin Unit (EU) L–1. When 500-10000 EU L–1 was in the glass capillary, pulsed currents were observed with an increase of the pulse height and pulse frequency with increasing of the endotoxin concentration. This is because the size of the endotoxin micelles as well as the micelle concentration were increased with the increase of the endotoxin concentration. The fact that pulses appeared far less frequently than expected indicates that only large micelles associated with each other were observed as a pulsed current. Although the current detection limit is 500 EU L–1, optimization of the pore diameter will make it possible to detect even smaller micelles, resulting in an increased the sensitivity. This method is expected to be applied to use as a rapid and simple sensor to detect endotoxin, especially in the continuous monitoring of dialysis solution.
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