Research about the Endotoxin Detection System Using of Quartz Crystal Microbalance

摘要

Hemodialysis is the most popular treatment method for chronic renal failure. However, hemodialysis patients must continue the treatment throughout their life spans; and long-term extracorporeal dialysis can lead to various complications and diseases, such as, for example, dialysis-related amyloidosis, which is one of the major refractory complications associated with dialysis and has recently been linked to endotoxin. Endotoxin is one of the major components of the cell walls of gram-negative bacteria; they have various biological activities. In addition, it is known that some amount of endotoxin exists in the living environment, and medicines are often contaminated with endotoxin. Dialysis-related amyloidosis results from the use of dialyzing fluids contaminated with endotoxin. This shows the importance of the detection and removal of endotoxin from dialyzing fluids. Currently, limulus amebocyte lysate (LAL) tests are used to detect the presence of endotoxin. However, there are several methodologies to conduct this test, for example, the gelatinization method, turbidimetric assay method, colorimetric assay method, and fluoroscopic method. And each of these requires 30-60 min for completion. Thus, LAL tests cannot be used as for real-time endotoxin detection, and a new detection method is necessary. In this research, we studied the adsorption reaction between ε-polylysine and endotoxin. ε-polylysine is a straight-chain molecule comprising 25-30 lysine residues; it is used as an antimicrobial agent and, moreover, cellulose beads with immobilized ε-polylysine are used the barrier filters for endotoxin removal. The adsorption of endotoxin on to the immobilized ε-polylysine will cause an increase in mass allowing the existence of endotoxin to be immediately detected with a quartz crystal microbalance (QCM). In this report, the immobilization of ε-polylysine on Au and Si substrates and its adsorptive activity are described. We used X-ray photoelectron spectroscopy (XPS) to confirm the immobilization of ε-polylysine, and atomic force microscopy (AFM) and QCM to measure the adsorptive activity. This molecular adsorption-type endotoxin sensor is aimed toward the realization of a real-time endotoxin detection system.