The molecular adsorption-type endotoxin detection system using immobilized ε-polylysine

Hemodialysis for chronic renal failure is the most popular treatment method with artificial organs. However, hemodialysis patients must continue the treatment throughout their life, the results of long term extracorporeal dialysis, those patients develop the various complications and diseases, for example, dialysis amyloidosis etc. Dialysis amyloidosis is one of the refractory complications, and endotoxin is thought to be the most likely cause of it, recently. Endotoxin is one of the major cell wall components of gram-negative bacteria, and it has various biological activities. In addition, it is known that a mount of endotoxin exists in living environment, and medicine is often contaminated with endotoxin. When contaminated dialyzing fluids are used to hemodialysis, above-mentioned dialysis amyloidosis is developed. Therefore, it is important that the detection and removal of endotoxin from dialyzing fluids. Until now, the measurement methods using Limulus Amebosyte Lysate (LAL) reagent were carried out as the tests for the presence of endtoxin. However, these methods include several different varieties of measurement techniques. The following are examples of them, gelatinization method, turbidimetric assay method, colorimetric assay method and fluoroscopic method. However, these techniques needed 30-60 minutes for the measurement. From these facts, they are not able to use as a "real-time endotoxin detector". The detection of endotoxin has needed to carry out immediately, for that reason, a new detection method is desired. In this research, we focused attention to adsorption reaction between ε-polylysine and endotoxin. ε-polylysine has the structure of straight chain molecule composed by 25-30 residues made by lysine, and it is used as an antimicrobial agent, moreover, cellulose beads with immobilized ε-polylysine is used as the barrier filter for endotoxin removal. The endotoxin is adsorbed to immobilized ε-polylysine, as the result of this reaction, the mass incrementation is occurred, and the existence of endtoxin can be detected immediately, by using of Quartz Crystal Microbalance (QCM). In this report, the immobilization of ε-polylysine onto the Au and Si substrate and its adsorptive activity are described. We use X-ray Photoelectron Spectroscopy (XPS) to confirm the ε-polylysine immobilization, and the adsorptive activity of immobilized ε-polylysine is measured by AFM and QCM. This molecular adsorption type endotoxin sensor aims to the realization of "real-time endotoxin detection system".