Research On Bactericidal Mechanism Of Human Lactoferrin And Construction Of Its New Detection Method By Bioelectrochemical Technique

Lactoferrin has many kinds of functions, including antimicrobial. However, thebactericidal mechanism of lactoferrin has not been distinctly elucidated. Sensitive,rapid and simple detection techniques are needed in the diagnosis of the disease, aswell as the product quality control, but the existing detection techniques all have someinsufficient. Therefore, this study is to solve two problems with the method ofbioelectrochemical technology: one is to validate the bactericidal mechanism, and theother is to build an electrochemical detection technology for lactoferrin.Firstly, a bioelectrochemical detection technique was used for lactoferrin and eachstep of the modified electrodes was characterized by cyclic voltammetry andelectrochemical impedance spectroscopy. The result showed that the linear segmentincreased from10pg/mLto1μg/mL with a regression equation of I=-10.94563-6.44048lgC (R2=0.9946，C in g/mL，I in μA)，and the LOD is3.3pg/mL.The optimum conditions for lactoferrin binding to the monoclonal antibody were37℃for incubation temperature and60min for incubation time. Immuno-electrodesshowed good stability up to4weeks under4℃and showed a high specificity andsensitivity to the detection of lactoferrin.Secondly, the bactericidal mechanism of lactoferrin was studied by employingbioelectrochemistry technique. The result showed that LF bound tolipopolysaccharides (LPS), causing release of LPS, which resulted in formation ofchannels in the mimic biomembrane. In addition, a survey of microenvironmentfactors for bactericidal activity of lactoferrin was conducted. Lactoferrin exhibitedgreater bactericidal function in the weakly alkaline conditions of pH8. Thebactericidal activity of lactoferrin was related to lactoferrin concentration and higherconcentration meant stronger bactericidal activity. Meanwhile, the bactericidalactivity decreased with a higher concentration metal ions. Divalent cations, especially the Ca2+showed greater influence on the activity than monovalent cations. In contrast,Fe3+had no effect on the activity.