ABSTRACT
Detection of biologically active compounds is one of the most important topics in molecular biology and biochemistry. One of the most promising detection methods is based on the application of surface plasmon resonance for label-free detection of biologically active compounds. This method allows one to monitor binding events in real time without labeling. The system can therefore be used to determine both affinity and rate constants for interactions between various types of molecules. Here, we describe the application of a surface plasmon resonance biosensor for label-free investigation of the interaction between an immobilized antigen bovine serum albumin (BSA) and antibody rabbit anti-cow albumin IgG1 (anti-BSA). The formation of a self-assembled monolayer (SAM) over a gold surface is introduced into this laboratory training protocol as an effective immobilization method, which is very promising in biosensing systems based on detection of affinity interactions. In the next step, covalent attachment via artificially formed amide bonds is applied for the immobilization of proteins on the formed SAM surface. These experiments provide suitable experience for postgraduate students to help them understand immobilization of biologically active materials via SAMs, fundamentals of surface plasmon resonance biosensor applications, and determination of non-covalent biomolecular interactions. The experiment is designed for master and/or Ph.D. students. In some particular cases, this protocol might be adoptable for bachelor students that already have completed an extended biochemistry program that included a background in immunology.
ABSTRACT
The aim of this study was to examine the effect of caffeine on the activity of lysozyme and some immune parameters of mice. The mice were divided into five groups. Group 1, the control group, was given water. The other four groups were administered various concentrations of caffeine by oral intubation (group 2, 2 mg x kg(-1); group 3, 20 mg x kg(-1); group 4, 40 mg x kg(-1); group 5, 200 mg x kg(-1)). It was found that the activity of lysozyme in the serum depended on the caffeine dose. Compared with the control (group 1), lysozyme activity was 1.4-times higher in group 2, 1.6-times higher in the group 3, and 1.8-times higher in groups 4 and 5 (P < 0.05). In group 3 a significant increase in spleen weight was detected and the spleen index was 2.1-times (P < 0.05) higher compared with control. In group 3 the number of monocytes and neutrophils was 2.5-times higher (P < 0.05) compared with control. In group 5 the caffeine increased the number of neutrophils 2.7-times and increased the number of eosinophils 4.6-times (P < 0.05) compared with control. Our study revealed that caffeine played an important role in the development of protective immune response.
