Aspects of "antigen-antibody" interaction of chicken infectious bronchitis virus determined by surface plasmon resonance.

Authors performed investigation on "antigen-antibody" interaction of chicken infectious bronchitis coronavirus (IBV) by a method based on the surface plasmon resonance (SPR). Presence of space-size effect related to a difference between antigen and antibody particle sizes has been theoretically grounded and experimentally proven. Herewith, the difference between responses of the SPR-sensor to specific and non-specific interactions is considerably less (up to 6.3 times) than the expected one (8 - 11 times). An impact of functionalization of sensor's sensitive element surface, as well as acidity of buffer solution on the activity of antigen-antibody interaction was studied here. The difference between sensor's responses to specific and non-specific interactions increased two-fold from 200 to 432ang sec due to this treatment. When changing the acidity of analyzed solution from pH7.3 to pH6.8, the corresponding difference between sensor's responses increased by 6.3 times from 194 up to 1235ang.sec. Thus, an impact of space-size effect on interaction between IBV antigen and specific antibody can be considerably (almost in 3 times) decreased by reducing the acidity of used buffer solution. The results of our investigation can be successfully applied to develop new methods for detection of pathogens and specific antibodies using SPR.

CD150-mediated Akt signalling pathway in normal and malignant B cells.

AIM: To study upstream and downstream events in CD150-mediated Akt signaling pathway in normal human B cells, EBV-transformed lymphoblastoid (LCL) and malignant Hodgkin's lymphoma (HL) B cell lines. METHODS: To access protein-protein interaction we applied immunoprecipitation, Western blot analysis and surface plasmon resonance (SPR) technique. A novel modification of SPR technique using reduced glutathione bound to golden surface was proposed. Immunostaining and isolation of cytoplasmic fractions and nuclear extracts were performed to detect proteins' localization in cells. Western blot analysis was performed to follow up the phosphorylation of proteins on specific sites and proteins' expression level. RESULTS: It was shown that CD150 ligation induced Akt activation in normal tonsillar B cells (TBC), SH2D1A positive LCL and HL B cell lines. The p85α subunit of PI3K co-precipitated with CD150 cytoplasmic tail. This direct association depends on tyrosine phosphorylation and is mediated by N terminal SH2 domain of p85α. CD150 initiated phosphorylation of FoxO1 transcription factor in normal B cells as well as in LCL MP-1 and HL cell line L1236. At the same time, CD150 ligation triggered GSK-3ß kinase phosphorylation only in immortalized LCL MP-1 and HL cell line L1236. CONCLUSIONS: We have demonstrated that CD150 receptor could trigger PI3K-mediated Akt signaling pathway in normal, EBV-transformed and malignant B cells. CD150-mediated phosphorylation of Akt downstream targets GSK-3ß and FoxO1 in EBV-transformed and HL cells could be one of the mechanisms to avoid apoptosis and support survival program in these immortalized B cells.