Monoclonal antibodies are powerful and versatile tools that enable the study of proteins in diverse contexts. They are often utilized to assist with identification of subcellular localization and characterization of the function of target proteins of interest. However, because there can be considerable sequence diversity between orthologous proteins in Xenopus and mammals, antibodies produced against mouse or human proteins often do not recognize Xenopus counterparts. To address this issue, we refined existing mouse monoclonal antibody production protocols to generate antibodies against Xenopus proteins of interest. Here, we describe several approaches for the generation of useful mouse anti-Xenopus antibodies to multiple Xenopus proteins and their validation in various experimental approaches. These novel antibodies are now available to the research community through the Developmental Study Hybridoma Bank (DSHB).

Monocytes are professional antigen presenting cells (APC). They serve as precursors of macrophages and dendritic cells (DC). We have used cytopathic (cp) and non-cytopathic (ncp) Bovine Viral Diarrhea Viruses (BVDV) to determine the genes and proteins expression levels in bovine monocytes. Four specific aims were accomplished in this study. The first aim was to assess the baseline expression of the proteins involved in professional antigen presentation in bovine monocytes. The results showed that the differential detergent fractionation (DDF) approach can provide interpretable and meaningful functional information in bovine monocytes. The second aim was to evaluate the role of in vitro cp and ncp BVDV infection in the expression of the selected bovine genes involved in professional antigen presentation. The results showed that both BVDV could escape innate immune responses by modulating toll-like receptor (TLR) gene expression, followed by pro-inflammatory, type I interferon (IFN), Th1/Th2 type cytokine genes expression, and decreasing the expression levels of CD80/CD86 in professional APC. The third objective was to determine how the two biotypes affect selective antigen uptake, receptor-mediated endocytosis and non-selective uptake, macropinocytosis in bovine monocytes. The results indicated that bovine monocytes use macropinocytosis for a bulklow uptake of soluble antigens. The final aim was to characterize protein profiles in peripheral blood monocytes infected with cp BVDV isolate in vitro. Comparative profiling of the membrane and cytosolic proteins related to professional antigen presentation were assessed. The results showed that 47 bovine proteins, involved in immune function of professional APC have been significantly altered after cp BVDV infection. Overall, we hypothesize that by modulating expression levels of multiple proteins and genes related to immune responses BVDV could significantly compromise immune defense mechanisms resulting in uncontrolled immune activation or suppression.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.