Anti-Beta 2 Microglobulin monoclonal antibody, clone H33-33 [PE]

Global aquaculture production continues to increase rapidly. One of the most important species of marine fish currently cultivated in Southern Europe is Solea senegalensis, reaching more than 300 Tn in 2017. In the present work, 14 Bacterial Artificial Chromosome (BAC) clones containing candidate genes involved in the immune system (b2m, il10, tlr3, tap1, tnf alpha, tlr8, trim25, lysg, irf5, hmgb2, calr, trim16, and mx), were examined and compared with other species using multicolor Fluorescence in situ Hybridization (mFISH), massive sequencing and bioinformatic analysis to determine the genomic surroundings and syntenic chromosomal conservation of the genomic region contained in each BAC clone. The mFISH showed that the groups of genes hmgb2-trim25-irf5-b2m; tlr3-lysg; tnf alpha-tap1, and il10-mx-trim16 were co-localized on the same chromosomes. Synteny results suggested that the studied BACs are placed in a smaller number of chromosomes in S. senegalensis that in other species. Phylogenetic analyses suggested that the evolutionary rate of immune system genes studied is similar among the taxa studied, given that the clustering obtained was in accordance with the accepted phylogenetic relationships among these species. This study contributes to a better understanding of the structure and function of the immune system of the Senegalese sole, which is essential for the development of new technologies and products to improve fish health and productivity.

Murine bone marrow-derived macrophages (M0) and M1- and M2-polarized macrophages are being widely used as a laboratory model for polarized macrophages related molecular mechanism analysis. Gene expression analysis based on reference gene normalization using RT-qPCR was a powerful way to explore the molecular mechanism. But little is known about reference genes in these cell models. So, the goal of this study was to identify reference genes in these types of macrophages. Candidate reference genes in murine bone marrow-derived and polarized macrophages were selected from microarray data using Limma linear model method and evaluated by determining the stability value using five algorithms: BestKeeper, NormFinder, GeNorm, Delta CT method, and RefFinder. Finally, the selected stable reference genes were validated by testing three important immune and inflammatory genes (NLRP1, IL-1 beta, and TNF-alpha) in the cell lines. Our study has clearly shown that Ubc followed by Eef1a1 and B2m respectively were recognized as the three ideal reference genes for gene expression analysis in murine bone marrow-derived and polarized macrophages. When three reference genes with strong different stability were used for validation, a large variation of a gene expression level of IL-1 beta, TNF-alpha and NLRP1 were obtained which provides clear evidence of the need for careful selection of reference genes for RT-qPCR analysis. Normalization of mRNA expression level with Ubc rather than Actb or Gusb by qPCR in macrophages and polarized macrophages is required to ensure the accuracy of the qPCR analysis.