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Analysis of anti-human CD22 human mAbs affinity and specificity on human PBMCs. a) Affinity determination of anti-human CD22 mAbs using SPR by flowing various concentration of CD22 antibody over CD22 chip-bound. b) Flow cytometry analysis of CD22 mAbs on human PBMC. Human PBMC were labeled with anti human CD19 (APC) and purified CD22 mAbs (clone γ1λ1, γ3λ3-5, γ23κ5-2 or γ27λ26) coupled with Alexa Fluor 568 fluorochrome or with a commercial mouse mAb anti-human CD22 (Ms anti-human CD22). (PPT 666 kb

Evidence of Transfer by Conjugation of Type IV Secretion System Genes between Bartonella Species and Rhizobium radiobacter in Amoeba

Background: Bartonella species cospeciate with mammals and live within erythrocytes. Even in these specific niches, it has been recently suggested by bioinformatic analysis of full genome sequences that Lateral Gene Transfer (LGT) may occur but this has never been demonstrated biologically. Here we describe the sequence of the B. rattaustraliani (AUST/NH4 T) circular plasmid (pNH4) that encodes the tra cluster of the Type IV secretion system (T4SS) and we eventually provide evidence that Bartonella species may conjugate and exchange this plasmid inside amoeba. Principal Findings: The T4SS of pNH4 is critical for intracellular viability of bacterial pathogens, exhibits bioinformatic evidence of LGT among bacteria living in phagocytic protists. For instance, 3 out of 4 T4SS encoding genes from pNH4 appear to be closely related to Rhizobiales, suggesting that gene exchange occurs between intracellular bacteria from mammals (bartonellae) and plants (Rhizobiales). We show that B. rattaustraliani and Rhizobium radiobacter both survived within the amoeba Acanthamoeba polyphaga and can conjugate together. Our findings further support the hypothesis that tra genes might also move into and out of bacterial communities by conjugation, which might be the primary means of genomic evolution for intracellular adaptation by cross-talk of interchangeable genes between Bartonella species and plant pathogens. Conclusions: Based on this, we speculate that amoeba favor the transfer of genes as phagocytic protists, which allows fo

The repertoire of ICE in prokaryotes underscores the unity, diversity, and ubiquity of conjugation

Horizontal gene transfer shapes the genomes of prokaryotes by allowing rapid acquisition of novel adaptive functions. Conjugation allows the broadest range and the highest gene transfer input per transfer event. While conjugative plasmids have been studied for decades, the number and diversity of integrative conjugative elements (ICE) in prokaryotes remained unknown. We defined a large set of protein profiles of the conjugation machinery to scan over 1,000 genomes of prokaryotes. We found 682 putative conjugative systems among all major phylogenetic clades and showed that ICEs are the most abundant conjugative elements in prokaryotes. Nearly half of the genomes contain a type IV secretion system (T4SS), with larger genomes encoding more conjugative systems. Surprisingly, almost half of the chromosomal T4SS lack co-localized relaxases and, consequently, might be devoted to protein transport instead of conjugation. This class of elements is preponderant among small genomes, is less commonly associated with integrases, and is rarer in plasmids. ICEs and conjugative plasmids in proteobacteria have different preferences for each type of T4SS, but all types exist in both chromosomes and plasmids. Mobilizable elements outnumber self-conjugative elements in both ICEs and plasmids, which suggests an extensive use of T4SS in trans. Our evolutionary analysis indicates that switch of plasmids to and from ICEs were frequent and that extant elements began to differentiate only relatively recently. According to the present results, ICEs are the most abundant conjugative elements in practically all prokaryotic clades and might be far more frequently domesticated into non-conjugative protein transport systems than previously thought. While conjugative plasmids and ICEs have different means of genomic stabilization, their mechanisms of mobility by conjugation show strikingly conserved patterns, arguing for a unitary view of conjugation in shaping the genomes of prokaryotes by horizontal gene transfer

Polymethacrylates.Material Selection For Medical Applications:Requirements For Several Kinds of Medical Applications

This chapter reviews several cases of methacrylate-based polymers used for medical applications. The main chemicals and fillers used for elaborating biomaterials are presented, together with the main synthesis reactions. Their properties are recalled and discussed using the well-established structure-properties relationships of polymer physicochemistry. Last, the main degradation mechanisms are recalled, together with their consequences on the engineering properties of polymethacrylates, in order to predict the long-term in vivo behavior of such complex materials

Antigen-specific single B cell sorting and expression-cloning from immunoglobulin humanized rats: a rapid and versatile method for the generation of high affinity and discriminative human monoclonal antibodies

International audienceBackground: There is an ever-increasing need of monoclonal antibodies (mAbs) for biomedical applications andfully human binders are particularly desirable due to their reduced immunogenicity in patients. We have applied astrategy for the isolation of antigen-specific B cells using tetramerized proteins and single-cell sorting followed byreconstruction of human mAbs by RT-PCR and expression cloning.Results: This strategy, using human peripheral blood B cells, enabled the production of low affinity human mAbsagainst major histocompatibility complex molecules loaded with peptides (pMHC). We then implemented thistechnology using human immunoglobulin transgenic rats, which after immunization with an antigen of interestexpress high affinity-matured antibodies with human idiotypes. Using rapid immunization, followed by tetramerbasedB-cell sorting and expression cloning, we generated several fully humanized mAbs with strong affinities,which could discriminate between highly homologous proteins (eg. different pMHC complexes).Conclusions: Therefore, we describe a versatile and more effective approach as compared to hybridoma generationor phage or yeast display technologies for the generation of highly specific and discriminative fully human mAbs thatcould be useful both for basic research and immunotherapeutic purposes