排序方式: 共有93条查询结果,搜索用时 15 毫秒
81.
Michael J Gramer Ewald TJ van den Bremer Muriel D van Kampen Amitava Kundu Peter Kopfmann Eric Etter David Stinehelfer Justin Long Tom Lannom Esther H Noordergraaf Jolanda Gerritsen Aran F Labrijn Janine Schuurman Patrick HC van Berkel Paul WHI Parren 《MABS-AUSTIN》2013,5(6):962-973
The manufacturing of bispecific antibodies can be challenging for a variety of reasons. For example, protein expression problems, stability issues, or the use of non-standard approaches for manufacturing can result in poor yield or poor facility fit. In this paper, we demonstrate the use of standard antibody platforms for large-scale manufacturing of bispecific IgG1 by controlled Fab-arm exchange. Two parental antibodies that each contain a single matched point mutation in the CH3 region were separately expressed in Chinese hamster ovary cells and manufactured at 1000 L scale using a platform fed-batch and purification process that was designed for standard antibody production. The bispecific antibody was generated by mixing the two parental molecules under controlled reducing conditions, resulting in efficient Fab-arm exchange of >95% at kg scale. The reductant was removed via diafiltration, resulting in spontaneous reoxidation of interchain disulfide bonds. Aside from the bispecific nature of the molecule, extensive characterization demonstrated that the IgG1 structural integrity was maintained, including function and stability. These results demonstrate the suitability of this bispecific IgG1 format for commercial-scale manufacturing using standard antibody manufacturing techniques. 相似文献
82.
Madureira P Baptista M Vieira M Magalhães V Camelo A Oliveira L Ribeiro A Tavares D Trieu-Cuot P Vilanova M Ferreira P 《Journal of immunology (Baltimore, Md. : 1950)》2007,178(3):1379-1387
Certain extracellular proteins produced by several pathogenic microorganisms interfere with the host immune system facilitating microbial colonization and were thus designated virulence-associated immunomodulatory proteins. In this study, a protein with B lymphocyte stimulatory activity was isolated from culture supernatants of Streptococcus agalactiae strain NEM316. This protein, with an apparent molecular mass of 45 kDa, was identified as GAPDH by N-terminal amino acid sequencing. The gapC gene was cloned and expressed in Escherichia coli for the production of a recombinant histidyl-tagged protein. The recombinant GAPDH (rGAPDH), purified in an enzymatically active form, induced in vitro an up-regulation of CD69 expression on B cells from normal and BCR transgenic mice. In addition, rGAPDH induced an increase in the numbers of total, but not of rGAPDH-specific, splenic Ig-secreting cells in C57BL/6 mice treated i.p. with this protein. These in vitro- and in vivo-elicited B cell responses suggest that the B cell stimulatory effect of rGAPDH is independent of BCR specificity. A S. agalactiae strain overexpressing GAPDH showed increased virulence as compared with the wild-type strain in C57BL/6 mice. This virulence was markedly reduced in IL-10-deficient and anti-rGAPDH antiserum-treated mice. These results suggest that IL-10 production, which was detected at higher concentrations in the serum of rGAPDH-treated mice, is important in determining the successfulness of the host colonization by S. agalactiae and they highlight the direct role of GAPDH in this process. Taken together, our data demonstrate that S. agalactiae GAPDH is a virulence-associated immunomodulatory protein. 相似文献
83.
Balassiano IT Bastos Mdo C Madureira DJ Silva IG Freitas-Almeida AC Oliveira SS 《Memórias do Instituto Oswaldo Cruz》2007,102(7):861-866
This study analyzed the involvement of tetA and tetE genes in the tetracycline resistance of 16 strains of genus Aeromonas, isolated from clinical and food sources. Polymerase chain reactions revealed that 37.5% of the samples were positive for tetA, and also 37.5% were tetE positive. One isolate was positive for both genes. Only the isolate A. caviae 5.2 had its resistance associated to the presence of a plasmid, pSS2. The molecular characterization of pSS2 involved the construction of its restriction map and the determination of its size. The digestion of pSS2 with HindIII originated two fragments (A and B) that were cloned separately into the pUC18 vector. The tetA gene was shown to be located on the HindIII-A fragment by PCR. After transforming a tetracycline-sensitive strain with pSS2, the transformants expressed the resistance phenotype and harbored a plasmid whose size was identical to that of pSS2. The results confirmed the association between pSS2 and the tetracycline resistance phenotype, and suggest a feasible dissemination of tetA and tetE among strains of Aeromonas. This study suggests the spreading tetA and tetE genes in Aeromonas in Brazil and describes a resistance plasmid that probably contributes to the dissemination of the resistance. 相似文献
84.
Self-incompatibility in passion fruit: cellular responses in incompatible pollinations 总被引:1,自引:0,他引:1
Hérika C. Madureira Telma N. S. Pereira Maura Da Cunha Denise E. Klein Marcos V. V. de Oliveira Leandro de Mattos Gonçalo A. de Souza Filho 《Biologia》2014,69(5):574-584
Self-incompatibility (SI) is a genetic mechanism in angiosperms that prevents selfing. The SI system in passion fruit (Passiflora edulis Sims) was investigated using hand pollinations. Pollen tube growth was inspected by microscopy, and sequence analysis of potential regulators of this process was carried out. The results revealed that the pollen tubes grew slowly and were often completely arrested in the stigma in an incompatible combination. Under these circumstances the pollen tube was rapidly and significantly rearranged, followed by the rapid deposition of callose in the stigma during the SI response. The structural changes in the pollen grain after an incompatible pollination were investigated using scanning electron microscopy. Furthermore, ultrastructural observations during incompatible interactions showed that the membrane system of the pollen tube was damaged, and fertilisation was not observed or was considerably delayed when compared to compatible interactions. The analysis presented here provides evidence that the passion fruit genome presents similar sequences to those encoding factors involved in SI in different species. These results suggest that, in the SI system of passion fruit, the rejection of an incompatible pollen grain is characterised by drastic structural changes in both pollen and pollen tube. 相似文献
85.
Liane FM Finotelo Paulo JS Amaral Julio C Pieczarka Edivaldo HC de Oliveira Alcides Pissinati Michaela Neusser Stephan Müller Cleusa Y Nagamachi 《BMC evolutionary biology》2010,10(1):189
Background
The New World monkey (Platyrrhini) subfamily Pitheciinae is represented by the genera Pithecia, Chiropotes and Cacajao. In this work we studied the karyotypes of Pithecia irrorata (2n = 48) and Cacajao calvus rubicundus (2n = 45 in males and 2n = 46 in females) by G- and C-banding, NOR staining and chromosome painting using human and Saguinus oedipus whole chromosome probes. The karyotypes of both species were compared with each other and with Chiropotes utahicki (2n = 54) from the literature. 相似文献86.
Ronald van Eijk Paul HC Eilers Remco Natté Anne-Marie Cleton-Jansen Hans Morreau Tom van Wezel Jan Oosting 《BMC bioinformatics》2010,11(1):67
Background
Multiplex Ligation-Dependent Probe Amplification (MLPA) is an application that can be used for the detection of multiple chromosomal aberrations in a single experiment. In one reaction, up to 50 different genomic sequences can be analysed. For a reliable work-flow, tools are needed for administrative support, data management, normalisation, visualisation, reporting and interpretation. 相似文献87.
88.
Pycnidiospores of Phyllosticta ampelicida, the causal agent of black rot of grape, were found to germinate only on substrata on which they were firmly attached. Such surfaces were poorly wettable and had advancing contact angles (straight thetaa) formed by a water drop of >80°, e.g., grape leaf, polystyrene, Teflon, polycarbonate, collodion, and glass treated with the silanes n-octadecyltrichlorosilane, dimethyldichlorosilane, or diphenyldichlorosilane. When pycnidiospores were deposited on more wettable surfaces they did not attach firmly and did not germinate. Such highly wettable surfaces had straight thetaa = 40° and were represented by heat-treated glass, cellophane, nutrient- and water-agars, polystyrene treated with UV-irradiation or sulfuric acid, and glass silanized with n-2-aminoethyl-3-aminopropyltrimethoxysilane, n-(trimethoxysilylpropyl)ethylenediamine triacetic acid trisodium, or 3-aminopropyltriethoxysilane. Adhesion of pycnidiospores was assessed with and without a hydraulic shearing force. Pycnidiospore adhesion occurred over several minutes in distilled deionized water, unless it was first acidified, which decreased attachment time to <0.03 s. Attachment of pycnidiospores treated with sodium azide, formaldehyde, or boiled in water for 10 min was similar to nontreated conidia. Possible mechanisms of adhesion of the conidia to surfaces include hydrophobic and ionic interactions. 相似文献
89.