1,490 research outputs found
Techniques for fractionation, isolation and characterization of subvisible and submicron particles in biopharmaceutical preparations
No full textThere is an increased interest from industry, academia and regulators for protein aggregates, subvisible and visible particles due to possible biological consequences, such as immunogenicity, altered bioactivity and modified pharmacokinetic profiles. Aggregates, subvisible and visible particles are important product instabilities, which might be present in every formulation of biotherapeutic products like monoclonal antibody solutions. Especially, the presence of subvisible particles in biotherapeutic products is currently a hot topic and it constantly gains more importance.
The ultimate goal of this thesis is to develop tools and techniques in order to be able to characterize well-defined size fractions of proteinaceous subvisible particles with various desired oxidation profiles using in vivo transgenic mouse model. Up to now only a few articles were published and the available data from in vitro and in vivo experiments on aggregates and subvisible particles is often conflicting and fragmented, which impedes the development of sound conclusions. Moreover, complex mixtures of monomers, aggregates, particles and other degradants were used to draw conclusions.
Only estimated values of protein particle density were used up to now in published studies although it is required to know the density of the measured particles in order to accurately calculate their dimensions and mass. The first aim of this thesis was therefore to develop a method to measure experimentally the protein particle density without extrapolation (Chapter 1). The density for commercially available standard beads (polystyrene, polymethacrylate and melamine) and a large bench of stressed proteinaceous samples was determined with the use of the resonant mass measurement instrument (RMM, Archimedes) and its ability to measure the buoyant mass of individual particles. Various fluids with increasing densities were implemented in order to determine the neutral buoyant mass where the particle density equals the fluid density.
Chapter 2 reports the development of a process to isolate well-defined subvisible fractions using differential centrifugation for a model IgG1 antibody. The process to separate four fractions in the submicron and micrometer size range was developed and successfully optimized through the use of a design of experiments. The centrifugation technique was compared to an already published fractionation method using a preparative fluorescence-activated cell sorter. Efficiency, advantages and drawbacks for both methods were compared and discussed (Chapter 2).
Oxidation profile of aggregates and subvisible particles seem to be an important attribute regarding induced biological consequences. That is why the next goal was to develop a method for selective oxidation of methionine and tryptophan residues in a model mAb in order to be able to delineate the effects and the contribution of individual protein modifications in the primary structure. This included a large set of experiments where different reaction conditions such as temperature of incubation, reaction time, type and concentration of oxidant (t-BHP, H2O2, AAPH) were evaluated in presence (or not) of a large excess of anti-oxidant (free amino acids) in order to protect the corresponding amino acid in a model antibody of the IgG1 subtype (Chapter 3).
To complete the work, unfractionated materials and well-defined size fractions (with well-established oxidation profile) were prepared using the established tools (Chapter 1-3). Those samples were deeply characterized and injected subcutaneously into wild type and transgenic mice for immunization. Anti-drug antibody levels were measured following ELISA in order to assess the immunogenic potential of those preparations (Chapter 4)
The PANS and PITM model: a new formulation of f_k
No full textThe partially averaged Navier-Stokes (PANS) model, proposed in Girimaji (2006), can be used to simulate turbulent flows either as a RANS, LES or DNS. The Partially Integrated Transport Model (PITM) is identical to PANS except that in PANS the diffusion coefficients in the k and epsilon are modified. Both models include f_k which denotes the ratio of modeled to total kinetic energy. In RANS, f_k=f_kH$-equivalence introduced by Friess et al. 2015
The PANS and PITM model: a new formulation of f_k
No full textThe partially averaged Navier-Stokes (PANS) model, proposed in Girimaji (2006), can be used to simulate turbulent flows either as a RANS, LES or DNS. The Partially Integrated Transport Model (PITM) is identical to PANS except that in PANS the diffusion coefficients in the k and epsilon are modified. Both models include f_k which denotes the ratio of modeled to total kinetic energy. In RANS, f_k=f_kH$-equivalence introduced by Friess et al. 2015
Predictors of mortality in ARDS patients referred to a tertiary care centre: a pilot study
No full textBackground and objective: In order to identify parameters predicting intensive care unit mortality in patients transferred to a specialized tertiary centre because of progressive acute respiratory distress syndrome, an observational pilot study was carried out involving 94 patients. Methods and Results: Forty-one patients (43.6%) died. Survival was defined as intensive care unit discharge. Survivors were younger (32.0 +/- 11.8 vs. 39.1 +/- 12.4 yr, P = 0.008), at admission they had a lower acute physiology and chronic health evaluation (APACHE) II score (21.7 +/- 5.4 vs. 25.4 +/- 5.2, P = 0.0009), higher PaO2/FiO2 (122 +/- 79 vs. 79 +/- 42 mmHg, P = 0.002), lower positive end-expiratory pressure (10.6 +/- 3.1 vs. 12.5 +/- 3.7 cmH(2)O, P = 0.02) and a lower Murray score (2.8 +/- 0.63 vs. 3.0 +/- 0.62, P = 0.04). No differences were observed for tidal volumes and peak inspiratory pressures. Days of hospitalization and mechanical ventilation prior to transferral were not related to survival. Multivariate analysis of variables assessed on admission detected only differences for age (P = 0.014) and APACHE II (P = 0.005). Odds ratio was 1.06 (95% confidence interval (CI): 1.013-1.119) for age and 1.21 (CI: 1.059-1.381) for APACHE II. Multivariate analysis of changes in respiratory parameters, APACHE II and Murray score during the first 3 days after transferral revealed a significant difference only for positive end-expiratory pressure (P < 0.008). Corresponding odds ratio was 2.40 (CI: 1.25-4.58) for an increase of 1 cmH(2)O/24 h. Conclusion: Age-related mortality in this small, but highly selected group of patients with established ARDS increased early in life even in a population with an overall mean age of 35.1 yr. APACHE II was the only clinical predictor for mortality on admission. The need for a substantial increase in positive end-expiratory pressure after transferral markedly reduced the chance to survive
Immune cells infiltration and the expression of growth associated protein-43 expression correlate with pain in chronic pancreatisis
No full textCorrigendum to “The 2016 update of the International Study Group (ISGPF) definition and grading of postoperative pancreatic fistula: eleven years after.” Surgery 2017. Mar; 161 (3):584–591. Epub Dec 28, 2016 (Surgery (2017) 161(3) (584–591), (S0039606016307577), (10.1016/j.surg.2016.11.014))
No full textThe authors regret that the name of author Charles R. Vollmer MD is incorrect in the final published version. The correct name Charles Vollmer. The authors would like to apologise for any inconvenience caused. Below is the correct order of authors: Claudio Bassi, MDa, Giovanni Marchegiani, MDa, Christos Dervenis, MD,b, Micheal Sarr, MDc, Mohammad Abu Hilal, MDd, Mustapha Adham, MDe, Peter Allen, MDf, Roland Andersson, MDg, Horacio J. Asbun, MDh, Marc G. Besselink, MDi, Kevin Conlon, MDj, Marco Del Chiaro, MDk, Massimo Falconi, MDl, Laureano Fernandez-Cruz, MDm, Carlos Fernandez-del Castillo, MDn, Abe Fingerhut, MDo, Helmut Friess, MDp, Dirk J Gouma, MDi, Thilo Hackert, MDq, Jakob Izbicki, MDr, Keith D. Lillemoe, MDn, John P. Neoptolemos, MDs, Attila Olah, MDt, Richard Schulick, MDu, Shailesh V. Shrikhande, MDv, Tadahiro Takada, MDw, Kyoichi Takaori, MDx, William Traverso, MDy, Charles Vollmer, MDz, Christopher L. Wolfgang, MDaa, Charles J. Yeo, MDbb, Roberto Salvia, MDa, Marcus Buchler, MDq, from the International Study Group on Pancreatic Surgery (ISGPS
Differential expression of connective tissue growth factor in inflammatory bowel disease.
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