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Biotechnology Progress

Paula G Miller, Ying I Wang, Glen Swan, Michael L Shuler
Transporting living complex cellular constructs through the mail while retaining their full viability and functionality is challenging. During this process, cells often suffer from exposure to suboptimal life-sustaining conditions (e.g. temperature, pH), as well as damage due to shear stress. We have developed a transport device for shipping intact cell/tissue constructs from one facility to another that overcomes these obstacles. Our transport device maintained three different cell lines (Caco2, A549, and HepG2 C3A) individually on transwell membranes with high viability (above 97%) for 48 h under simulated shipping conditions without an incubator...
June 9, 2017: Biotechnology Progress
Changhao Li, Jingya Li, Christopher Q Lan, Dankui Liao
Cell density and morphology changes were tested to examine the effects of salts including NaHCO3 , NaCl, KHCO3 and KCl at 160 mM on protozoa. It was demonstrated that ionic stress rather than osmotic stress led to protozoa cell death and NaHCO3 was shown to be the most effective inhibitor. Deformation of cells and cell shrinkage were observed when protozoan cells were exposed to PEG or any of the salts. However, while PEG treated cells could fully recover in both number and size, only a small portion of the salt-treated cells survive and cell size was 36% -58% smaller than regular...
June 8, 2017: Biotechnology Progress
Alejandro Rodriguez-Sanchez, Juan Carlos Leyva-Diaz, Alejandro Gonzalez-Martinez, Jose Manuel Poyatos
Three pilot-scale bioreactors were started up and operated under salinity-amended urban wastewater feeding. The bioreactors were configured as membrane bioreactor and two different hybrid, moving bed biofilm reactor-membrane bioreactor and operated with a hydraulic retention time of 9.5 h, a solid residence time of 11.75 days and a total solids concentration of 2500 mg L(-1) . The three systems showed excellent performance in suspended solids, BOD5 , and COD removal (values of 96-100%, 97-99%, and 88-90%, respectively), but poor nitrogen removal (values of 20-30%)...
June 8, 2017: Biotechnology Progress
Amin S M Salehi, Mark T Smith, Song-Min Schinn, Jeremy M Hunt, Christina Muhlestein, Joann Diray-Arce, Brent L Nielsen, Bradley C Bundy
Emancipating sense codons toward a minimized genetic code is of significant interest to science and engineering. A key approach toward sense codon emancipation is the targeted in vitro removal of native tRNA. However, challenges remain such as the insufficient depletion of tRNA in lysate-based in vitro systems and the high cost of the purified components system (PURE). Here we used RNase-coated superparamagnetic beads to efficiently degrade E. coli endogenous tRNA. The presented method removes >99% of tRNA in cell lysates, while partially preserving cell-free protein synthesis activity...
June 8, 2017: Biotechnology Progress
Flor González-Cataño, Luz Tovar-Castro, Eduardo Castaño-Tostado, Carlos Regalado-Gonzalez, Blanca García-Almendarez, Anaberta Cardador-Martínez, Silvia Amaya-Llano
No abstract text is available yet for this article.
June 1, 2017: Biotechnology Progress
Ming Dong, Tara D Gonzalez, Meghan M Klems, Lisa M Steinberg, Wilfred Chen, Eleftherios T Papoutsakis, Brian J Bahnson
Methanol:coenzyme M methyltransferase is an enzyme complex composed of three subunits, MtaA, MtaB and MtaC, found in methanogenic archaea and is needed for their growth on methanol ultimately producing methane. MtaABC catalyzes the energetically favorable methyl transfer from methanol to coenzyme M to form methyl coenzyme M. Here we demonstrate that this important reaction for possible production of methanol from the anaerobic oxidation of methane can be reversed in vitro. To this effect, we have expressed and purified the Methanosarcina barkeri MtaABC enzyme, and developed an in vitro functional assay that demonstrates MtaABC can catalyze the energetically unfavorable (ΔG° = 27 kJ/mol) reverse reaction starting from methyl coenzyme M and generating methanol as a product...
May 27, 2017: Biotechnology Progress
Ingrid K Leth, Karen A McDonald
A chemically defined media was developed for growing Agrobacterium tumefaciens at large scale for commercial production of recombinant proteins by transient expression in plants. Design of experiments was used to identify major and secondary effects of ten media components: sucrose, ammonium sulfate ((NH4 )2 SO4 ), magnesium sulfate heptahydrate (MgSO4 *7H2 O), calcium chloride dihydrate (CaCl2 *2H2 O), iron (II) sulfate heptahydrate (FeSO4 *7H2 O), manganese (II) sulfate monohydrate (MnSO4 *H2 O), zinc sulfate heptahydrate (ZnSO4 *7H2 O), sodium chloride (NaCl), potassium chloride (KCl) and a sodium/potassium phosphate buffer (Na2 HPO4 /KH2 PO4 )...
May 27, 2017: Biotechnology Progress
Kerensa J Klottrup, Guillermo Miro-Quesada, Layla Flack, Ivan Pereda, Pamela Hawley-Nelson
The manufacturing process for biotherapeutics is closely regulated by the Food and Drug Administration (FDA), European Medicines Agency (EMA) and other regulatory agencies worldwide. To ensure consistency of the product of a manufacturing cell line, International Committee on Harmonization guidelines (Q5D, 1997) state that the cell substrate should be derived from a single cell progenitor, i.e., clonal.Cell lines in suspension culture may naturally revert to cell adhesion in the form of doublets, triplets and higher order structures of clustered cells...
May 27, 2017: Biotechnology Progress
Michael Sokolov, Jonathan Ritscher, Nicola MacKinnon, Jonathan Souquet, Hervé Broly, Massimo Morbidelli, Alessandro Butté
This work investigates the insights and understanding which can be deduced from predictive process models for the product quality of a monoclonal antibody based on designed high-throughput cell culture experiments performed at milliliter (ambr-15(®) ) scale. The investigated process conditions include various media supplements as well as pH and temperature shifts applied during the process. First, principal component analysis (PCA) is used to show the strong correlation characteristics among the product quality attributes including aggregates, fragments, charge variants, and glycans...
May 27, 2017: Biotechnology Progress
Md Ruhul Kuddus, Megumi Yamano, Farhana Rumi, Takashi Kikukawa, Makoto Demura, Tomoyasu Aizawa
Snakin-1 (SN-1) is a cysteine-rich plant antimicrobial peptide and the first purified member of the snakin family. SN-1 shows potent activity against a wide range of microorganisms, and thus has great biotechnological potential as an antimicrobial agent. Here, we produced recombinant SN-1 in Escherichia coli by a previously developed coexpression method using an aggregation-prone partner protein. Our goal was to increase the productivity of SN-1 via the enhanced formation of insoluble inclusion bodies in E...
May 27, 2017: Biotechnology Progress
Daniel Strauss, Joshua Goldstein, Tomoko Hongo-Hirasaki, Yoshiro Yokoyama, Naokatsu Hirotomi, Tomoyuki Miyabayashi, Dominick Vacante
Virus filtration provides robust removal of potential viral contaminants and is a critical step during the manufacture of biotherapeutic products. However, recent studies have shown that small virus removal can be impacted by low operating pressure and depressurization. To better understand the impact of these conditions and to define robust virus filtration design spaces, we conducted multivariate analyses to evaluate parvovirus removal over wide ranges of operating pressure, solution pH, and conductivity for three mAb products on Planova™ BioEX and 20N filters...
May 27, 2017: Biotechnology Progress
Cristina García, Manuel Rendueles, Mario Díaz
Besides its properties as an antioxidant, stabilizer or acidifier, lactobionic acid has emerged as a potential prebiotic compound, raising the possibility of being included together with the probiotic microorganism Lactobacillus casei in novel functional fermented foods with synbiotic characteristics. Their manufacturing strategy could benefit from the recently implemented microbial synthesis of lactobionic acid by the strong producer Pseudomonas taetrolens, employing residual dairy whey as raw material. The phenomenon of amensalism established between Pseudomonas and Lactobacillus makes simultaneous fermentation unfeasible...
May 27, 2017: Biotechnology Progress
Michael D Hughson, Thayana A Cruz, Rimenys J Carvalho, Leda R Castilho
The pressures to efficiently produce complex biopharmaceuticals at reduced costs are driving the development of novel techniques, such as in downstream processing with straight-through processing (STP). This method involves directly and sequentially purifying a particular target with minimal holding steps. This work developed and compared 6 different 3-step STP strategies, combining membrane adsorbers, monoliths and resins, to purify a large, complex and labile glycoprotein from Chinese hamster ovary (CHO) cell culture supernatant...
May 27, 2017: Biotechnology Progress
Samanta Hernández-García, María Inmaculada García-García, Francisco García-Carmona
Among enzyme immobilization techniques, the preparation of crosslinked enzyme aggregates has shown promising results in biocatalysis, because they are easy to prepare, versatile and cheap. The method involves the precipitation of enzymes with ammonium sulfate or an organic solvent and subsequent crosslinking with glutaraldehyde. However, the Schiff base produced with glutaraldehyde is reversible and can be broken with acids or bases, releasing proteins to the reaction medium. To solve this problem, we propose replacing glutaraldehyde with diepoxide compounds to obtain an irreversible secondary amine bond...
May 27, 2017: Biotechnology Progress
Chaohong Shi, Nengwu Zhu, Naixin Kang, Pingxiao Wu, Xiaoping Zhang, Yanhong Zhang
Biorecovery is emerging as a promising process to retrieve gold from secondary resources. The present study aimed to explore the uptake pattern of Pycnoporus sanguineus biomass for gold, identify the effective functional groups in gold recovery process, and thus further intensify the process via microbial surface modification. Results showed that P. sanguineus biomass could effectively recover gold with the formation of highly crystal AuNPs without any exogeneous electron donor. Under the conditions of various initial gold concentrations (1...
May 25, 2017: Biotechnology Progress
Marco P Carvalho, Elisabete C Costa, Ilídio J Correia
Drug screening is currently demanding for realistic models that are able to reproduce the structural features of solid tumors. 3D cell culture systems, namely spheroids, emerged as a promising approach to provide reliable results during drug development. So far, Liquid Overlay Technique (LOT) is one of the most used methods for spheroids assembly. It comprises cellular aggregation due to their limited adhesion to certain biomaterials, like agarose. However, researchers are currently improving this technique in order to obtain spheroids on surfaces that mimic cancer extracellular matrix (ECM), since cell-ECM interactions modulate cells behavior and their drug resistance profile...
May 25, 2017: Biotechnology Progress
Nicholas Moscatello, Ruiquan Qi, Mahmoud Kamal Ahmadi, Blaine A Pfeifer
Yersiniabactin (Ybt) is a mixed nonribosomal peptide-polyketide natural product that binds a wide range of metals with the potential to impact processes requiring metal retrieval and removal. In this work, we substantially improved upon the heterologous production of Ybt and an associated anthranilate analog through systematic screening and optimization of culture medium components. Specifically, a Plackett-Burman design-of-experiments methodology was used to screen 22 components and to determine those contributing most to siderophore production...
May 25, 2017: Biotechnology Progress
Yashas Rajendra, Sowmya Balasubramanian, Neil A McCracken, Dawn L Norris, Zhirui Lian, Matthew G Schmitt, Christopher C Frye, Gavin C Barnard
Generating purified protein for GLP toxicology studies (GLP-Tox) represents an important and often rate limiting step in the biopharmaceutical drug development process. Toxicity testing requires large amounts of therapeutic protein (>100g), typically produced in a single 500L - 2,500L bioreactor, using the final CHO clonally-derived cell line (CDCL). One approach currently used to save time is to manufacture GLP-Tox material using pools of high-producing CHO CDCLs instead of waiting for the final CDCL. Recently, we reported CHO pools producing mAb titers >7 g/L using piggyBac-mediated gene integration (PB CHO pools)...
May 25, 2017: Biotechnology Progress
Hongcheng Liu, Christine Nowak, Bruce Andrien, Mei Shao, Gomathinayagam Ponniah, Alyssa Neill
Glycosylation of the conserved asparagine residue in the CH2 domain is the most common posttranslational modification of recombinant monoclonal antibodies. Ideally, a consistent oligosaccharide profile should be maintained from early clinical material to commercial material for the development of recombinant monoclonal therapeutics, though variation in the profile is a typical result of process changes. The risk of oligosaccharide variation posed to further development is required to be thoroughly evaluated based on its impact on antibody structure, stability, efficacy and safety...
May 25, 2017: Biotechnology Progress
Patrícia Aparecida Tozetti, Samia Rigotto Caruso, Amanda Mizukami, Taisa Risque Fernandes, Fernanda Borges da Silva, Fabiola Traina, Dimas Tadeu Covas, Maristela Delgado Orellana, Kamilla Swiech
Choosing the culture system and culture medium used to produce cells are key steps towards a safe, scalable and cost-effective expansion bioprocess for cell therapy purposes. The use of AB human serum (AB HS) as an alternative xeno-free supplement for mesenchymal stromal cells (MSC) cultivation has increasingly gained relevance due to safety and efficiency aspects. Here we have evaluated different scalable culture systems to produce a meaningful number of umbilical cord matrix-derived MSC (UCM MSC) using AB HS for culture medium supplementation during expansion and cryopreservation to enable a xeno-free bioprocess...
May 9, 2017: Biotechnology Progress
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