Reduction of medium consumption in perfusion mammalian cell cultures using a perfusion rate equivalent concentrated nutrient feed

Media preparation for perfusion cell culture processes contributes significantly to operational costs and the footprint of continuous operations for therapeutic protein manufacturing. In this study, definitions are given for the use of a perfusion equivalent nutrient feed stream which, when used in combination with basal perfusion medium, supplements the culture with targeted compounds and increases the medium depth. Definitions to compare medium and feed depth are given in this article. Using a concentrated nutrient feed, a 1.8-fold medium consumption (MC) decrease and a 1.67-fold increase in volumetric productivity (PR) were achieved compared to the initial condition. Later, this strategy was used to push cell densities above 100 × 10⁶ cells/ml while using a perfusion rate below 2 RV/day. In this example, MC was also decreased 1.8-fold compared to the initial condition, but due to the higher cell density, PR was increased 3.1-fold and to an average PR value of 1.36 g L⁻¹ day⁻¹ during a short stable phase, and versus 0.46 g L⁻¹ day⁻¹ in the initial condition. Overall, the performance improvements were aligned with the given definitions. This multiple feeding strategy can be applied to gain some flexibility during process development and also in a manufacturing set-up to enable better control on nutrient addition.     

Andean and California condors possess dissimilar genetic composition but exhibit similar demographic histories

While genetic diversity of threatened species is a major concern of conservation biologists, historic patterns of genetic variation are often unknown. A powerful approach to assess patterns and processes of genetic erosion is via ancient DNA techniques. Herein, we analyzed mtDNA from historical samples (1800s to present) of Andean Condors (Vultur gryphus) to investigate whether contemporary low genetic variability is the result of recent human expansion and persecution, and compared this genetic history to that of California condors (Gymnogyps californianus).We then explored historic demographies for both species via coalescent simulations. We found that Andean condors have lost at least 17% of their genetic variation in the early 20th century. Unlike California condors, however, low mtDNA diversity in the Andean condor was mostly ancient, before European arrival. However, we found that both condor species shared similar demographies in that population bottlenecks were recent and co‐occurred with the introduction of livestock to the Americas and the global collapse of marine mammals. Given the combined information on genetic and demographic processes, we suggest that the protection of key habitats should be targeted for conserving extant genetic diversity and facilitate the natural recolonization of lost territories, while nuclear genomic data should be used to inform translocation plans.     

Before and After Prozac: Psychiatry as Medicine,and the Historiography of Depression

Culture, Medicine, and Psychiatry - This article examines the historiography of depression, with an eye to illuminating wider issues in the social study of psychiatry and depression. It argues that...     

Soil organic carbon fractions in the Great Plains of the United States: an application of mid-infrared spectroscopy

Biogeochemistry - Spectroscopy is a powerful means of increasing the availability of soil data necessary for understanding carbon cycling in a changing world. Here, we develop a calibration...     

Cell engineering method using fluorogenic oligonucleotide signaling probes and flow cytometry

Objective

Chromovert® Technology is presented as a new cell engineering technology to detect and purify living cells based on gene expression.

Methods

The technology utilizes fluorogenic oligonucleotide signaling probes and flow cytometry to detect and isolate individual living cells expressing one or more transfected or endogenously-expressed genes.

Results

Results for production of cell lines expressing a diversity of ion channel and membrane proteins are presented, including heteromultimeric epithelial sodium channel (αβγ-ENaC), sodium voltage-gated ion channel 1.7 (NaV1.7-αβ1β2), four unique γ-aminobutyric acid A (GABA_A) receptor ion channel subunit combinations α1β3γ2s, α2β3γ2s, α3β3γ2s and α5β3γ2s, cystic fibrosis conductance regulator (CFTR), CFTR-Δ508 and two G-protein coupled receptors (GPCRs) without reliance on leader sequences and/or chaperones. In addition, three novel plasmid-encoded sequences used to introduce 3′ untranslated RNA sequence tags in mRNA expression products and differentially-detectable fluorogenic probes directed to each are described. The tags and corresponding fluorogenic signaling probes streamline the process by enabling the multiplexed detection and isolation of cells expressing one or more genes without the need for gene-specific probes.

Conclusions

Chromovert technology is provided as a research tool for use to enrich and isolate cells engineered to express one or more desired genes.

     

Preclinical Assessment of the Analgesic Pharmacology of NKTR-181 in Rodents

Cellular and Molecular Neurobiology - Pharmacological evaluation of the mu-opioid receptor (MOR) agonist properties of NKTR-181 in rodent models. Graded noxious stimulus intensities were used in...     

Elicitation Improves the Leaf Area,Enzymatic Activities,Antioxidant Activity and Content of Secondary Metabolites in Achillea millefolium L. Grown in the Field

Salicylic acid (SA) is a plant hormone that stimulates the growth and metabolism of plants, also acting as an abiotic elicitor. This study aimed to evaluate the effect of SA on leaf production, leaf area and synthesis of secondary compounds in yarrow plants. The experiments were conducted under field conditions in two consecutive years and f-received SA foliar applications (T1-control; T2-1.0 mmol L⁻¹ applications at 20, 60 and 100 days after planting (DAP) and T3-1.0 mmol L⁻¹ applications at 100 DAP during 3 days). The exogenous application of SA resulted in increases in leaf area (total and specific), number of leaves and leaf mass ratio of yarrow plants, polyphenolic compounds, phenylalanine ammonia-lyase and chalcone synthase enzymes and the antioxidant activity of the plant extract. The HPLC–DAD–MS/MS analysis of phenolic compounds revealed increases in the amounts of quinic acid and rutin. The results of this research lead us to affirm that SA exerted both the hormonal effect on number of leaves and leaf area, and also acted as eliciting substance.