Genetic Engineering & Biotechnology News

DEC 2018

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GENengnews.com | Genetic Engineering & Biotechnology News | DECEMBER 2018 | 19 measure the IgG production rate of single cells in a sample population, using a custom- ized pair of IgG-specific fluorescent probes (Figure 2A). To validate the assay, five popu- lations of picodroplets generated from me- dium containing varying concentrations of IgG were pooled together, detected by Cyto- Mine and analyzed using Cyto-Mine Studio Software. Five discrete picodroplet popula- tions could be resolved, corresponding to the different IgG concentrations, confirming that the assay could quantitate IgG secretion (Figures 2B & 2C). The IgG assay was then used to screen a heterogeneous pool of Chinese hamster ovary (CHO) cells, stably transfected to express human IgG, to identify and isolate the highest antibody-producing clones. The population of picodroplets to the upper left of the scatterplot (Figure 2D) with high acceptor-to-donor fluorescence ratio was gated for collection. The bright cluster to the lower right represents the bulk of data points, which comprises picodroplets con- taining low- or nonproducing cells and empty picodroplets. Antigen-Specicity Assay for Antibody Discovery Sphere Fluidics also carried out a study to screen hybridomas for antigen-specific clones generated from a mouse immu- nized with human tumor necrosis factor-α (TNF-α). The detection probes comprised fluorescently conjugated human TNF-α and a mouse IgG-Fc acceptor probe, en- abling specific detection of only mouse IgG-recognizing human TNF-α (Figure 3A). The choice of probes permits simulta- neous screening for antigen-specificity and immunoglobulin isotype. Titration experiments were undertaken with culture medium picodroplets contain- ing different concentrations of anti-human TNF-α IgG. Different titers could be re- solved into discrete picodroplet populations, and the control IgG experiment demonstrat- ed the fidelity of the assay for detecting only antigen-specific IgG (Figures 3B & 3C). A mixed population of hybridoma cells was analyzed with the validated detection probes to find TNF-α-specific IgG-producing clones. A subpopulation of cells with a high acceptor-to-donor fluorescence ratio, indi- cating secretion of human TNF-α-specific IgG, was gated for collection, while the ma- jority of picodroplets were diverted to waste (Figure 3D). Beyond Antibody Discovery Biopharmaceutical organizations are un- der increasing pressure to streamline their antibody discovery and cell line development processes, with unmet needs for increased throughput, shortened timelines, reduced costs, and improved proof of monoclonal- ity. Next-generation, single-cell analysis plat- forms like the Cyto-Mine allow researchers to screen hundreds of thousands of individu- al cells or up to 40 million cells (in pools) for secreted target protein, to isolate high-value cells or pools of interest, and to dispense with high viability into microplate wells—and to accomplish these tasks more efficiently by using one integrated and automated system. The system can be tailored to a range of re- quirements and workflows in bioproduction and other biological areas, such as genome editing, single-cell disease diagnostics, and metagenomics. All could benefit from the improved efficiencies that are achieved by harnessing picodroplet technology. How a Top-5 Pharma Doubled the Speed of Cell Line Development Single cell cloning (SCC) is a critical and high value process for an increasing number of applications including cell engineering; stable cell lines for therapeutic mAbs and biosimilars; vector production for gene therapy; and many more. During this webinar, Tom Kelly from Janssen (part of Johnson & Johnson) will talk about his use of Solentim's VIPS™ (Veriƒed in-situ Plate Seeding) system, the world's ƒrst All-in-One Single Cell Cloning System. He will explain why Janssen moved away from older generation CLD technology using Clonepix, which was based on detecting and picking high-producer clones in semi-solid media. Tom will explain how, through using VIPS - in combination with improvements in vector design and transfection methods—Janssen's clones are all high producers and only one round of cloning is now necessary. Tom will show examples from Janssen's own data to illustrate each of the key metrics for a single cell cloning device: • Seeding eŒciency • Cloning eŒciency • Ghost wells (false negatives) He will talk about the overall beneƒts that VIPS has provided, why they have purchased multiple VIPS systems and their vision for the future. The webinar will also explain how VIPS sits at the heart of an integrated portfolio of Solentim products that enable small CLD teams to go from single clone isolation to Master Cell Bank. A live Q&A session will follow the presentation, o"ering you a chance to pose questions to our expert panelists. Free Registration! www.genengnews.com/VIPS View It Now! On Demand DURATION: 60 minutes COST: Complimentary Speakers Tom Kelly Scientist, Janssen R&D Ian Taylor, Ph.D. Commercial Director, Solentim Produced with support from Webinars Bioprocessing Frank F. Craig, Ph.D. (frank.craig@ spherefluidics.com), is CEO of Sphere Fluidics. Website: www.spherefluidics.com. Figure 3. Cyto-Mine antigen-specicity assay. (A) Antigen-specic IgG secreted from the encapsulated cell is recognized by the •uorescently conjugated antigen and the IgG-specic probe. (B) Scatterplots of FRET signal from picodroplets with the indicated concentrations of anti-human TNF-α IgG, detected by •uorescently conjugated human TNF-α and an IgG-specic acceptor probe. (C) Representative titration curve generated from data in B. (D) Scatterplot of FRET signal generated from hybridomas screened for secretion of anti-human TNF-α IgG. A C B D

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