3 Ways Microfluidics Is Transforming Biologics
3 Ways Microfluidics Is Transforming Biologics
New technologies are advancing what is possible in the biopharmaceutical industry, and droplet microfluidics is at the forefront of several exciting developments. At Sphere Fluidics’ Discovery Day, which took place at Granta Park, Cambridge on 3 March 2020, we heard from some of the leading experts and researchers in the sector and learnt about how picodroplet technology has redefined processes and workflows in biologics discovery and development.
1. Application of Cyto-Mine® to Antibody Screening
Darren Schofield, a Senior Scientist within the Antibody Discovery and Protein Engineering Department at AstraZeneca, kicked off the Discovery Day’s Antibody Discovery and Development track. Darren focused on innovations including the application of Sphere Fluidics’ Cyto-Mine® as an emerging antibody discovery technology and the sequencing of antibody repertoires from immunised animals at a single cell level. In the recent study performed on a frozen vial of mouse lymphocytes, AstraZeneca used Sphere Fluidics’ Cyto-Mine® to screen up to 40 million cells from immunized animals for subsequent single cell antibody sequencing animals. Researchers first used a bulk IgG FRET assay to detect all IgG-secreting cells. Then in the second screening round, they used an antigen-specific FRET assay to detect cells secreting antigen-specific antibodies. AstraZeneca completed this two-step process in just two days.
An overview of the Cyto-Mine® Antibody Discovery process
Darren was followed by GlaxoSmithKline’s Irene Sanjuan-Nandin and Manjunath Hegde, who while reviewing microfluidic single B cell screening strategies, demonstrated that Cyto-Mine® is a robust method for identifying rare antigen-specific B cells. Their talk also explored a range of challenges and opportunities in the therapeutic monoclonal antibody discovery process, such as the development of multi-parametric screening assays required for antibody discovery.
2. Application of Cyto-Mine® to Cell Line Development
Jonathan Dempsey, the Managing Director of Dempsey Consulting and founding Partner of Pathway Biopharma Consulting, introduced the Cell Line Development track by providing an overview of CHO cell lineage, and the transformative potential of genomics and synthetic biology in optimizing host cell lines.
Alison Young, a Staff Scientist within the Mammalian Cell Culture group at FUJIFILM Diosynth Biotechnologies, spoke further on the topic of host cell line optimization, explaining the steps to obtain a host with improved biomanufacturing attributes using directed evolution strategies. Alison also presented a cell line development approach using Sphere Fluidics’ Cyto-Mine® to combat cellular heterogeneity. This process involves optimizing the input cell concentration by diluting transfected pools to a low density before encapsulating single cells into picodroplets. Then a fluorescent assay is carried out to select picodroplets that contain a high-expressing single cell these are then imaged and individually dispensed into different wells of a 96-well plate. The imaging functionality of Cyto-Mine® quickly identifies if a picodroplet contains one cell or two and provides documented evidence to prove that a specific cell line is derived from a single cell progenitor.
A summary of how Cyto-Mine® accelerates Cell Line Development
3. Application of Cyto-Mine® to Genome Editing
New product developments and emerging scientific research were explored in the closing talk by Sphere Fluidics’Group Leader of Cell and Molecular Biology, Xin Liu. Building upon the earlier examples of using Cyto-Mine® as a screening tool in antibody discovery and cell line development, Xin demonstrated the application of Cyto-Mine® as a tool for both cell engineering and screening in genome editing. In a genome editing experiment in Cyto-Mine®, scientists co-encapsulated an HCT116 cell line (Horizon Discovery, UK) together with a viral vector, which contained Cas9 eGFP, and a construct to tag the protein, actin with RFP. Successful cell transduction was highlighted as green fluorescent protein expression. After a 16-hour incubation, using Cyto-Mine®, cells were sorted and selected based on the expression of green fluorescence and single cells were dispensed into multi-well plates. One day after the single cell was dispensed, the green fluorescence inside the cells can still be seen, confirming the expression of Cas9 eGFP, after six days and 13 days culture, the single cells had grown to form monoclonal colonies. At day 13 in culture, the colonies were investigated to identify the genome edits. Successful genome-editing events were confirmed by the presence of red fluorescence indicating that the RFP tagged actin protein was expressed.
An example of genome editing in Cyto-Mine®
Heading into the evening, the attendees had the opportunity to discuss their single cell research and receive expert advice from the Sphere Fluidics’ science team. Overall, the event provided excellent insight into the beginnings of the community, fostering open discussions and knowledge sharing to accelerate progress in biopharmaceutical discovery.