Establishing New Standards in Efficient Single Cell Cloning of hiPSCs

Life Sciences, Laboratory Technology,
  • Tuesday, September 22, 2020

Drug discovery, diagnostic, and therapeutic efforts are increasingly using human-induced pluripotent stem cells (hiPSCs) and related technologies. However, the inability to robustly manipulate hiPSCs as single cells has remained a significant biological and technical hurdle. Current techniques rely on inefficient methods, such as limiting dilution (LD), which are time consuming, expensive, incompatible with the sensitivity of hiPSCs, and ultimately, ineffective in addressing concerns about clonality. For example, hiPSC gene-editing efficiencies can be low, thus requiring the generation and screening of hundreds of clones, which in turn must be validated for clonal origins.

Solentim has previously established the benefits of its proven VIPS™ commercial single cell seeding platform over manual LD for the creation of master cell banks, which helped shape industry standards for clonality of therapeutic CHO and HEK cell lines. Until now, the technology has been incompatible with hiPSCs due to their inherent sensitivity and particular culture needs, such as specialized pre-plated matrices and daily feeding schedules. Here we present a robust and clinically relevant workflow for the single-cell subcloning of hiPSCs using the VIPS seeding platform in combination with a new soluble dispensing matrix, MatriClone™(Solentim). MatriClone is an animal component-free matrix dispensed at the time of seeding, which allows for cell attachment and growth without the need for pre-coating culture plates. The VIPS instrument is uniquely able to perform the dual functions of high efficiency single-cell seeding and whole-well imaging to document outgrowth and verify clonal origin. The combination of this instrument and dispensing matrix provides a novel solution for improving workflows for cell reprogramming or gene editing of hiPSCs.

In this webinar, we demonstrate that numerous hiPSC cell lines can be successfully subcloned in a robust and automated fashion using the VIPS instrument in conjunction with optimized culture conditions, thereby reducing time and cost while most importantly maintaining the integrity of clonal biology. Firstly, we show that the VIPS plus MatriClone combination results in a several-fold improvement in clonal colony outgrowth of single seeded hiPSCs when compared with manual LD. Secondly, a number of hiPSC subclones were identified from presumed healthy and disease-affected backgrounds using daily whole well imaging on the VIPS and selected for expansion approximately 10-14 days post-seeding. Expression of pluripotency was confirmed for each of the sub-cloned lines. Finally, we demonstrate that subclones that maintained genomic integrity after extended culture and successfully differentiated into mixed cortical neurons.

The robustness of this workflow has immediate potential to impact standards, consistency, and confidence of clonality in both academic and pharma research. Furthermore, future translation of this workflow could positively effect GMP standards and re-establish expectations with the regulatory bodies for the development and production of advanced cell models, cellular diagnostics, and cell therapeutics, including clonality documentation to accompany future IND submissions.

Speakers

http://Dr.%20Philip%20Manos

Philip Manos, Founder, President & Scientific Director, EverCell Bio, Inc.

Dr. Manos is a stem cell expert with a successful history of technology development and business operations within both academic and commercial organizations. Since entering the human induced pluripotent stem cell (hiPSC) field in its infancy in 2008, he has developed a strong publication record focused on hiPSC technology, including reprogramming via modified mRNA for derivation of the first clinically relevant hiPSCs. Dr. Manos began his career at the the Harvard Stem Cell Institute hESC Core facility. He then went on to help establish reprogramming and hiPSC disease modeling platforms at both the Novartis Institute for Biomedical Research and the Neural Stem Cell Institute before serving as the Director of Operations for StemCultures. Using his vast range of experience centered on hiPSC technology translation and business development, Dr. Manos founded EverCell Bio in 2019 to establish a new standard of personalized stem cell technologies and services.

Message Presenter
http://Dr.%20Ian%20Taylor

Dr. Ian Taylor, Chief Business Officer, Solentim Limited

Dr. Taylor has a PhD in Biochemistry. He has more than 25 years working in the conceptualization and commercialization of novel high value life-science instrumentation. Prior to joining Solentim when it was founded ten years ago, Dr. Taylor was Commercial Director at Genetix PLC where he was responsible for the development and sales of the ClonePix FL for selection and isolation of high value monoclonal antibody-producing clones. Prior to that, Dr. Taylor was part of the senior leadership team for PerkinElmer Life Sciences.

Message Presenter

Who Should Attend?

  • Stem cell cores
  • CDMOs for iPSCs
  • Disease model groups in Pharma R&D
  • Groups involved in gene editing or reprogramming for iPSC
  • Therapeutic iPSC
  • Cell therapy groups

What You Will Learn

  1. High efficiency single cell isolation of iPSCs
  2. High viability for colony outgrowth
  3. Combination with MatriClone substrate
  4. Full documentation of clonality

Xtalks Partner

Solentim

Solentim is the trusted global leader for solutions to create, isolate, and characterize the highest value cells for the development of new biological medicines, including cell and gene therapies, monoclonal antibodies, vaccines and stem cells. For our customers, which include most of the world’s leading biotherapeutics companies, we are the partner of choice for facilitating the highest level of assurance in cell line development for therapeutic protein and viral vector production. Our portfolio of proven and innovative technologies, combined with our unparalleled experience, ensures our customers achieve accelerated Master Cell Bank development and are confidently prepared for regulatory review.

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