(1273-B) Discovering novel biologics via automated high-throughput flow cytometry

Abstract: A crucial step in the identification of therapeutic proteins is to assess the binding of candidates to their targets in a cellular context, providing target engagement information on native epitopes and in physiologically relevant conditions. Manual approaches to plate-based flow cytometry screens are time-consuming, labor-intensive and lack robust reproducibility. To accelerate our discovery process, we developed a novel method for automated 96-well antibody screening on a Hamilton Microlab STARlet liquid handler. Our Hamilton method utilizes on-deck, fully-automated buffer transfers, mixes, and primary & secondary antibody additions, and the Cytkick Max upgrade to our Attune flow cytometer features higher sample flow rates and HT optimized wash and mix settings. These factors greatly reduce scientist interaction time from approximately 4 hours to 2 hours, and total assay time from 8 hours to 6 hours. This transforms our previous workflow into a high-throughput flow cytometry screening experiment manageable within an 8-hour workday, removing previous scheduling constraints from the planning process, and reducing the time needed for larger experiments. Building on this process, we are also integrating an on-deck centrifuge and chilled plate holders which will further reduce scientist hands on time to approximately 1 hour, within a 5 to 6 hour total process. This new process will require intervention only for cell preparation and loading onto a plate hotel located near the flow cytometer. We are also transferring this process to a larger Hamilton STARplus deck, allowing us to generate 8 flow cytometry-ready plates per run, compared to our current maximum of 4 plates per run, in the same 6-hour period. By allowing larger sample sets to be run faster and more frequently, our existing scientists’ bandwidth can manage 1.5 to 2 times higher throughput per day, multiplied by approximately 2 times higher assay frequency per week. This impacts project timelines by reducing turnaround time for large sample sets from 1 month to 2 or 3 weeks, reduces the need for follow-ups and reruns by allowing for richer data (more replicates or more conditions of interest) to be captured on the first pass, and widens this leg of the pipeline workflow by making it more feasible for flow cytometry sample sets from multiple projects to be run in the same week, by the same scientist. While our team intends to further integrate with a STARplus, on-deck cooling system, Agilent V-spin, and hoteling system on the Attune flow cytometer, this workflow concept can remain highly modular and adaptable, and ultimately be applied to any lab with sufficient liquid handler deck space, a benchtop centrifuge, and a flow cytometer with 96-well plate capabilities.