Multidrop combi peristaltic bulk dispenser

Prior to cell plating, 1,536 well plates are filled with 4 µL of neuronal culture medium consisting of Neurobasal-E, B-27, L-Glutamine, pen/strep (all Life Technologies) using a Multidrop Combi peristaltic bulk dispenser (ThermoFisher). Each plate is then “pinned” with 23 nL of a compound at a set stock concentration in DMSO that is delivered from a library plate using a robotic pintool transfer tool (Wako). Cells are resuspended at a concentration of 2.5 × 10⁵ cells/mL, filtered through a 10 µm nylon filter (Amazon.com: Small Parts), dispensed at a final well culture volume of 8 µL (1,000 cells/well), and incubated for up to 21 days. Since we are using such low volumes for extended cultures, the plates are covered with a Breathe-Easy gas-permeable membrane (Diversified Biotech) and a Microclime vapor barrier lid (Labcyte).

1536 well plates were filled with 4 µL of neuronal culture medium consisting of Neurobasal-E, 2% B-27, 0.5 mM L-Glutamine, and 1X final penicillin/streptomycin (all Life Technologies) using a Multidrop Combi peristaltic bulk dispenser (ThermoFisher). 23 nL of a stock compound solution 10 mM-170 nM in DMSO was transferred to each well in the culture plate from a library plate using a robotic pintool transfer tool (Wako) resulting in final concentrations ranging from 25 µM-420 pM. Cells were resuspended at a concentration of 1.25 × 10⁵ cells/mL, filtered, then dispensed into plates at a final well culture volume of 8 µL (1000 cells/well). Plate were incubated for 7days in 95% humidity at 37°C.