Scientists using cell culture for drug discovery, toxicology, stem cell biology, and basic research realize the critical importance of 3-dimensional (3D) models. Data from cells cultured in a non-physiologic, monolayer format on plastic surfaces has long been suspected to differ from true in vivo physiology, and evidence supporting how this difference is slowing the pace of scientific discovery is mounting.

In the past, there were few affordable, reliable choices for 3D culture and almost none that were amenable to high throughput screening. Spheroids, self-assembled microscale cell aggregates, are superior models of avascular in vivo microtumors. Using hanging drop technology, a drop of cell suspension is pipetted into the top of each well.
The cell suspension forms a stable drop below where the cells aggregate into spheroids.

Two critical steps must be accomplished to facilitate spheroid formation for 3D culture in the HDPs. These include dispensing cells and medium, and tracking spheroid formation. Dispensing into the plates can be manual or automated. Automated dispensing is faster than manual pipetting, which is the preferred method, especially with the 384-well plate. Cell imaging in the hanging drop is possible with a 4x or 10x
objective, or a long working distance 20x objective.

Here we demonstrate how dispensing and imaging procedures were accomplished to facilitate higher throughput in 3D HDP processing. Automating each of these processes provides a 3D solution that is less labor intensive and more reproducible than previous methods, and further promotes the use of this spheroid formation method.


Brad Larson1, Nicky Slawny2, and Peter Banks1
1BioTek Instruments, Inc., Winooski, Vermont, USA ? 23D Biomatrix, Inc., Ann Arbor,
Michigan, USA