3D cellular models have increased in significance as a tool of cell biology as cells are allowed to grow in vitro in the way they would in vivo.
Spheroids: a 3D Cellular Environment to Screen Compounds
We use Ultra Low Attachment (ULA) plates to generate and culture spheroids. These plates have an ultra-low attachment surface coating and a well geometry that favors the formation of a single spheroid within each well. Spheroids of different sizes develop gradients of oxygen, nutrients and metabolites, creating a hypoxic core and replicating cells on the outer edges.
Drug potency and efficacy is monitored in either an imaging-based assay (on a wide-field imaging plate reader) or by using commercially available assays that track parameters such as cell viability.
Advantages of 3D spheroids are:
- Retained 3D architecture of cellular environment in culture along with associated physiological functions
- Uniformity in size and shape make them desirable for high throughput screening formats in drug discovery projects
- Generate highly reproducible results
- Increased viability of cells that are otherwise difficult to culture, including primary cells
- Ability of spheroids to replicate the hypoxic core during tumor growth make them reliable models for cancer drug discovery projects
- Allows for the investigation of cellular changes such as cell toxicity or cell viability in real-time
Our track record
We have grown numerous cells in a 3D format:
- MCF7 breast cancer cell loose spheroid
- HEK embryonic kidney loose spheroid
- U2OS osteosarcoma loose spheroid
- HCT 116 colon tight spheroid
- U87 glioblastoma tight spheroid
- HT-29 colon tight spheroid
We treated the spheroids with various drug treatments and have monitored their response using Draq 7 (which only penetrates dead cells), Calcein AM (which only penetrates live cells) and Hoeshct for an overall cell count. We have also used to CellTire Glo to determine viability by looking at ATP levels.
Our bioscience division developed a 3D micro-tissue system from electrospun material that can be used in conjunction with well plates for higher throughput screening. Electrospun material mimics the natural extracellular matrix and provides an ideal substrate for cells to adhere to.
We re-engineered electrospun material to form micro-scaffold islands on to which we seed, grow and differentiate cells prior to performing more conventional assays in well plates. Cells grow on, around and into the material, forming a micro-island of adherent cells that are effectively “micro-tissues in solution”.
The incorporation of iron particles into fibers during manufacture results in scaffolds that can be physically manipulated using magnetism.
Advantages of 3D electrospun scaffolds are:
- Robust and reproducible 3D culture environment
- Applicable to any cell type (recombinant, human primary cells and iPSCs plus differentiation)
- Movable: from vessels-to-well and well-to-well with magnetism
- Scalable to any assay throughput
- Integrates seamlessly with all current screening and assay workflows
Our Track Record
- Proteins were isolated from cells on scaffolds, run in Western Blotting and probed with antibodies to Pax-6, Ctip-2, and beta tubulin. Shown are Western blots of proteins isolated 14 day post-culture in differentiation media showing expression of mature cortical neurone proteins
- Optical Coherent Tomography was used to examine the growth of cells in scaffold material. HCT116, A549, U87 an U937 cells were seeded into scaffold material and allowed to grow for 24, 48 and 72 hours post seeding. Shown here for HCT116 cells, after 24hrs, the cells have infiltrated the material and by 72 hrs, the material is completely engulfed by the cells
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