A continuous perfusion microplate for cell culture.

Area of Science:

• Biotechnology
• Cell Biology
• Microfluidics

Background:

• Traditional cell culture methods often lack the dynamic microenvironment found in vivo.
• Continuous fluid perfusion is crucial for mimicking physiological conditions but typically requires complex external pumping systems.

Purpose of the Study:

• To develop and validate a novel perfusion microplate system for continuous fluid exchange between wells without external pumps.
• To assess the utility of this microplate for various cell culture applications, including co-culture and toxicity studies.

Main Methods:

• A 96-well microplate design with fluidically connected source, sample (cell culture), and waste wells was developed.
• Fluid perfusion was driven by hydrostatic pressure and capillary action through cellulose membrane wicks.
• Cell viability assays were performed using hepatocytes (C3A cells) and EOC 20 cells in co-culture with LADMAC cells.
• Drug metabolism and toxicity studies were conducted using primary human hepatocytes and colon cancer cells (HCT 116).

Main Results:

• The perfusion microplate demonstrated effective continuous fluid perfusion driven by hydrostatic pressure and wicking.
• Hepatocyte viability in the microplate was comparable to traditional daily media exchange.
• Co-cultured cells (EOC 20 and LADMAC) showed equal viability, indicating successful conditioned media transfer.
• A prodrug (Tegafur) metabolized by hepatocytes in the source well induced toxicity in adjacent colon cancer cells.

Conclusions:

• The perfusion microplate is a valuable tool for cell culture, simplifying media exchange and reducing labor.
• It effectively supports co-cultures and enables in vivo-like studies of drug metabolism and toxicity.