48-Well Micro Chemotaxis Chamber

Protocol tips

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	Incubate in humidified atmosphere and 5% CO2 at 37°C for 4 hours

Protocol tips
Incubate in humidified atmosphere and 5% CO2 at 37°C for 4 hours

Publication protocol

HT-1080 cell migration was measured using a 48-well micro chemotaxis chamber (Neuro Probe, Gaithersburg, MD, USA) equipped with a 8 µm pore-size polycarbonate membrane filter (Whatman, GE Healthcare Bio-Sciences Corp., Florham Park, NJ, USA). Fifty µL of cell suspension (1×106 cells/mL) in culture medium containing 10% FBS was placed into the upper chamber. Extracellular matrix (ECM) gel from Engelbreth-Holm-Swarm murine sarcoma (Sigma-Aldrich) diluted in serum-free medium at 0.05 mg/mL was placed in the bottom chamber as chemoattractant, except for the blank sample where it was omitted. After a 4-hour-long incubation in humidified atmosphere and 5% CO2 at 37°C, the chamber was disassembled and cells from the upper face of the filter were dislodged. The membrane was dried, methanol-fixed and stained with Toluidine Blue (Sigma-Aldrich). The amount of cells migrated through the membrane was quantified by densitometry using the Kodak Image Station 4000MM (Carestream Health, Inc., Rochester, NY, USA) and software provided by the manufacturer.

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