Cell migration / Invasion cell type - A549

Cell Invasion or Cell Migration assays are technically challenging to set up as the gradient between the two compartments equilibrates in time during the assay. It is also problematic to view cells and for cells to migrate through a non-physiologic polycarbonate or polypropylene filter. Care must be taken while loading the well with cells to form a single cell suspension. Precaution must be taken while trypsinization (under-trypsinization can lead to cell clumping while over-trypsinization could strip off adhesion molecules necessary for migration). This leads to difficulty in getting significant results, when only small numbers of cells cross the filter or when the distribution and/or staining of the cells is uneven.

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Found 3 matching solutions for this experiment

Upstream tips
Store all components at 4ºC.
Protocol tips
Incubate for 24 hours in a cell culture incubator at 37C.

Retain the medium in the 24-well migration plate that contains chemoattractant(s) and cells that migrated through the membrane and into the medium
Downstream tips
Read fluorescence with a fluorescence plate reader at 480 nm/520 nm.
Upstream tips
Do not freeze.
Protocol tips
Incubate 48 hours in a culture incubator.

Stain invasive cells on lower surface of the membrane by dipping inserts in the staining solution for 20 minutes.
Upstream tips
Coat with 100 µl basement membrane extract coating solution at 37°C for 4 h.
Protocol tips
Incubate at 37C in a CO2 incubator for 24 hours
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