Viral Growth in Human Nasal Epithelial Cells Cultured at an Air-Liquid Interface

Begin with human nasal epithelial cells covered with mucus, cultured on a transwell insert in a multi-well plate.

The basal side of the culture contacts medium while the apical surface is exposed to air, simulating the air-liquid interface of the nasal cavity.

Wash the culture with buffer to remove the mucus layer.

Add coronavirus, suspended in serum-free medium, to the culture.

Incubate the plate with occasional rocking to ensure viral adsorption.

Remove the viral inoculum and wash with buffer to remove unbound viruses.

Remove the insert and replenish the well with fresh medium.

Incubate the plate to allow viral infection.

The coronavirus binds to the cell-surface receptor, enters by endocytosis, and releases its genome into the cytoplasm.

The viral genome replicates and produces proteins, which assemble into new viruses.

New viruses are released apically, spread to infect nearby cells, and amplify viral growth.

Grow and differentiate the nasal air-liquid interface, or ALI cultures, in transwells having a basal and an apical chamber. Expand the dissociated human sinonasal cells under submerged condition, following which, remove the media from the apical chamber to allow differentiation of th