# Influence of Insert Brand and Culture Method on Ciliary Activity and Epithelial Cell Types in Human Nasal Air–Liquid Interface Cell Cultures

## TL;DR

This study compares how different cell culture inserts and culture methods affect nasal cell cultures, finding that insert brand does not significantly influence results.

## Contribution

The study provides a direct comparison of three insert brands and an alternative culture setup for nasal epithelial cell cultures.

## Key findings

- Ciliary activity and cell type composition were similar across all three insert brands.
- The inverted culture setup showed higher ciliary beat frequency and lower active ciliated area compared to the conventional setup.
- All three insert types produced comparable cell cultures regardless of culture method.

## Abstract

Cultures of primary human nasal epithelial cells (hNECs) differentiated at the air–liquid interface (ALI) represent a sophisticated and widely used model of the human upper respiratory epithelium. Despite the availability of various cell culture insert types and the well-established understanding that different culture media influence the cell culture characteristics, the possible impact of the insert brand remains rather underexplored. We cultured hNECs from nineteen healthy adult donors on three distinct brands of commercially available inserts—Corning® Transwell®, CELLTREAT®, and ThinCert®—and compared the ciliary activity and cellular composition of the cultures using high-speed video microscopy and flow cytometry, respectively. Additionally, we employed an alternative method of hNEC culture setup—the inverted condition—wherein the hNECs were seeded on the basal side of the insert with the idea to avoid mucus accumulation. Our results show that ciliary activity and cell type composition did not differ between insert types for both culture conditions. However, we found a higher ciliary beat frequency and a lower active (ciliated) area in the inverted setup compared to the conventional setup across all three insert brands. These findings indicate that all three mentioned insert types yield comparable cell cultures.