# Cryopreserved Kidney Epithelial (Vero) Cell Monolayers for Rapid Viral Quantification, Enabled by a Combination of Macromolecular Cryoprotectants

## TL;DR

This paper introduces a new method using frozen Vero cells to quickly measure virus levels, reducing the time needed for testing.

## Contribution

The novel use of macromolecular cryoprotectants and cryopreserved Vero cell monolayers enables rapid viral quantification.

## Key findings

- Cryopreserved Vero cells with polyampholytes achieved high post-thaw recovery and viability.
- Influenza viral plaques were detected in 24 hours using cryopreserved cells.
- Chemical ice nucleation reduced variability between wells and improved consistency.

## Abstract

Plaque assays quantify the amount of active, replicating
virus
to study and detect infectious diseases by application of samples
to monolayers of cultured cells. Due to the time taken in thawing,
propagating, plating, counting, and then conducting the assay, the
process can take over a week to gather data. Here, we introduce assay-ready
cryopreserved Vero monolayers in multiwell plates, which can be used
directly from the freezer with no cell culture to accelerate the process
of plaque determination. Standard dimethyl sulfoxide cryopreservation
resulted in just 25% recovery, but addition of polyampholytes (macromolecular
cryoprotectants) increased post-thaw recovery and viability in 12-
and 24-well plate formats. Variability between individual wells was
reduced by chemically induced ice nucleation to prevent supercooling.
Cryopreserved cells were used to determine influenza viral plaques
in just 24 h, matching results from nonfrozen controls. This innovation
may accelerate viral detection and quantification and facilitate automation
by eliminating extensive cell culturing.