High-Throughput Determination of Infectious Virus Titers by Kinetic Measurement of Infection-Induced Changes in Cell Morphology.
Int J Mol Sci
; 25(15)2024 Jul 24.
Article
in En
| MEDLINE
| ID: mdl-39125646
ABSTRACT
Infectivity assays are the key analytical technology for the development and manufacturing of virus-based therapeutics. Here, we introduce a novel assay format that utilizes label-free bright-field images to determine the kinetics of infection-dependent changes in cell morphology. In particular, cell rounding is directly proportional to the amount of infectious virus applied, enabling rapid determination of viral titers in relation to a standard curve. Our kinetic infectious virus titer (KIT) assay is stability-indicating and, due to its sensitive readout method, provides results within 24 h post-infection. Compared to traditional infectivity assays, which depend on a single readout of an infection endpoint, cumulated analysis of kinetic data by a fit model results in precise results (CV < 20%) based on only three wells per sample. This approach allows for a high throughput with ~400 samples processed by a single operator per week. We demonstrate the applicability of the KIT assay for the genetically engineered oncolytic VSV-GP, Newcastle disease virus (NDV), and parapoxvirus ovis (ORFV), but it can potentially be extended to a wide range of viruses that induce morphological changes upon infection. The versatility of this assay, combined with its independence from specific instruments or software, makes it a promising solution to overcome the analytical bottleneck in infectivity assays within the pharmaceutical industry and as a routine method in academic research.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Viral Load
Limits:
Animals
/
Humans
Language:
En
Journal:
Int J Mol Sci
Year:
2024
Document type:
Article
Affiliation country:
Germany
Country of publication:
Switzerland