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Rotavirus concentration from cell culture harvests: trypsin treatment followed by hydroextraction
Abstract:
An 80- to 150-ml amount of calf or simian rotavirus-containing cell culture harvests of MA-104 cells were treated with 50 microgram of trypsin per ml and hydroextracted overnight (4 degrees C) with polyethylene glycol 6,000. The concentrate was resuspended in 8 to 10 ml of tryptose phosphate broth and plaque assayed. Between 85 and 97% of the input virus could be recovered with a concentration of up to 15-fold.
Insights
This study details a method to concentrate rotavirus from cell cultures using trypsin and polyethylene glycol, achieving high recovery rates for improved viral quantification. This technique enhances rotavirus (RV) detection in research settings.
Area of Science:
- Virology
- Cell Biology
- Biotechnology
Background:
- Rotavirus (RV) is a leading cause of severe diarrheal disease in infants worldwide.
- Accurate quantification of infectious rotavirus particles is crucial for vaccine development and epidemiological studies.
- Current concentration methods can be labor-intensive and may lead to significant viral loss.
Purpose of the Study:
- To develop and optimize a simple, efficient method for concentrating rotavirus from cell culture harvests.
- To evaluate the recovery rate and concentration factor of the optimized method.
- To facilitate improved rotavirus detection and analysis in laboratory settings.
Main Methods:
- Cell culture harvests containing calf or simian rotavirus (RV) were treated with trypsin.
- Virus was concentrated using overnight hydroextraction with polyethylene glycol (PEG) 6,000 at 4°C.
- The concentrated virus was resuspended and plaque assayed for quantification.
Main Results:
- The method successfully concentrated rotavirus from MA-104 cell cultures.
- Recovery rates of input virus ranged from 85% to 97%.
- A concentration factor of up to 15-fold was achieved.
Conclusions:
- The described method provides an effective means to concentrate rotavirus from large-volume cell cultures.
- This technique offers high viral recovery and significant concentration, aiding in sensitive rotavirus detection.
- The optimized protocol is suitable for routine laboratory use in rotavirus research.