The baculovirus expression vector system (BEVS) is a powerful tool to produce recombinant protein on both laboratory and industrial scales for various applications such as production of vaccines and pharmaceutical proteins. It displays several advantages over other expression systems, including the capacity for high expression levels, large DNA insertion, and post-translational modifications of numerous proteins similar to those in mammalian cells. To improve the efficiency of protein production, optimization of a series of parameters is required. Among them, the ratio of input virus to the number of cells in the culture, also called the multiplicity of infection (MOI), is one of the most important factors. Therefore, accurate measurement of virus titer before protein production is much essential.
Common methods used to calculate baculovirus titer contain plaque assay and end point dilution. However, both methods are significantly affected by cell types, period of virus incubation and many other factors. Besides, the accuracy of the titer is greatly dependent on the operator’s skill and experience. These bioassays are also time-consuming, often requiring 4–7 days for the determination of the titer. Here, QVirusTM Platform has developed a simple and rapid baculovirus titer assay method. A high throughput, flow cytometric assay is utilized to quantitate infectious baculovirus particles produced in insect cell culture, allowing for accurate virus titer determination for multiple conditions in less than 24 hours.
The titer (infectious units (IU) per milliliter) of a baculovirus stock is determined by measuring the expression of surface glycoprotein 64 (gp64, a baculovirus envelope protein) on the surface of infected insect cells eight hours post infection (hpi). The assay detects the cell surface expression of gp64 after a short incubation period allowing for a single round of infection. Then, virus infected cells are assessed by flow cytometric analysis for gp64 expression. Our method measures the number of infected cells, in triplicate, over a series of nine dilutions of the baculovirus stock ranging from 1:2 to 1:100,000. The assay is formatted for 96 well plates and requires a minimum volume of baculovirus stock.
QVirusTM Platform is pleased to offer baculovirus titer assay services to help our customers maximize expression and reproducibility. If you have any special requirements, please feel free to contact us.
1. Qi J, et al. Rapid baculovirus titration assay based on viable cell side scatter (SSC). Analytica Chimica Acta, 2015, 879:58-62.