Microassay for interferon, using [3H]uridine, microculture plates, and a multiple automated sample harvester

Area of Science:

• Virology
• Immunology
• Biochemistry

Background:

• Interferon (IFN) is crucial for antiviral defense.
• Accurate and efficient assays are needed to quantify IFN activity.
• Existing methods can be time-consuming or require large sample volumes.

Purpose of the Study:

• To develop and validate a microassay for quantifying interferon activity.
• To assess the reliability and scalability of the developed microassay.
• To compare the microassay results with a standard plaque reduction assay.

Main Methods:

• Utilized microculture wells with target cells.
• [3H]uridine incorporation measured vesicular stomatitis virus (VSV) replication.
• Automated sample harvester collected radioactively labeled viral RNA.
• Liquid scintillation spectrophotometry quantified radioactivity.

Main Results:

• Interferon titers determined by the microassay correlated well with plaque reduction assays at low VSV concentrations (100 PFU).
• A 2-fold decrease in IFN titer was observed for every 10-fold increase in VSV challenge concentration (range 10^2 to 10^5 PFU).
• The microassay demonstrated high repeatability and suitability for processing numerous samples.

Conclusions:

• The developed microassay provides a reliable and repeatable method for quantifying interferon activity.
• The assay is adaptable for high-throughput screening of interferon samples.
• Care must be taken when interpreting results with varying challenge virus concentrations.