WinterVomitingLab:Protocols/In-solution inactivation test

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Overview

• Virus inactivation using a liquid sanitizer solution.

Materials

• Sanitizer(s)
• Neut/elute buffer
• Neutralization buffer
• Centrifuge bottles

Procedure

1. Prepare neutralization tubes containing 900 μl of desired neutralization buffer for each sample.
2. Prepare sanitizer solutions.
3. For samples-
   1. Add 100 μl of prepared virus stock to sample tube and incubate for 1 minute (or appropriate time) while inverting the tube.
   2. Remove 100 μl from tube and add to the tube containing neutralization buffer.
   3. Store samples at -70°C until further processing.
4. For neutralization controls-
   1. Add 100 μl of sanitizer to the tube containing neutralization buffer and incubate for 1 minute while inverting the tube.
   2. Remove 10 μl from tube, and add 10 μl of prepared virus stock.
   3. Store samples at -70°C until further processing.
5. For cytotoxicity controls-
   1. For 10⁰ control, add 100 μl of sanitizer to 900 μl of neut/elute buffer and incubate for 1 minute while inverting tube.
   2. For 10^-1 control, remove 100 μl from 10⁰ control and add to 900 μl PBS.
   3. Store samples at -70°C until further processing.
6. For negative controls-
   1. Add 100 μl of sanitizer to the tube containing neutralization buffer and incubate for 1 minute while inverting tube.
   2. Store samples at -70°C until further processing.
7. For positive controls-
   1. Add 100 μl of prepared virus stock to 900 μl PBS and incubate for 1 minute while inverting tube.
   2. Store samples at -70°C until further processing.
   3. Perform a serial dilution of the virus stock just before plaque assay.
8. Plaque assay controls (performed with each plaque assay to aid in troubleshooting if questionable results are obtained)-
   1. Negative control with agar
   2. Negative control without agar (liquid culture)
   3. Positive control (-2 dilution of virus stock) with agar
   4. Positive control (-2 dilution of virus stock ) without agar (liquid culture)

References

Relevant papers and books

Contact

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