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

  1. []

Contact

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