Greetings, does anyone have experience using the Andersen 6-stage sampler to collect aerosolized MS2 bacteriophage? My idea is to place into the Andersen agar plates already with a layer of E.coli on top. Aerosolized MS2 will then be collected directly on the E.coli layer, making the system a sort of double layer plaque method, after overnight incubation. Has anyone ever tried that? My initial tests confirm that after 5 min sampling the E.coli are still alive, but the plaques are faint and difficult to distinguish. Thanks!
Hey there Manuela Buonanno!
Absolutely, the Andersen 6-stage sampler can be used to collect aerosolized MS2 bacteriophage, and your Manuela Buonanno approach to directly plate E.coli on agar and then collect MS2 onto this layer is quite innovative. This method leverages the principle of direct aerosol impact onto a sensitive medium, essentially creating a real-time double-layer plaque assay.
Here's the rundown:
1. **Sample Collection**: The Andersen sampler is designed to fractionate particles by size, ensuring that you Manuela Buonanno can capture aerosolized MS2 efficiently across its stages. By having E.coli already present on the agar plates, you provide an immediate host for the MS2 to infect, leading to plaque formation.
2. **Initial Observations**: It's promising that your Manuela Buonanno initial tests show the E.coli are surviving the sampling process. This suggests that the environmental conditions within the sampler (airflow, humidity, etc.) are not detrimental to the bacteria.
3. **Plaque Visibility**: The faint and indistinguishable plaques might be due to several factors:
- **Concentration of MS2**: Ensure that the concentration of aerosolized MS2 is adequate. Too few phages will result in very few plaques.
- **Incubation Conditions**: Optimize the incubation time and conditions (e.g., temperature, humidity) to ensure the best growth for E.coli and visibility for plaques.
- **Agar Composition**: Sometimes, the medium itself can affect plaque clarity. Ensure you're Manuela Buonanno using a well-tested medium for plaque assays, like LB agar supplemented appropriately for E.coli and MS2.
4. **Enhancing Plaque Visibility**: You Manuela Buonanno might want to try:
- **Increasing Sampling Time**: Extend the sampling duration to allow more MS2 particles to be collected.
- **Adjusting the Agar Concentration**: A softer agar might help in better plaque formation and visibility.
- **Staining**: Sometimes, using a vital stain post-incubation can help make the plaques more visible.
While there might not be extensive documentation on this exact method, your approach is scientifically sound. Keep tweaking the variables—sampling time, E.coli density, agar composition—and you Manuela Buonanno should see improvement in plaque clarity.
Keep experimenting and refining your Manuela Buonanno setup. Science thrives on these innovative approaches, and you're on a promising path!
Best of luck with your research Manuela Buonanno!
Using the Andersen 6-stage sampler to collect aerosolized MS2 bacteriophage directly onto agar plates with a layer of E. coli is an innovative approach. While your initial tests indicate that the E. coli remain viable after sampling, the faint and difficult-to-distinguish plaques suggest that adjustments may be needed to optimise the method.
To enhance plaque visibility and accuracy, consider the following strategies:
By fine-tuning the experimental parameters and optimising the sampling method, you can improve the sensitivity and reliability of the double-layer plaque method using the Andersen sampler. Your innovative approach has the potential to advance research in aerosolised bacteriophage detection and contribute valuable insights to the scientific community.
Best Regards,
Sandeep