SEA-PHAGES | Syringe filter failure?

Link to this post \| posted 24 Sep, 2021 19:35
cheryl.brown@mnstate.edu	We have been plagued with recurrent contamination problems on plates. One of the confusing factors was the fact that the populations of contaminants was varied. We have widespread, but inconsistent, contamination. I just ran a test yesterday, following the student protocols. I soaked the soil, ran it through a coffee filter, and then through a syringe filter. One sample was more plant material, so the broth needed no force to go through. The other sample was a fine, silty soil, and considerable pressure was needed to get it filtered. There was no obvious break or breach, as the same pressure was needed the entire time. I then plated the filtrate with sterile top agar and no culture. The "forced" plate was covered with bacteria. Is there a change in protocol needed? Additional pre-filtering?

Link to this post | posted 24 Sep, 2021 19:35

We have been plagued with recurrent contamination problems on plates. One of the confusing factors was the fact that the populations of contaminants was varied. We have widespread, but inconsistent, contamination.

I just ran a test yesterday, following the student protocols. I soaked the soil, ran it through a coffee filter, and then through a syringe filter. One sample was more plant material, so the broth needed no force to go through. The other sample was a fine, silty soil, and considerable pressure was needed to get it filtered. There was no obvious break or breach, as the same pressure was needed the entire time. I then plated the filtrate with sterile top agar and no culture.

The "forced" plate was covered with bacteria.

Is there a change in protocol needed? Additional pre-filtering?

Link to this post \| posted 26 Sep, 2021 19:51
viknesh	cheryl.brown@mnstate.edu We have been plagued with recurrent contamination problems on plates. One of the confusing factors was the fact that the populations of contaminants was varied. We have widespread, but inconsistent, contamination. I just ran a test yesterday, following the student protocols. I soaked the soil, ran it through a coffee filter, and then through a syringe filter. One sample was more plant material, so the broth needed no force to go through. The other sample was a fine, silty soil, and considerable pressure was needed to get it filtered. There was no obvious break or breach, as the same pressure was needed the entire time. I then plated the filtrate with sterile top agar and no culture. The "forced" plate was covered with bacteria. Is there a change in protocol needed? Additional pre-filtering? Hi Cheryl, These filters are not designed to work under pressure. As soon as it becomes difficult to push liquid through the filter, the filter pores are essentially clogged/blocked and you've reached the filtration capacity of that filter. Even with steady force, you risk micro-ruptures and eventually a full rupture of the membrane. For clay or silty samples, which always clog the filters, I spin the soil suspension (in 50 ml conical tubes) at 4 krpm 10 minutes. This is usually fast enough to pellet much of the particulate matter so that I can avoid clogging the filters too quickly. Once it clogs, I either use a new filter or just enrich with however little filtrate I am able to get. Often, it is easy to obtain at least 10 ml of filtrate before these samples (spun at 4krpm) clog the filter. I hope this is helpful. Vic

Link to this post | posted 26 Sep, 2021 19:51

viknesh

cheryl.brown@mnstate.edu
We have been plagued with recurrent contamination problems on plates. One of the confusing factors was the fact that the populations of contaminants was varied. We have widespread, but inconsistent, contamination.

I just ran a test yesterday, following the student protocols. I soaked the soil, ran it through a coffee filter, and then through a syringe filter. One sample was more plant material, so the broth needed no force to go through. The other sample was a fine, silty soil, and considerable pressure was needed to get it filtered. There was no obvious break or breach, as the same pressure was needed the entire time. I then plated the filtrate with sterile top agar and no culture.

The "forced" plate was covered with bacteria.

Is there a change in protocol needed? Additional pre-filtering?

Hi Cheryl,

These filters are not designed to work under pressure. As soon as it becomes difficult to push liquid through the filter, the filter pores are essentially clogged/blocked and you've reached the filtration capacity of that filter. Even with steady force, you risk micro-ruptures and eventually a full rupture of the membrane.

For clay or silty samples, which always clog the filters, I spin the soil suspension (in 50 ml conical tubes) at 4 krpm 10 minutes. This is usually fast enough to pellet much of the particulate matter so that I can avoid clogging the filters too quickly. Once it clogs, I either use a new filter or just enrich with however little filtrate I am able to get. Often, it is easy to obtain at least 10 ml of filtrate before these samples (spun at 4krpm) clog the filter.

I hope this is helpful.

Vic

Link to this post \| posted 27 Sep, 2021 14:50
cheryl.brown@mnstate.edu	Thank you. As a follow-up, are you aware of anyone else having difficulties with VWR filters recently? We have been using the same filters for 3 years and this fall is the first time we have had the problem. It seems like a QA failure. We ran a test with the current filters and some others on hand, and, using the same force in both, the other filters (also .22 micron syringe filters), only the VWR ones had the contamination problem. I have some .2 micron PES filters on hand. Can those be used in this application? We are also going to tweak the pre-filter/coffee filter step to further decrease particles in samples.

Link to this post | posted 27 Sep, 2021 14:50

cheryl.brown@mnstate.edu

Thank you. As a follow-up, are you aware of anyone else having difficulties with VWR filters recently? We have been using the same filters for 3 years and this fall is the first time we have had the problem. It seems like a QA failure.

We ran a test with the current filters and some others on hand, and, using the same force in both, the other filters (also .22 micron syringe filters), only the VWR ones had the contamination problem.

I have some .2 micron PES filters on hand. Can those be used in this application?

We are also going to tweak the pre-filter/coffee filter step to further decrease particles in samples.

Link to this post \| posted 27 Sep, 2021 14:56
viknesh	cheryl.brown@mnstate.edu Thank you. As a follow-up, are you aware of anyone else having difficulties with VWR filters recently? We have been using the same filters for 3 years and this fall is the first time we have had the problem. It seems like a QA failure. We ran a test with the current filters and some others on hand, and, using the same force in both, the other filters (also .22 micron syringe filters), only the VWR ones had the contamination problem. I have some .2 micron PES filters on hand. Can those be used in this application? We are also going to tweak the pre-filter/coffee filter step to further decrease particles in samples. I've personally not heard any recent complaints about VWR filters, but perhaps others will chime in. The 0.2 um PES filters will work fine. Vic

Link to this post | posted 27 Sep, 2021 14:56

viknesh

cheryl.brown@mnstate.edu
Thank you. As a follow-up, are you aware of anyone else having difficulties with VWR filters recently? We have been using the same filters for 3 years and this fall is the first time we have had the problem. It seems like a QA failure.

We ran a test with the current filters and some others on hand, and, using the same force in both, the other filters (also .22 micron syringe filters), only the VWR ones had the contamination problem.

I have some .2 micron PES filters on hand. Can those be used in this application?

We are also going to tweak the pre-filter/coffee filter step to further decrease particles in samples.

I've personally not heard any recent complaints about VWR filters, but perhaps others will chime in. The 0.2 um PES filters will work fine.

Vic

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Syringe filter failure?