This video documents small volume (~1 L) filtration of microbial biomass from the water column.
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Method Article
Small Volume (1-3L) Filtration of Coastal Seawater Samples
In This Article
Summary
Abstract
The workflow begins with the collection of coastal marine waters for downstream microbial community, nutrient and trace gas analyses. For today s demonstration samples were collected from the deck of the HMS John Strickland operating in Saanich Inlet. This video documents small volume (~1 L) filtration of microbial biomass from the water column. The protocol is an extension of the large volume sampling protocol described earlier, with one major difference: here, there is no pre-filtration step, so all size classes of biomass are collected down to the 0.22 μm filter cut-off. Samples collected this way are ideal for nucleic acid analysis. The set-up, filtration, and clean-up steps each take about 20-30 minutes. If using two peristaltic pumps simultaneously, up to 8 samples may be filtered at the same time. To prevent biofilm formation between sampling trips, all filtration equipment must be rinsed with dilute HCl and deionized water and autoclaved immediately after use.
Protocol
- Set up of filtration apparatus
- Before filtering, first wipe away any condensation from the full bottles and weigh them
- Place an autoclave bin on one side of the pump, and the waste flask on the other.
- Then place a full sample bottle in the autoclave bin; however, do not remove the cap of the bottle yet.
- Pass the tubing through the peristaltic pump so that the luer end of the tubing is in the flask.
- Using tweezers, remove the cotton from a sterile 25 ml pipette, and attach the pipette to the other end of the tubing.
- Filtration of seawater
- To begin filtering seawater, open the ....
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Discussion
It is imperative to wipe down benches and pumps with a damp towel after use to remove any lingering traces of salt water that would otherwise corrode equipment. Although the procedure is simple, the time requirement can be extensive depending on the concentration of biomass or debris in the sample being filtered.
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Disclosures
The authors have nothing to disclose.
Acknowledgements
We would like to thank the Canadian Foundation for Innovation, the British Columbia Knowledge Development Fund and the National Sciences and Engineering Research Council (NSERC) of Canada for supporting ongoing studies on low oxygen regions of coastal and open ocean waters. D.A.W. was supported by fellowships from NSERC, Killam and the TULA foundation funded Centre for Microbial Diversity and Evolution.
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Materials
| Name | Company | Catalog Number | Comments |
|---|---|---|---|
| Peristaltic pump | Masterflex (Cole Palmer) | 77913-20, 77200-62, 77200-04 | |
| Tygon LFL tubing | Masterflex (Cole Palmer) | 06429-24 | For filtration apparatus |
| Hose clamps | Cole-Parmer | 0683204 | For filtration apparatus |
| 1/4” x 3/16” pipe adaptors | Cole-Parmer | 31702-20 | For filtration apparatus |
| Polypropylene prefilter housing | ADVANTEC | 501200 | For filtration apparatus |
| Metal luer fitting (Hose end to Male luer lock) | Cole-Parmer | 31507-27 | For filtration apparatus |
| 2“ Magnets | For filtration apparatus | ||
| 3 port carboy caps | For filtration apparatus | ||
| GV 0.22 μm STERIVIX filters | EMD Millipore | SVGV01R5 | |
| 47 mm diameter 2.7 μm GF/D filters | Whatman, GE Healthcare | 1823047 | |
| Lysis buffer | 0.75 M sucrose, 40 mM EDTA, 50 mM Tris (pH 8.3) | ||
| 0.1 % HCl | For post-filtration rinse | ||
| Autoclaved water | For post-filtration rinse |
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