Isolating MPB from sediment: taking the new protocol for a spin

The final step: a glass fiber filter containing cleaned, filtered MPB ready for stable isotope analysis.

This week I tried a new diatom separation protocol. This may not sound exciting, but visually, it is. The MPB collected from the pond is a foamy, brown layer of detritus, diatoms, cyanobacteria, microbes, and even some tiny invertebrates mixed with very fine sediment (See "Mystery of the Fuzz," May 31 2010). Sediments and detritus, however, can contribute to noise in measurements of carbon isotopic composition (because they too contain carbon, sometimes with very different signatures than whatever they're mixed with). SI analysis also requires a certain weight of the compound of interest, and having sediment mixed in with a sample will also make it heavier with material that is not important.
Because of these two issues, people have come up with a few different ways of separating sediment from their samples. The first protocol I'm testing employs very fine mesh filters and a high-density liquid made with silica.
Diatoms and cyanobacteria are luckily smaller than most sediment particles, so the first step is to run the sludge through a mesh to remove large pieces of sediment and little inverts. The resulting foggy water sample is filtered again through a smaller mesh which retains the diatoms, and these are in turn rinsed into a beaker with a small amount of water. Now comes the fun part: the contents of this little beaker are added to a plastic centrifuge tube, which contains a silica liquid that is denser than water. We then centrifuge the diatom/silica/small sediment mixture. Since any remaining sediment particles are heavier than the silica, they spin to the bottom.

Diatoms and cyanos, however, have a lower density than the silica, and they remain at the top of the tube. Centrifuge tube containing colloidal silica and MPB/sediment. The MPB is the bright green layer floating at the top, and there's a lump of sediment at the bottom.

We can then scoop the green-brown layer off the top of the silica, rinse it, and end up with a small, concentrated sample of MPB algae free of sediment. We'll see how these samples turn out!