A simple protocol for extracting proteins from phytoplankton

• Water samples were filtered through glass fibre filters (47 mm diameter GF/F (Glass Fibre)) to trap the phytoplankton cells.
  
• The frozen GF/F filter was always kept on ice.
  
• After a slight thaw, the filter was folded with tweezers and inserted into a 15 ml plastic centrifugation vial.
  
• Total cellular proteins present on the frozen GF/F filter were solubilised in 750 µl of solubilisation buffer (100 mM Tris-HCl, pH 8.3, 320 mM sucrose, 1 mM EDTA, 1% sodium dodecyl sulfate and 0.5% dithiothreitol).
  (Note: addition of protease inhibitors such as PefaBloc should be considered).
  
• Cells were disrupted by 30 s of continuous sonication until a smooth paste was obtained. Big chunks of filters should not be apparent. (The vial was always kept on ice during the sonication in order to avoid heating).
  
• The tube was centrifuged and the extract supernatant retrieved.
  

Extraction efficiencies can be estimated by serial re-extractions of the pellet with fresh additions of solubilisation buffer.