Modified Filter-Transfer-Freeze ("FTF") Technique for Raman Microspectroscopic Analysis of Single Cells V.2

Published Protocol Version 2

   A method for concentrating aquatic microbes for single-cell analysis by Raman microspectroscopy. Modified from Hewes and Holm-Hansen (1983): A Method for Recovering Nanoplankton from Filters for Identification with the Microscope: The Filter-Transfer-Freeze (FTF) Technique. Limnology and Oceanography. 28(2): 389-394.

     Many materials commonly used in microscopy to examine individual microorganisms contribute contaminating Raman scattered emissions, e.g., most types of membranes and filters, glass slides, cover slips, immersion oil, glycerol, etc. and are therefore unsuitable for this application. We developed a protocol that enables single-cell analysis while minimizing extraneous Raman scattered photons.  The technique transfers cells from popular polycarbonate membranes to mirror-finished 304 stainless steel slides (1x3x0.0235') supplied by Stainless Supply® (Monroe, NC USA).