CLASI-FISH: Principles of combinatorial labeling and spectral imaging

Alex M. Valm, Jessica L. Mark Welch, Gary G. Borisy
2012 Systematic and Applied Microbiology  
Although the number of phylotypes present in a microbial community may number in the hundreds or more, until recently, fluorescence in situ hybridization has been used to label, at most, only a handful of different phylotypes in a single sample. We recently developed a technique, CLASI-FISH for Combinatorial Labeling and Spectral Imaging -Fluorescence in situ Hybridization, to greatly expand the number of distinguishable taxa in a single FISH experiment. The CLASI technique involves labeling
more » ... robes of interest with combinations of probes coupled with spectral imaging to allow the use of fluorophores with highly overlapping excitation and emission spectra. Here, we present the basic principles and theory of CLASI-FISH along with some guidelines for performing CLASI-FISH experiments. We further include a protocol for creating fluorescence spectral reference standards, a vital component of successful CLASI-FISH.
doi:10.1016/j.syapm.2012.03.004 pmid:22521573 pmcid:PMC3407316 fatcat:7ax6gdgjfbaunm7occbveo6ez4