Ultrastructural analysis of Caenorhabditis elegans cilia

In addition to organizing centrosomes, centrioles have an evolutionarily conserved role as basal bodies in the formation of cilia. The discovery of a range of human genetic disorders linked to cilia dysfunction has led to a resurgence of interest in this cellular organelle. The nematode Caenorhabditis elegans possesses several unique features (highly stereotypical morphology, dispensability of cilia for organismal viability and fertility) that make it an attractive model to study cilia assembly and function. However, both the adult worm and the embryo present particular challenges for electron microscopy (EM), which remains the gold standard for high-resolution morphological studies. Here, we present a step-by-step guide for the ultrastructural analysis of C. elegans cilia, including optimized protocols for standard chemical fixation as well as high-pressure freezing/freeze substitution and further processing for serial-section transmission electron microscopy and electron tomography.