Experimental model for studing the primary cilia in tissue culture cells

In HeLa, PK (pig kidney embryo cells), 3T3, PtK₁ cells and rat embryo fibroblasts (REF), antibodies against acetylated tubulin stained centrioles, primary cilia, some cytoplasmic microtubules and microtubule bundles of the residual body. The primary cilia were stained more intensely than cytoplasmic microtubules and could easily distinguished from them. This makes it possible to detect the primary cilia in cultured cells and to estimate their number by light microscopy. The four studied cultures had 1/4 to 1/3 of interphase cells with detectable primary cilia, and only in HeLa cells the primary cilia were very rare. Comparison of electron microscopic and immunofluorescense data showed that the frequency of primary cilia in four tissue cultures determined by these two methods was the same. Therefore, one can use antibodies against acetylated tubulin for studying the primary cilia. In synchronized mitotic fibroblasts (3T3 and REF) the first primary cilia appeared 2 h after the cells had been plated on coverslips, which is 1 h after the cells had entered the interphase. Four hours after plating the number of ciliated cells reached the average level for nonsynchronous population. This model can be used for further investigation of the expression of primary cilia.