Primary cell lines of the red-eared slider were established from somatic slider tissues cultivated in appropriate medium for a certain period. Exponential cell growth started after 2 weeks of continuous cultivating. The cells were of various morphology resembling keratinocytes, fibroblasts, and melanocytes, some of them gathered to form dense conglomerates similar to so-called “embryoid bodies”. The cell lines were maintained for 3–7 passages before being taken for cytogenetic manipulations. Red-eared slider cell lines derived in this way can be cryopreserved and successfully recovered to provide a source of fixed cells for 3D imaging as well as a source of chromosomes and chromatin fiber preparations for cytological and cytogenetic studies. The protocol for red-eared slider cells was developed on the basis of the experiments described.