Further Development of an Equine Cell Line that can be Propagated over 100 Times

Abstract

Cell lines originating from horses are necessary for isolation and propagation of equine herpesviruses (EHV). Although we established an equine-derived cell line, FHK-Tcl3, propagation ceased after fewer than 40 passages. In this study, FHK-Tcl3 cell propagation continued beyond 40 passages, achieving over 100 passages. FHK-Tcl3 cells were then cloned by limiting dilution at the 100th passage. Cloned cells were termed FHK-Tcl3.1. FHK-Tcl3.1 cells grew well and were propagated every 3 to 4 days by splitting 1:5. In addition, EHV-1, -2 and -4 showed a clear cytopathic effect (CPE) in FHK-Tcl3.1 cells, and this CPE was very similar to those seen in parental FHK-Tcl3 and primary fetal horse kidney cells. FHK-Tcl3.1 cells continue to propagate and the current passage record is over 100 times after cloning. Therefore, this cell appears to have been immortalized. FHK-Tcl3.1 cells have potential for growth and diagnosis of various equine viruses, including equine herpesviruses.

Keywords: cell line; equine herpesvirus.

Fig. 1.

Growth of cloned cells. Vertical error bars indicate standard deviation of means.

Fig. 2.

CPE in FHK-Tcl3.1 cells after infection with EHV-1, -2 and -4. FHK-Tcl3.1 cells were infected with EHV-1 (A), EHV-4 (B), EHV-2 (C) or only medium (D). Cytopathic morphological changes were observed after 32 hr (EHV-1), 72 hr (EHV-4) and 153 hr (EHV-2).