Use of Refrigeration as a Practical means to Preserve Viability of in Vitro-Cultured IDE8 Tick Cells

Abstract

In vitro cultivation of the IDE8 cell line, derived from embryonic Ixodes scapularis ticks, constitutes an important system for the study of tick-borne pathogens, as these cells support growth of rickettsial species which are not normally transmitted by this tick. However, since cryopreservation of IDE8 cells is not always successful, there is a need to develop alternative ways to preserve these cells. In the present study, a suspension of IDE8 cells in culture medium was kept under refrigeration at 4 degrees C for up to 60 days. Every 15 days, the suspension was mixed and aliquots were re-cultured in 2-ml tubes, under standardized conditions. In addition, three techniques for cryopreservation, using two different cryoprotectants (DMSO and glycerol), were evaluated. Medium changes were carried out every week and subculturing every 2 weeks. The development of cultures and their respective subcultures, after returning to standard culture temperature, was evaluated by percentage viability and by cellular morphology evaluated in Giemsa-stained cytocentrifuge smears. All cultures and subcultures appeared healthy, showing growth rates comparable to cultures that had not been kept under refrigeration. The results demonstrated that storage under refrigeration at 4 degrees C is an efficient method for preservation of IDE8 cells for up to 60 days and that refrigeration may be preferable to cryopreservation for short-term preservation of IDE8 cells.

Tick cell lines: tools for tick and tick-borne disease research

“…They divide relatively slowly, can be maintained at high cell densities (10 6 -10 7 cells ml À1 ), and many lines do not require regular subculture, making them particularly suitable for isolation of slow-growing microorganisms. Short-term storage at 12 8C [3] or even 4 8C [59] is often preferable to cryopreservation, as frozen tick cells can be difficult to resuscitate reliably [3,57]; however, cells cryopreserved in liquid nitrogen can survive for at least 12 years [18]. Individual cultures of some tick cell lines can be extremely long-lived, surviving for several years with regular medium changes and occasional subcultures, reflecting the ability of ixodid ticks to exist for extremely long periods of time between blood meals in nature.…”

In vitro establishment and propagation of a Brazilian strain ofAnaplasma marginale with appendage in IDE8 (Ixodes scapularis) cells

“…The IDE8 line had been established from embryonic Ixodes scapularis ticks (14) and recently we have reported efficient ways to preserve this cell line by refrigeration and cryopreservation (3). Few other isolates of A. marginale have been established in vitro in IDE8 cells (4,5,15,24).…”

In vitro cultivation of Anaplasma marginale and A. phagocytophilum in tick cell lines: a review

“…Cryopreservation of tick cells in liquid nitrogen can be difficult; however, some cell lines can be preserved under refrigeration for up to one month (BASTOS et al, 2006;LALLINGER et al, 2010).…”