Cold storage and cryopreservation of tick cell lines.
BACKGROUND
Tick cell lines are now available from fifteen ixodid and argasid species of medical and veterinary importance. However, some tick cell lines can be difficult to cryopreserve, and improved protocols for short- and long-term low temperature storage will greatly enhance their use as tools in tick and tick-borne pathogen research. In the present study, different protocols were evaluated for cold storage and cryopreservation of tick cell lines derived from Rhipicephalus (Boophilus) decoloratus, Rhipicephalus (Boophilus) microplus, Ixodes ricinus and Ixodes scapularis. For short-term cold storage, cells were kept under refrigeration at 6 degrees C for 15, 30 and 45 days. For cryopreservation in liquid nitrogen, use of a sucrose-phosphate-glutamate freezing buffer (SPG) as cryoprotectant was compared with dimethylsulfoxide (DMSO) supplemented with sucrose. Cell viability was determined by the trypan blue exclusion test and cell morphology was evaluated in Giemsa-stained cytocentrifuge smears.


RESULTS
Cold storage at 6 degrees C for up to 30 days was successful in preserving R. (B.) microplus, R. (B.) decoloratus, I. ricinus and I. scapularis cell lines; lines from the latter three species could be easily re-cultivated after 45 days under refrigeration. While cell lines from all four tick species cryopreserved with 6% DMSO were successfully resuscitated, the R. (B.) decoloratus cells did not survive freezing in SPG and of the other three species, only the R. (B.) microplus cells resumed growth during the observation period.


CONCLUSIONS
This constitutes the first report on successful short-term refrigeration of cells derived from R. (B.) decoloratus, R. (B.) microplus, and I. ricinus, and use of SPG as an alternative to DMSO for cryopreservation, thus making an important contribution to more reliable and convenient tick cell culture maintenance.
Lallinger
Gertrud
G
Lehrstuhl für Vergleichende Tropenmedizin und Parasitologie, Ludwig-Maximilians-Universitaet Muenchen, Leopoldstr, 5, 80802, Munich, Germany. Lygia.Passos@lmu.de.
Zweygarth
Erich
E
Bell-Sakyi
Lesley
L
Passos
Lygia Mf
LM
Parasitology. 2004;129 Suppl:S3-14
15938502
Exp Appl Acarol. 2002;28(1-4):177-85
14570129
J Med Entomol. 1975 Jan 10;11(6):698-706
1123829
Front Biosci (Landmark Ed). 2009 Jan 01;14:2857-77
19273240
J Parasitol. 1991 Dec;77(6):1006-8
1779279
Exp Appl Acarol. 2009 Nov;49(3):209-19
19252822
Trends Parasitol. 2007 Sep;23(9):450-7
17662657
J Med Entomol. 2007 Nov;44(6):1091-101
18047211
Exp Appl Acarol. 2006;39(3-4):347-52
16779573
J Parasitol. 1994 Aug;80(4):533-43
8064520
J Bacteriol. 1950 Apr;59(4):509-22
15436424
Vet Parasitol. 2003 Feb 28;112(1-2):63-73
12581585
J Comp Pathol. 2004 May;130(4):285-93
15053931
Exp Appl Acarol. 1989 Aug;7(3):219-29
2766897