Trehalose, an easy, safe and efficient cryoprotectant for the parasitic protozoan Trypanosoma brucei

Abstract

Trehalose, a non-permeating cryoprotective agent (CPA), has been documented as less toxic and highly efficient at cryopreserving different kinds of cells or organisms. In the present study, trehalose was evaluated for its application in cryopreservation of both Trypanosoma brucei procyclic and bloodstream form cells. The cryopreservation efficiency was determined by the motility of trypanosomes after thawing, as well as a subsequent recovery and infectivity assessment. The viability of trypanosomes from cultivation that were frozen in a serial concentrations of trehalose showed similar results to classical CPAs of glycerol and DMSO. Nevertheless, trypanosomes cryopreserved in 0.2M trehalose showed the best growth characteristic during subsequent cultivation. In addition, CPA cocktails with trehalose and permeating CPA glycerol or DMSO were developed and evaluated. Interestingly, trypanosomes in host (mouse) blood cryopreserved in 0.4M trehalose plus 5% glycerol showed higher infectivity than those preserved in trehalose/DMSO cocktails as well as individually. Further investigations showed that, in comparison with slow freezing at -80°C, flash freezing in liquid nitrogen provided better cryopreservation for bloodstream form cells than slow freezing. In conclusion, trehalose is an easy, safe and efficient CPA for cryopreservation of T. brucei and potentially for other protozoan species and cells.

Comparison of Sucrose and Trehalose for Protein Stabilization Using Differential Scanning Calorimetry

“…However, although the exact mechanisms for protein stabilization at both low and high temperatures are somewhat unknown, different models for stabilization have been proposed. Since it has been experimentally verified that the disaccharides sucrose and trehalose are particularly successful stabilizers, especially the latter, as shown in other works, ,− most models have been proposed to explain their stabilizing role.…”

Trypanosoma congolense: In Vitro Culture and Transfection

“…Cryopreservation is a way of preserving trypanosome strains in the laboratory without the need for continuous culture and also provides a safeguard against loss through contamination with other microorganisms or other disaster. We routinely cryopreserve using glycerol as cryoprotectant, but DMSO and trehalose may also be used (Wen et al, 2016). For best results, cultures should be dense and nearing the end of logarithmic growth when cryopreserved.…”