Advances in Microwave-Assisted Freeze Substitution

Dorward, DW; Raae-Nielsen, J.; Hansen, BT; Nair, Vila; Fischer, ER

doi:10.1017/s1431927613004790

Cited by 4 publications

(1 citation statement)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Besides optimizing individual FS protocols, efforts were made to accelerate FS on the basis of novel technical know-how. Dorward et al 9,10 introduced microwave-assisted FS combining a laboratory microwave oven with a cryo-thermal platform and accelerated the FS of chemically prefixed tissue culture cells. 11 In 2011, McDonald and Webb 12 demonstrated the tremendous potential of sample agitation for acceleration of the FS process.…”

Section: Introductionmentioning

confidence: 99%

Agitation Modules: Flexible Means to Accelerate Automated Freeze Substitution

Reipert

Goldammer

Richardson

et al. 2018

J Histochem Cytochem.

View full text Add to dashboard Cite

For ultrafast fixation of biological samples to avoid artifacts, high-pressure freezing (HPF) followed by freeze substitution (FS) is preferred over chemical fixation at room temperature. After HPF, samples are maintained at low temperature during dehydration and fixation, while avoiding damaging recrystallization. This is a notoriously slow process. McDonald and Webb demonstrated, in 2011, that sample agitation during FS dramatically reduces the necessary time. Then, in 2015, we (H.G. and S.R.) introduced an agitation module into the cryochamber of an automated FS unit and demonstrated that the preparation of algae could be shortened from days to a couple of hours. We argued that variability in the processing, reproducibility, and safety issues are better addressed using automated FS units. For dissemination, we started low-cost manufacturing of agitation modules for two of the most widely used FS units, the Automatic Freeze Substitution Systems, AFS(1) and AFS2, from Leica Microsystems, using three dimensional (3D)-printing of the major components. To test them, several labs independently used the modules on a wide variety of specimens that had previously been processed by manual agitation, or without agitation. We demonstrate that automated processing with sample agitation saves time, increases flexibility with respect to sample requirements and protocols, and produces data of at least as good quality as other approaches.

show abstract

Section: Introductionmentioning

confidence: 99%