Drosophila larvae were sandwiched between two 3 mm aluminum slit carriers with the 100 µm cavities facin each other and high-pressure frozen with a Leica EM HPM100. No ethanol as synchronization media was used, 1-hexadecene was used as filler. The wholes of the slit carriers were filled with filter tips dipped in 1-hexadecene to keep the carrier sandwich complete after freezing.
The carrier sandwich was mounted on a self-made 3mm holder in the VCT100 loading station and transferred into an ACE900 prototype using the VCT100 shuttle and the VCT500 dock mounted on the ACE900 (connected by the adapter plate). The cryo-stage was set to -120°C and the holder was sitting there for 15min to reach equilibrium (since the sample came from