Cold Box for 2mL Cryotubes

Comparable to a $200 commercial cold box, this design cost me less than $3 of materials to make. Designed to hold 5x glycerol stocks in cryotubes to allow work without thawing, allowing the stocks to be reused until empty. Tube dimensions are 11mm in diameter and sit 30mm into the box, with room for the caps to sit under the lid for additional insulation.

Printing:
-Print the cryobox top at 100 percent infill.
-Print the cryobox bottom at 15 percent infill.
-Print the cryobox lid at 100 percent infill.
-Before proceeding, dry fit both parts to confirm they nest cleanly and that all tubes fit properly into their slots.

Fill Port Creation:
-Using the smallest drill bit available, drill a very small hole through the top surface of the box.
-This hole will serve as the fill port for the thermal buffer solution.

Initial Sealing:
-Apply a thin, continuous layer of silicone sealant to all internal surfaces that will contact liquid.
-Allow the silicone to cure in open air for 24 hours.

Final Assembly:
-After the initial cure, apply additional silicone sealant to the mating surfaces in a continuous bead.
-Press the top and bottom together firmly to fully seal the chamber. Wipe away excess silicone.
-Allow the assembled unit to cure for another 24 hours.

Filling the Chamber:
-Prepare a 15 to 20 percent glycerol in water solution.
-Using a syringe, inject the solution through the fill port until the chamber is completely filled.
(This glycerol concentration freezes into a slushy consistency, which provides better thermal buffering than pure alcohol or commercial gel ice pack materials. Optionally use IPA or ice pack gel, understanding that it will not work quite as well)

Sealing the Fill Port:
-Coat the threads of a small screw with silicone sealant.
-Insert the screw into the fill hole and tighten gently.
-Apply a small amount of additional silicone over the screw head if needed.
-Allow the sealant to fully cure before placing the box into the freezer to chill overnight before using.

To allow better contact between tube slots and cryotubes, can optionally add about 250 microliters of the glycerol solution to each slot. This will turn into a slush that will fill any voids when a tube is inserted, maximizing working time.