Evaluation Of An Automated 3‐D Printed Sperm Cryopreservation Device For Use With Eastern Oysters (Crassostrea Virginica): Urop Final Report February 15, 2021

Many populations of Eastern oysters (Crassostrea virginica) in Louisiana are vulnerable to environmental changes. This has created a need for cryopreservation to preserve genetic diversity and offset declining availability of wild stocks for use in breeding. Eastern oysters have been subjected to over‐harvesting and depletion of their main natural habitats. Oyster hatcheries and farms exist but can only do so much when it comes to preserving genetic diversity. They are limited not only in the availability of wild stocks to breed with, but also in maintaining these critical genetic lines as live animals. Sperm repositories would allow preservation of samples to ensure viability and longevity of the Eastern oyster genetic lines. Cryopreservation plays a critical role in this, but currently its accessibility to breeding programs is limited. Existing methods for cryopreservation can be expensive, and breeding programs do not have access to the necessary machines and devices. To address these issues, the goal of this research was to develop a 3‐D printed Auto‐Position Cooling Device (APCD) along with electronic components to automate freezing hardware for use with portable cryogenic shipping dewars. The objectives of this project were to: 1) develop a 3‐D printed device that can accommodate French straws and has the capability to be integrated with a linear actuator; 2) develop a robust user interface (for input of user commands and data, and real‐time monitoring) based on an LCD screen and navigation buttons; 3) construct a temperature measurement system based on sample and chamber thermocouples, and digital conversion and linearization modules; 4) develop and algorithm to control sample height using a microcontroller (Arduino) and a linear actuator to provide calibration procedures and freezing profiles based on recorded temperatures, and 5) perform cryopreservation of sperm from Eastern oysters to evaluate feasibility of freezing and recovering biological samples. A portable version was fabricated with a microcontroller (Arduino Uno), 3‐ dimenstional (3‐D) printed components, Liquid Crystal Display (LCD), linear actuator, and a thermocouple. This device can control the freezing rate through a user‐interface based on different heights within the dewar. Because of the fabrication methods and use of consumerlevel electronics this device was inexpensive (~$200 total) and was easy to use with minimal training.