PhD 3

Non-invasive visualization of changes in reproductive cells during cryopreservation

Location: Institute for Multiphase Processes, Leibniz University Hannover, Germany

Main Supervisor: Prof. Birgit Glasmacher. For any questions, please contact Dr. Marc Müller, e-mail: mueller@imp.uni-hannover.de

Participating researchers: Dr. Marc Müller (LUH), Prof. Alexander Petrenko (NASU), Dr. Fabian Will (LLS Rowiak)

Objectives:

  1. Develop and validate a Raman microscopy system with temperature controlled chamber for monitoring CPA/VA diffusion into the cells. Understanding of diffusion of CPAs/VAs into the cells is of utmost importance to ensure efficient cryopreservation. The diffusion chamber will be designed, manufactured at the institutional mechanical workshop and validated with reproductive cells. The diffusion of CPA/VA will be monitored in a dynamic perfusion chamber following specific RAMAN peaks of the respective components of the CPA/VA solution (DMSO, ethylene glycol, formamide, propylene glycol, sucrose, trehalose). The calculated diffusion coefficients will be compared to those from the analysis of CPA/VA diffusion using cryomicroscopy based on changes in cell volume (NASU partner).

  2. Characterize structural changes of sperm and oocytes due to cryopreservation using different cooling rates and CPAs/VAs. Among the structural changes appearing during cooling/warming procedures as well as addition and removal of CPAs/VAs we will focus on analysis of cytoplasmic lipid reduction, membrane fatty acid composition alteration, cholesterol and aquaporin content.

Expected Results:

  1. Improved understanding of the diffusion in sperm and oocytes during CPA/VA loading

  2. Working setup for visualization of CPA/VA loading at different temperatures

Planned secondments:  (Host, Supervisor, timing, length, and purpose)

  1. NASU, Petrenko, M12, 2 M, Training in cryo-preservation techniques, experiments on diffusion of CPA/VA via cryomicroscopy

  2. NURE, Avrunin, M18, 2 M, Training in image analysis techniques incl. application of neural networks

  3. IMV, Gavin-Plagne, M24, 1 M, Training in industrial cryopreservation standards

Enrollment in Doctoral Degree: Yes (LUH)