Optimization of cryoprotectant addition and removal procedures for vitrification of adherent mammalian cells Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/3484zn375

Descriptions

Attribute NameValues
Creator
Abstract or Summary
  • Cryopreservation of adherent cells may be advantageous for cell types that are difficult to preserve in suspension or when it is necessary to preserve characteristics of the adherent cultured cells. Vitrification is a promising procedure for the preservation of adherent cells that prevents ice crystal formation and the resulting dissociation and morphological damage. To successfully vitrify adherent cells, high concentrations of CPA are required which increases the likelihood of osmotic and toxic damage. In this dissertation, we describe a rational design strategy that predicts mathematically optimized CPA addition and removal procedures based on the minimization of a toxicity cost function. These rationally designed procedures rely on the accurate knowledge of cell biophysical parameters. We validate an in situ calcein fluorescence quenching method for the determination of membrane permeability parameters for adherent cells. We also describe the determination of osmotic tolerance limits for adherent cells. We use rational design strategies to determine CPA addition and removal procedures for adherent endothelial cells, neuronal cells, and induced pluripotent stem cells as well as oocytes. Also, we provide experimental support for the feasibility of these methods using adherent endothelial cells. The mathematical methods and experimental procedures outlined in this dissertation are important tools for the design of addition and removal procedures for concentrated CPA solutions. This dissertation is an important step toward successful design and implementation of vitrification strategies for adherent cells and tissues.
Resource Type
Date Available
Date Copyright
Date Issued
Degree Level
Degree Name
Degree Field
Degree Grantor
Commencement Year
Advisor
Committee Member
Academic Affiliation
Non-Academic Affiliation
Keyword
Subject
Rights Statement
Peer Reviewed
Language
Replaces

Relationships

Parents:

This work has no parents.

Last modified

Downloadable Content

Download PDF

Items