Currently, millions of serum biospecimens are being stored in biorepositories across the nation, while tens of thousands of new biospecimens are added to the pool daily. These biospecimens are stored for future research, mainly for proteinaceous biomarker discovery and verification (e.g. for diagnostic, therapeutic, and epidemiologic outcomes). The success of biomarker research not only depends upon the availability of the tools (proteomic, peptidomic, lipidomic and metabolomic technologies) to extract information from biospecimens, but also on the availability of """"""""high quality"""""""" biospecimens. However, in most biorepositories, serum is stored by freezing without following any preservation protocol;the samples are directly placed in -20, -40 or -80oC freezers, in the absence of any cryoprotectant, where they experience very slow cooling (1-2o C/min). It is well known that these conditions impose very harsh chemical and physical stresses on macromolecules, altering their characteristics (structure and activity), often irreversibly. Recent evidence has shown that some of the most promising proteinaceous cancer biomarkers are indeed very susceptible to freeze/thaw and frozen state storage. Therefore, it is plausible that frozen state storage may cause most of the potential biomarker information in the stored sera biospecimens to be lost forever. Our long-term goal is to eliminate the requirement for frozen state storage and develop the techniques to store serum biospecimens at room temperature using isothermal vitrification technology. Isothermal vitrification is the process by which liquids doped with sugars are desiccated to a """"""""glass"""""""" (a very viscous fluid). In this state, biochemical reactions are halted, degradation of the specimen is stopped, and macromolecules are stabilized in their native states. Isothermal vitrification will eliminate the exposure of the proteinaceous biomarkers to freeze/thaw stresses and to frozen state storage damage and thus will substantially increase the quality of the stored biospecimens. It will also present a more economical alternative to freezing, since storage of specimens in freezer-farms will no longer be needed. We will accomplish our goal by achieving the following four Aims:
Aim #1. Develop a panel of lyoprotectant chemicals to be added to serum samples for isothermal vitrification: Demonstrate that the lyoprotectant cocktail enables rapid and uniform vitrification while stabilizing sera proteins.
Aim #2. Determine the retention and elution efficiency of sera proteins following vitrification by adsorption in standard filter paper using the developed lyoprotectant cocktail.
Aim #3. Determine whether specific sera proteins are altered following optimized vitrification and elution conditions from the optimal matrix;document protein depletion, aggregation and degradation.
Aim #4. Develop and validate an electrospun sponge for streamlined vitrification of serum samples at sera collection sites.
The advent of genomics and proteomics for personalized medicine has placed biospecimen research on the forefront of NIH priorities, since research projects are only as good as the biospecimens used. We are developing methodology to stably store liquid biospecimens, such as sera, at room temperature. This technology will eliminate the need for long-term storage of serum samples in freezer farms, which are very costly to operate.
|Solivio, Morwena J; Less, Rebekah; Rynes, Mathew L et al. (2016) Adsorbing/dissolving Lyoprotectant Matrix Technology for Non-cryogenic Storage of Archival Human Sera. Sci Rep 6:24186|
|Twomey, Alan; Kurata, Kosaku; Nagare, Yutaka et al. (2015) Microheterogeneity in frozen protein solutions. Int J Pharm 487:91-100|
|Less, Rebekah; Boylan, Kristin L M; Skubitz, Amy P N et al. (2013) Isothermal vitrification methodology development for non-cryogenic storage of archival human sera. Cryobiology 66:176-85|
|Twomey, Alan; Less, Rebekah; Kurata, Kosaku et al. (2013) In situ spectroscopic quantification of protein-ice interactions. J Phys Chem B 117:7889-97|