Silk fibroin is a high molecular weight amphiphilic protein that self-assembles into robust biomaterials with remarkable properties including stabilization of biologicals and tunable release kinetics correlated to processing conditions. Cells, antibiotics, monoclonal antibodies and peptides, among other biologics, have been encapsulated in silk using various processing approaches and material formats. The mechanistic basis for the entrapment and stabilization features, along with insights into the modulation of release of the entrained compounds from silks will be reviewed with a focus on stabilization of bioactive molecules.

Advanced personalized medical diagnostics depend on the availability of high-quality biological samples. These are typically biofluids, such as blood, saliva, or urine; and their collection and storage is critical to obtain reliable results. Without proper temperature regulation, protein biomarkers in particular can degrade rapidly in blood samples, an effect that ultimately compromises the quality and reliability of laboratory tests. Here, we present the use of silk fibroin as a solid matrix to encapsulate blood analytes, protecting them from thermally induced damage that could be encountered during nonrefrigerated transportation or freeze-thaw cycles. Blood samples are recovered by simple dissolution of the silk matrix in water. This process is demonstrated to be compatible with a number of immunoassays and provides enhanced sample preservation in comparison with traditional air-drying paper approaches. Additional processing can remediate interactions with conformational structures of the silk protein to further enhance blood stabilization and recovery. This approach can provide expanded utility for remote collection of blood and other biospecimens empowering new modalities of temperature-independent remote diagnostics.lood contains a variety of proteins, enzymes, lipids, metabolites, and peptides, which can be interrogated as biomarkers for health screening, monitoring, and diagnostics. The integrity of these blood components and thus the quality of information attained from their analysis is determined by the storage conditions from sampling until analysis, or the so-called pre-analytical phase (1, 2). This phase often includes time-consuming processing steps and the requirement for a continuous cold storage. Without temperature regulation, blood-derived biospecimens degrade quickly, accounting for up to 67% of all laboratory testing errors (2, 3). Further, when blood-derived materials are frozen, decreases in thermodynamic free energy and unfavorable ice crystal-protein interactions (4) can occur during subsequent thawing (5, 6), which can further compromise analyte integrity as a gold-standard methodology.As an alternative route, blood and blood derivatives can be dried via newer approaches such as isothermal vitrification (7), lyophilization (8), or on silica chips (9). Isothermal vitrification and lyophilization are inherently resource-intensive techniques and thus not suitable for field use. Silica chips are designed for enrichment of selected fractions of the low-molecular-weight serum proteome, but are not broadly protective at elevated temperature (10). An inexpensive alternative used since the 1960s are dried blood spots (DBS) (11), a paper card system which captures blood components among cellulose fibers as the water phase evaporates. These drying measures decrease sample weight by >90%, thus decreasing transport burden, and in theory can enhance long-term sample stability by decreasing water-dependent analyte degradation caused by hydrolysis and enzyme activity (12). Unfortunately, DBS stored in ...

The use of mRNA and miRNA as diagnostic parameters and therapeutic agents has drawn wide interest both clinically and in research. However, RNA is a labile molecule, which requires strict storage conditions, often including cold temperatures or dry environments, in order to preserve RNA integrity. Achieving this requires huge costs for storage and added difficulty in transport. To address these issues, we introduce a system to encapsulate and store it long-term in dried silk fibroin matrices. At temperatures up to 45 °C, mRNA samples stored in lyophilized silk matrices showed good stability over 1 week, as measured by real-time PCR to assess transcript recovery. While the presence of the silk interfered with the generation of cDNA required for quantitation at roughly 1% w/v (400:1 silk:RNA mass), this phenomenon was resolved by the use of commercial RNA purification kits for silk concentrations up to 4% w/v. A higher concentration of silk correlated with increased thermal protection. As an alternative to lyophilization, RNA was simply air-dried in the presence of aqueous fibroin to create storage matrices. While air-dried matrices composed of low silk content were not protective, higher concentrations were protective and progressively lost additional water over time of storage because of the overall hydrophobic nature of the system. Taken together, these findings provide a new and potentially simpler method for preserving RNA samples for long-term storage and transportation, acting primarily on a water exclusion mechanism.

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Preliminary studies have shown that silk fibroin can protect biomacromolecules from thermal degradation, but a deeper understanding of underlying mechanisms needed to fully leverage the stabilizing potential of this matrix has not been realized. In this study, we investigate stabilization of plasma C-reactive protein (CRP), a diagnostic indicator of infection or inflammation, to gain insight into stabilizing mechanisms of silk. We observed that the addition of antiplasticizing excipients that suppress β-relaxation amplitudes in silk matrices resulted in enhanced stability of plasma CRP. These observations are consistent with those made in sugar-glass-based protein-stabilizing matrices and suggest fundamental insight into mechanisms as well as practical strategies to employ with silk protein matrices for enhanced stabilization utility.