Efficient screening of high-signal and low-background antibody pairs in the bio-bar code assay using prion protein as the target.

The bio-bar code assay is an assay for ultrasensitive detection of proteins. The main technical hurdle in bio-bar code assay development is achieving a dose-dependent, reproducible signal with low background. We report on a magnetic bead ELISA screening mechanism for characterizing antibody pairs that are effective for use in the bio-bar code assay. The normal isoform of prion protein was utilized as the target protein as dozens of antibodies have been developed against it. The development of an ultrasensitive assay for the detection of the various isoforms of PrP has the potential to enable significant advances in the diagnosis and understanding of transmissible spongiform encephalopathies, including transmission mechanisms, disease pathology, and potential therapeutics. With prion protein as the target, the magnetic bead ELISA identified pairs with high background and low signal in the bio-bar code assay. The magnetic bead ELISA was effective as a screening mechanism because it reduced assay time and cost and allowed for understanding of pair characteristics such as development times and signal-to-noise ratios.

Effects of different post-spin stretching conditions on the mechanical properties of synthetic spider silk fibers.

Spider silk is a biomaterial with impressive mechanical properties, resulting in various potential applications. Recent research has focused on producing synthetic spider silk fibers with the same mechanical properties as the native fibers. For this study, three proteins based on the Argiope aurantia Major ampullate Spidroin 2 consensus repeat sequence were expressed, purified and spun into fibers. A number of post-spin draw conditions were tested to determine the effect of each condition on the mechanical properties of the fiber. In all cases, post-spin stretching improved the mechanical properties of the fibers. Aqueous isopropanol was the most effective solution for increasing extensibility, while other solutions worked best for each fiber type for increasing tensile strength. The strain values of the stretched fibers correlated with the length of the proline-rich protein sequence. Structural analysis, including X-ray diffraction and Raman spectroscopy, showed surprisingly little change in the initial as-spun fibers compared with the post-spin stretched fibers.