No abstract

As a result of thermal instability, some live attenuated viral (LAV) vaccines lose substantial potency from the time of manufacture to the point of administration. Developing regions lacking extensive, reliable refrigeration ("cold-chain") infrastructure are particularly vulnerable to vaccine failure, which in turn increases the burden of disease. Development of a robust, infectivity-based high throughput screening process for identifying thermostable vaccine formulations offers significant promise for vaccine development across a wide variety of LAV products. Here we describe a system that incorporates thermal stability screening into formulation design using heat labile measles virus as a prototype. The screening of >11,000 unique formulations resulted in the identification of liquid formulations with marked improvement over those used in commercial monovalent measles vaccines, with <1.0 log loss of activity after incubation for 8h at 40°C. The approach was shown to be transferable to a second unrelated virus, and therefore offers significant promise towards the optimization of formulation for LAV vaccine products.

This study investigated the growth of Helicobacter (H.) pylori in Brucella broth supplemented with either IsoVitaleX (1% vol/vol), hemin (.01% wt.vol), agar (0.3% wt/vol), or blood agar blocks (1.5% wt/vol agar). IsoVitaleX was found to significantly shorten the lag phase, while hemin inhibited the growth within the first 24 hours but later acted as a growth stimulant. There was a tendency toward stronger growth when blood agar blocks were added to the medium. Subsequent electron microscopic evaluation revealed that cells of H. pylori were attached to blood agar block surfaces. In contrast, the supplementation of Brucella broth with agar did not significantly increase the cell density. When H. pylori was grown in the presence of IsoVitaleX, strongly stainable electron-dense bodies (140-200 nm) were seen in the cytoplasms. Incubation of cultures on rotary shakers at 10 rpm significantly enhanced growth. The addition of glycerol (15% vol/vol) or fetal bovine serum (15% vol/vol) showed good ultrastructural preservation of bacteria with undamaged cell walls and cytoplasmic membranes, and cytoplasms were ribosome-dense. Cell counts revealed that cultures stored in glycerol or fetal bovine serum had a significantly lower loss in viability when compared with cultures stored without cryopreservatives. Unprotected cells of H. pylori showed on electron micrographs, clumping, cell lysis, and flagellar damage. Finally, the survival rates of H. pylori after multiple thawing from storage at -80 degrees C were best in Brucella broth/glycerol, Brucella broth/fetal bovine serum, and Brucella broth without cryopreservative (in descending order).

Background: This study examined whether hierarchical clustering could be used to detect cell states induced by treatment combinations that were generated through automation and high-throughput (HT) technology. Data-mining techniques were used to analyze the large experimental data sets to determine whether nonlinear, non-obvious responses could be extracted from the data. Methods: Unary, binary, and ternary combinations of pharmacological factors (examples of stimuli) were used to induce differentiation of HL-60 cells using a HT automated approach. Cell profiles were analyzed by incorporating hierarchical clustering methods on data collected by flow cytometry. Data-mining techniques were used to explore the combinatorial space for nonlinear, unexpected events. Additional small-scale, follow-up experiments were performed on cellular profiles of interest. Results: Multiple, distinct cellular profiles were detected using hierarchical clustering of expressed cell-surface antigens. Data-mining of this large, complex data set