Comprehensive characterization of the antigen-specific B cells induced during infections or following vaccination would facilitate the discovery of novel antibodies and inform how interventions shape protective humoral responses. The analysis of human B cells and their antibodies has been performed using flow cytometry to evaluate memory B cells and expanded plasmablasts, while microtechnologies have also provided a useful tool to examine plasmablasts/plasma cells after vaccination. Here we present an integrated analytical platform using arrays of subnanoliter wells (nanowells) for constructing detailed profiles for human B cells comprising the immunophenotypes of the cells, the distribution of isotypes of secreted antibodies, the specificity and relative affinity for defined antigens, and for a subset of cells, the genes encoding the heavy and light chains. The approach combines on-chip image cytometry, microengraving, and single-cell RT-PCR. Using clinical samples from HIV-infected subjects, we demonstrate that the method can identify antigen-specific neutralizing antibodies, is compatible with both plasmablasts/plasma cells and activated memory B cells, and is well-suited for characterizing the limited numbers of B cells isolated from tissue biopsies (e.g., colonic biopsies). The technology should facilitate detailed analyses of human humoral responses for evaluating vaccines and their ability to raise protective antibody responses across multiple anatomical compartments.

Since signals via CD40 and the B cell receptor are known to synergize to induce B cell activation, we have analyzed the pocket protein / E2F complexes in mouse B lymphocytes following stimulation by anti‐IgM, anti‐CD40, alone or together. We find that E2F4 and DP1 form the predominant E2F heterodimers in the G0 and G1 phases of the cell cycle, complexed with hypophosphorylated p130. During late G1 and S phase this complex is replaced by at least three different E2F complexes, one of which is an E2F complex containing p107 or pRB as well as two “free” E2F complexes consisting of E2F4 / DP1 and E2F1 – 3 / DP1. These effects were mirrored by the levels and phosphorylation status of the three pocket proteins. We also observed an increase in electrophoretic mobility of DP1 and E2F4 as B cells progressed from G0 into early G1, resulting from their dephosphorylation. This is known to correlate with a decrease in DNA binding capacity of these proteins and could also be important for derepression of genes negatively regulated through E2F sites in their promoters. These results therefore indicate that the pRB / E2F pathway integrates proliferative signals emanating from the sIgM and CD40 receptors.

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Engine design studies are often conducted with one of two distinct focuses on two different scales: overall engine analysis or localized surface analysis. However, recent engine research requires a more combined approach. Classical tribological experiments and models have been developed usually for general contacting surfaces under specific operating conditions. They attempt to simulate the actual engine environment, which varies significantly with component design and engine operating conditions. Macroscopic engine studies, on the other hand, yield information pertaining to the overall engine system, such as component dynamics, lubricant flow, pressures, and temperatures. These macroscopic operating parameters serve as boundary conditions to the localized surface or tribochemical problem. This paper discusses the macroscopic oil transport processes in the engine and presents several current engine performance problems, such as in friction reduction, modelling wear with antiwear tribofilms, and minimizing deposit formation and emission of ash-related species. It also illustrates applications of the combined analysis of bulk oil transport in the engine with material surface phenomena and tribochemistry. A specific example involves the oil supply to the piston ring-pack and changes in oil and additive characteristics in that region.

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