To facilitate the construction, functional characterization, and use of immunoadsorbents, we have developed a flow cytometry method that allows rapid assessment of large numbers of particle-bound antibodies. Protein G derivitized POROS beads were used to bind affinity-purified antibodies specific for synthetic peptides designed from human plasma proteins. The antibodies were covalently coupled to the beads and used to capture and release synthetic peptides that had been labeled at the C-terminus with the fluorochrome Alexa Fluor 488. Antibody coupling and specificity of antigen binding and release were measured by analysis of the POROS affinity beads by flow cytometry. The affinity-capture matrixes were also used through several antigen-binding and release cycles without loss of peptide binding efficiency. The ability to produce and characterize extremely small amounts of POROS affinity matrices will facilitate their use in protein microchemical procedures such as protein chip technology, monoclonal antibody screening and mass spectrometry, applications where analytes are limiting or present in low abundance in complex mixtures.

A detailed understanding of changes in blood protein biomarkers occuring in individuals over time would enable truly personalized approaches to health and disease monitoring. Such measurements could reveal smaller, earlier departures from normal baseline levels of biomarkers thus allowing better disease detection and treatment monitoring. Current practice, however, generally involves infrequent, sporadic biomarker testing, and this undersampling likely fails to capture important biological phenomena. Here we report the use of a robust multiplex immunomass spectrometric method (SISCAPA) to measure a panel of clinically-relevant proteins in a unique collection of 1,522 dried blood spots collected longitudinally by 8 individuals over periods of up to 9 years, with daily sampling during some intervals. Analytical workflow CVs of 2-6% for most assays were achieved by normalizing DBS plasma volume using a set of 3 minimally varying proteins, facilitating temporal analysis of both high-and low-amplitude biomarker changes compared to personalized baselines. The biomarkers included a panel of 9 positive and 5 negative acute phase response (inflammatory) proteins, allowing longitudinal analysis of inflammation markers associated with major and minor infections, influenza vaccination, recovery from hip-replacement surgery and Crohn's disease. The results illustrate complex time-dependent "biomarker trajectories" on multiple timescales and provide a basis for detailed personalized models of inflammation dynamics. The striking stability of most biomarker protein levels over time, combined with the convenience of self-sampling and low cost of multiplexed measurements using mass spectrometry, provide a new window into the temporal dynamics of disease processes. The extensive results obtained using this high throughput approach offer a new source of precision biomarker 'big data' amenable to machine learning approaches and application to more personalized health monitoring.

Spleen cells from Trypanosoma congolense-infected mice showed a drastic depression in their capacity to respond to B and T lymphocyte mitogens and to allogeneic spleen cells in mixed lymphocyte cultures. Spleen cells from infected mice were also poor stimulators in mixed lymphocyte cultures. The poor responsiveness or stimulation capacity was not due simply to dilution of relevant B or T lymphocytes by the large number of null cells found in the spleens of infected animals. These null cells expressed approximately eight times more H-2 antigen than spleen cells from normal (uninfected) mice and were devoid of Ia antigens.