Last data update: Jan 27, 2025. (Total: 48650 publications since 2009)
Records 1-3 (of 3 Records) |
Query Trace: McDonald MA[original query] |
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Development of a new peptide-bead coupling method for an all peptide-based Luminex multiplexing assay for detection of Plasmodium falciparum antibody responses
Wakeman BS , Shakamuri P , McDonald MA , Weinberg J , Svoboda P , Murphy MK , Kariuki S , Mace K , Elder E , Rivera H , Qvarnstrom Y , Pohl J , Shi YP . J Immunol Methods 2021 499 113148 Using a recombinant protein antigen for antibody testing shows a sum of antibody responses to multiple different immune epitopes existing in the protein antigen. In contrast, the antibody testing to an immunogenic peptide epitope reflects a singular antibody response to the individual peptide epitope. Therefore, using a panel of peptide epitopes provides an advantage for profiling multiple singular antibody responses with potential to estimate recent malaria exposure in human infections. However, transitioning from malaria immune epitope peptide-based ELISA to an all peptide bead-based multiplex Luminex assay presents some challenges including variation in the ability of different peptides to bind beads. The aim of this study was to develop a peptide coupling method while demonstrating the utility of these peptide epitopes from multiple stage antigens of Plasmodium falciparum for measuring antibodies. Successful coupling of peptide epitopes to beads followed three steps: 1) development of a peptide tag appended to the C-terminus of each peptide epitope consisting of beta-alanine-lysine (x 4)--cysteine, 2) bead modification with a high concentration of adipic acid dihydrazide, and 3) use of the peptide epitope as a blocker in place of the traditional choice, bovine serum albumin (BSA). This new method was used to couple 12 peptide epitopes from multiple stage specific antigens of P. falciparum, 1 Anopheles mosquito salivary gland peptide, and 1 Epstein-Barr virus peptide as an assay control. The new method was applied to testing of IgG in pooled samples from 30 individuals with previously repeated malaria exposure in western Kenya and IgM and IgG in samples from 37 U.S. travelers with recent exposure to malaria. The new peptide-bead coupling method and subsequent multiplex Luminex assay showed reliable detection of IgG to all 14 peptides in Kenyan samples. Among 37 samples from U.S. travelers recently diagnosed with malaria, IgM and IgG to the peptide epitopes were detected with high sensitivity and variation. Overall, the U.S. travelers had a much lower positivity rates of IgM than IgG to different peptide epitopes, ranging from a high of 62.2% positive for one epitope to a low of only 5.4% positive for another epitope. In contrast, the travelers had IgG positive rates from 97.3% to 91.9% to various peptide epitopes. Based on the different distribution in IgM and IgG positivity to overall number of peptide epitopes and to the number of pre-erythrocytic, erythrocytic, gametocytic, and salivary stage epitopes at the individual level, four distinct patterns of IgM and IgG responses among the 37 samples from US travelers were observed. Independent peptide-bead coupling and antibody level readout between two different instruments also showed comparable results. Overall, this new coupling method resolves the peptide-bead coupling challenge, is reproducible, and can be applied to any other immunogenic peptide epitopes. The resulting all peptide bead-based multiplex Luminex assay can be expanded to include other peptide epitopes of P. falciparum, different malaria species, or other diseases for surveillance, either in US travelers or endemic areas. |
The molecular mechanism of induction of unfolded protein response by Chlamydia
George Z , Omosun Y , Azenabor AA , Goldstein J , Partin J , Joseph K , Ellerson D , He Q , Eko F , McDonald MA , Reed M , Svoboda P , Stuchlik O , Pohl J , Lutter E , Bandea C , Black CM , Igietseme JU . Biochem Biophys Res Commun 2019 508 (2) 421-429 The unfolded protein response (UPR) contributes to chlamydial pathogenesis, as a source of lipids and ATP during replication, and for establishing the initial anti-apoptotic state of host cell that ensures successful inclusion development. The molecular mechanism(s) of UPR induction by Chlamydia is unknown. Chlamydia use type III secretion system (T3SS) effector proteins (e.g, the Translocated Actin-Recruiting Phosphoprotein (Tarp) to stimulate host cell's cytoskeletal reorganization that facilitates invasion and inclusion development. We investigated the hypothesis that T3SS effector-mediated assembly of myosin-II complex produces activated non-muscle myosin heavy chain II (NMMHC-II), which then binds the UPR master regulator (BiP) and/or transducers to induce UPR. Our results revealed the interaction of the chlamydial effector proteins (CT228 and Tarp) with components of the myosin II complex and UPR regulator and transducer during infection. These interactions caused the activation and binding of NMMHC-II to BiP and IRE1alpha leading to UPR induction. In addition, specific inhibitors of myosin light chain kinase, Tarp oligomerization and myosin ATPase significantly reduced UPR activation and Chlamydia replication. Thus, Chlamydia induce UPR through T3SS effector-mediated activation of NMMHC-II components of the myosin complex to facilitate infectivity. The finding provides greater insights into chlamydial pathogenesis with the potential to identify therapeutic targets and formulations. |
Teaching population health: a competency map approach to education
Kaprielian VS , Silberberg M , McDonald MA , Koo D , Hull SK , Murphy G , Tran AN , Sheline BL , Halstater B , Martinez-Bianchi V , Weigle NJ , de Oliveira JS , Sangvai D , Copeland J , Tilson HH , Scutchfield FD , Michener JL . Acad Med 2013 88 (5) 626-37 A 2012 Institute of Medicine report is the latest in the growing number of calls to incorporate a population health approach in health professionals' training. Over the last decade, Duke University, particularly its Department of Community and Family Medicine, has been heavily involved with community partners in Durham, North Carolina, to improve the local community's health. On the basis of these initiatives, a group of interprofessional faculty began tackling the need to fill the curriculum gap to train future health professionals in public health practice, community engagement, critical thinking, and team skills to improve population health effectively in Durham and elsewhere. The Department of Community and Family Medicine has spent years in care delivery redesign and curriculum experimentation, design, and evaluation to distinguish the skills trainees and faculty need for population health improvement and to integrate them into educational programs. These clinical and educational experiences have led to a set of competencies that form an organizational framework for curricular planning and training. This framework delineates which learning objectives are appropriate and necessary for each learning level, from novice through expert, across multiple disciplines and domains. The resulting competency map has guided Duke's efforts to develop, implement, and assess training in population health for learners and faculty. In this article, the authors describe the competency map development process as well as examples of its application and evaluation at Duke and limitations to its use with the hope that other institutions will apply it in different settings. |
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