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Interleukin(IL)-4 is produced by T cells and other leukocytes and is a critical mediator of monocyte and B cell responses. During routine flow cytometry panel validation for the investigation of intracellular cytokines, we observed unique IL-4 expression patterns associated with the widely available monoclonal antibody 8D4-8. Namely, IL-4 (8D4-8) expression was observed in the absence of cellular activation and enhanced following staurosporine exposure. Mass spectrometry analysis of immunoprecipitates from peripheral blood lymphocytes (PBL) revealed that 8D4-8 cross-reacts with the ubiquitous cytoskeletal protein myosin-9. We confirmed these results by western blotting immunoprecipitates, using immunofluorescence among staurosporine-treated Caco-2 cells, and by surface-labeling PBL for 8D4-8 and myosin-9 and analyzing by flow cytometry. Although previously reported from several independent groups, we found no evidence to support the hypothesis that IL-4 is produced by apoptotic cells. Rather, this appears to have been myosin-9. Our data indicate clone 8D4-8 should not be used in the flow cytometric study of IL-4. Furthermore, our work calls for a reevaluation of previous flow cytometric studies that have used this clone for IL-4 analysis and highlights the importance of validation in antibody-based assays.

The Youth Enjoy Science program at the University of Nebraska Medical Center has engaged American Indian/Alaska Native youth in mentored cancer research internships from 2017 to 2022. The primary purpose of this study was to examine mentor and mentee lived experiences of participation in Youth Enjoy Science research education internships and to provide insights that can inform mentorship practices in research education programs for American Indians/Alaska Natives. We conducted semi-structured interviews with current and former Youth Enjoy Science mentees (n=8) and mentors (n=8). Following a narrative inquiry research approach, we analyzed interview transcripts and collectively re-storied interview data. Participants described program characters, settings, problems, actions to address the problems identified, and resolutions that led to various recommendations for ways to raise contextual awareness between mentees and mentors.