Abstract
Abstract The study of immune cell function in non-lymphoid tissue and tumors promises to elucidate novel strategies to treat immune disorders, infectious diseases, and cancer. To address the challenge of isolating leukocytes from complex and variable tissues and tumors, we have developed a new protocol to isolate particle-free, human CD45+ leukocytes. Using the EasySep™ Release Human CD45 Positive Selection Kit, leukocytes are labeled with antibody complexes linked to magnetic particles and separated using an EasySep™ magnet. The magnetic particles are then removed from the desired cells by resuspension in EasySep™ Release Buffer and a final magnetic separation. To assess performance, NRG-3GS mice were first engrafted with human CD34+ cells followed by xenotransplant with human breast (MDA-MB-231) or ovarian (SKOV3) cancer cell lines. In humanized mouse lungs, bone marrow and spleen, the starting and isolated human CD45+ frequency ranges were 6.0 – 57.2% and 90.9 – 99.4%, respectively (n = 3). Starting with human tumor xenografts, tumor infiltrating leukocytes were enriched from a starting range of 0.4 – 18.0% to 76.6 – 92.7% (n = 4). The final immune cell frequencies are representative of the starting population, and further separation of immune subsets can be achieved with additional downstream isolation. Humanized mouse models of clinical disease are instrumental in furthering our understanding of complex mechanisms of disease progression and resolution. This new kit for the isolation of human immune cells from tissues and tumors will facilitate further examination of the roles of immunity in disease and the evaluation of immune-based treatment strategies.
Published Version
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