Natural IgM producing phagocytic B cells (NIMPABs) are a rare population of immune cells that can produce antibodies to broadly target and eliminate cancer cells via phosphatidylcholine (PtC)-specific phagocytosis. A novel, dual-labeled lipid-shelled superparamagnetic iron oxide nanoparticle (SPION)-based (SLNP) system was developed to trigger specific phagocytotic behavior in and subsequent enrichment of NIMPABs for a potential immunotherapy. Here we propose the design of an in vitro model to assess cell-SLNP interactions with J774A.1 monocyte cell line and the optimization of SLNP formulation. First, we developed and examined the morphology, size, concentration, purification, sterilization, and storage conditions of oleylamine-coated SPIONs and SLNPs using various microscopy methods, spectroscopy, dynamic light scattering, and zeta potential. Our data confirmed SPIONs are magnetic and 7-8 nm in diameter. SLNPs containing SPIONs retained the magnetic property, and are typically measured between 100-120 nm in diameter, and had a positive zeta potential. Fluorescence labeling of the SLNPs did not affect their properties. Second, we examined cytotoxicity and phagocytosis of SLNPs with J774A.1 cells. Our preliminary data showed that significant percentage of phagocytosis can be observed as early as 2 hours. However, longer than 2 hour incubation resulted in significant cytotoxic effects. The source of SLNP cytotoxicity was examined with transmission electron microscopy and characterization techniques, identifying high un-encapsulated free oleylamine-coated SPION content in SLNP samples contributing to a positive zeta potential for these samples. Simplified SLNPs with oleic acid-coated SPIONs were synthesized and resulting examined particles had a negative zeta potential, reduced free SPION content and improved SPION incorporation into SLNP cores. Based on these findings, SLNP criteria, characterization techniques, and cell assays were revised to establish a rigorous, standardized workflow essential for determining the optimal SLNP formulation. Future work must continue to modify SLNP formulation with information obtained from all characterization techniques and cellular assays outlined in the in vitro model. Once optimized, selective SLNP-mediated isolation of NIMPAB cells can be validated ex vivo with murine peritoneal cavity washout cells and then human peripheral blood samples. / 2024-08-26T00:00:00Z
Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/45056 |
Date | 26 August 2022 |
Creators | McPhillips, Marissa L. |
Contributors | Wong, Joyce Y., Ngo, John T . |
Source Sets | Boston University |
Language | en_US |
Detected Language | English |
Type | Thesis/Dissertation |
Page generated in 0.013 seconds