Microneedle-Based Immune Monitoring Platform Samples Cells and Interstitial Fluid from Tissue in Situ


Anasuya Mandal


A. Mandal(1, 4), A. V. Boopathy(4), J. H. Van(2), P. T. Hammond(1,4,5) & D. J. Irvine(2,3,4,5,6,7)

Author Affiliation: 

"1 Department of Chemical Engineering, MIT 2 Department of Biological Engineering, MIT 3 Department of Materials Science and Engineering, MIT 4 Koch Institute for Integrative Cancer Research, MIT 5 Institute for Soldier Nanotechnologies, MIT 6 Ragon Institute of MGH, MIT, and Harvard 7 Howard Hughes Medical Institute, MD, USA"


"Current protocols for immune system monitoring involve the collection of cells from blood or cerebrospinal fluid. However, since major populations of immune cells reside within tissues, these invasively-obtained body fluid samples are, at best, indirect indicators of the status of the immune system. Direct tissue sampling through biopsies is difficult to incorporate into long-term, repetitive, longitudinal immune monitoring. Delayed-type hypersensitivity tests (e.g., Mantoux tuberculin test) query the presence of antigen-specific cells in the skin, but do not provide information about the phenotype and functional characteristics of responding immune cells. Here we report a technology that addresses several of these challenges simultaneously, with the synergistic goals of providing enhanced diagnostic methods for sampling and analyzing the function of the immune system, and providing a greater insight into the status of the immune system than state of the art assays. We fabricated hydrogel-coated microneedles capable of sampling both cells and interstitial fluid from tissues, permitting the quantification of constituents like antigen-specific IgG. Previous work from our group has focused on transdermal delivery of vaccines using microneedles in a variety of different approaches, using murine and non-human primate models. Complementary to recent reports using microneedles for glucose monitoring and circulating biomarker detection, we report here for the first time, microneedles enabling parallel monitoring of both cellular and humoral immune responses. Microneedles were fabricated by coating the surface of poly-L-lactide solid polymer microneedles with alginate solutions containing antigen and/or adjuvant, followed by crosslinking of the polysaccharide layer with calcium. Incorporation of chemoattractants or adjuvants in the alginate coating of the microneedles promoted immune cell recruitment into the alginate layers of microneedles within 8 hours of application, as evidenced via confocal microscopy. In preliminary studies, using a subcutaneous alginate gel injection model in C57Bl/6 mice, we determined that 48 hours was the optimal time of sampling, using a chemoattractant dose of 2 - 4µg. Injected alginate gels showed enhanced infiltration of CD4+ T cells under the influence of inflammatory chemoattractants (CCL21 and CXCL10) and showed increasing infiltration of cells with time, over 48 hours. We then applied these optimized conditions to sample cells from skin using alginate-coated microneedles. Cells were released from retrieved microneedle arrays by dissolving the alginate layer in the presence of EDTA, enabling subsequent phenotypic analysis through antibody staining and flow or imaging cytometry. Using mice immunized with the model antigen, ovalbumin, we demonstrated that sampling microneedles, applied for 12 hours following an intradermal injection of antigen and adjuvants 48 hours prior, sampled antigen-specific CD8+ cells at the injection site, at levels comparable to blood. We were also able to reliably quantitate antigen-specific IgG from the interstitial fluid collected on the same microneedle patch. This approach allows ex vivo analysis of cells retrieved directly from the local tissue environment and enables the investigation of antigen-specific cells for diagnostic purposes as well as answering spatio-temporal questions related to immunology in local tissue environments. This simple and non-invasive sampling approach should facilitate longitudinal monitoring of antigen-specific immune cell populations in the skin relevant for a variety of infectious and autoimmune diseases such as lupus."