Microfluidic Squeezing Enables MHC Class I Antigen Presentation by Diverse Immune Cells to Elicit CD8+ T Cell Responses with Antitumor Activity

J Immunol. 2022 Feb 15;208(4):929-940. doi: 10.4049/jimmunol.2100656. Epub 2022 Jan 28.


CD8+ T cell responses are the foundation of the recent clinical success of immunotherapy in oncologic indications. Although checkpoint inhibitors have enhanced the activity of existing CD8+ T cell responses, therapeutic approaches to generate Ag-specific CD8+ T cell responses have had limited success. Here, we demonstrate that cytosolic delivery of Ag through microfluidic squeezing enables MHC class I presentation to CD8+ T cells by diverse cell types. In murine dendritic cells (DCs), squeezed DCs were ∼1000-fold more potent at eliciting CD8+ T cell responses than DCs cross-presenting the same amount of protein Ag. The approach also enabled engineering of less conventional APCs, such as T cells, for effective priming of CD8+ T cells in vitro and in vivo. Mixtures of immune cells, such as murine splenocytes, also elicited CD8+ T cell responses in vivo when squeezed with Ag. We demonstrate that squeezing enables effective MHC class I presentation by human DCs, T cells, B cells, and PBMCs and that, in clinical scale formats, the system can squeeze up to 2 billion cells per minute. Using the human papillomavirus 16 (HPV16) murine model, TC-1, we demonstrate that squeezed B cells, T cells, and unfractionated splenocytes elicit antitumor immunity and correlate with an influx of HPV-specific CD8+ T cells such that >80% of CD8s in the tumor were HPV specific. Together, these findings demonstrate the potential of cytosolic Ag delivery to drive robust CD8+ T cell responses and illustrate the potential for an autologous cell-based vaccine with minimal turnaround time for patients.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adoptive Transfer
  • Animals
  • Antigen Presentation*
  • Antigen-Presenting Cells / immunology*
  • Antigen-Presenting Cells / metabolism
  • Antigens, Neoplasm / immunology
  • CD8-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / metabolism
  • Cell Culture Techniques
  • Female
  • Histocompatibility Antigens Class I / immunology*
  • Humans
  • Immunization
  • Immunophenotyping
  • Leukocytes, Mononuclear / immunology
  • Leukocytes, Mononuclear / metabolism
  • Lymphocytes, Tumor-Infiltrating / immunology
  • Lymphocytes, Tumor-Infiltrating / metabolism
  • Mice
  • Mice, Knockout
  • Microfluidics* / methods
  • Models, Biological
  • Neoplasms / immunology*
  • Neoplasms / metabolism
  • Neoplasms / pathology
  • T-Lymphocyte Subsets / immunology
  • T-Lymphocyte Subsets / metabolism


  • Antigens, Neoplasm
  • Histocompatibility Antigens Class I