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@ARTICLE{Hazin:126676,
      author       = {J. Hazin$^*$ and G. Moldenhauer$^*$ and P. Altevogt$^*$ and
                      N. R. Brady$^*$},
      title        = {{A} novel method for measuring cellular antibody uptake
                      using imaging flow cytometry reveals distinct uptake rates
                      for two different monoclonal antibodies targeting {L}1.},
      journal      = {Journal of immunological methods},
      volume       = {423},
      issn         = {0022-1759},
      address      = {Amsterdam [u.a.]},
      publisher    = {Elsevier Science},
      reportid     = {DKFZ-2017-02704},
      pages        = {70 - 77},
      year         = {2015},
      abstract     = {Monoclonal antibodies (mAbs) have emerged as a promising
                      tool for cancer therapy. Differing approaches utilize mAbs
                      to either deliver a drug to the tumor cells or to modulate
                      the host's immune system to mediate tumor kill. The rate by
                      which a therapeutic antibody is being internalized by tumor
                      cells is a decisive feature for choosing the appropriate
                      treatment strategy. We herein present a novel method to
                      effectively quantitate antibody uptake of tumor cells by
                      using image-based flow cytometry, which combines image
                      analysis with high throughput of sample numbers and sample
                      size. The use of this method is established by determining
                      uptake rate of an anti-EpCAM antibody (HEA125), from single
                      cell measurements of plasma membrane versus internalized
                      antibody, in conjunction with inhibitors of endocytosis. The
                      method is then applied to two mAbs (L1-9.3, L1-OV52.24)
                      targeting the neural cell adhesion molecule L1 (L1CAM) at
                      two different epitopes. Based on median cell population
                      responses, we find that mAb L1-OV52.24 is rapidly
                      internalized by the ovarian carcinoma cell line SKOV3ip
                      while L1 mAb 9.3 is mainly retained at the cell surface.
                      These findings suggest the L1 mAb OV52.24 as a candidate to
                      be further developed for drug-delivery to cancer cells,
                      while L1-9.3 may be optimized to tag the tumor cells and
                      stimulate immunogenic cancer cell killing. Furthermore, when
                      analyzing cell-to-cell variability, we observed L1 mAb
                      OV52.24 rapidly transition into a subpopulation with
                      high-internalization capacity. In summary, this novel
                      high-content method for measuring antibody internalization
                      rate provides a high level of accuracy and sensitivity for
                      cell population measurements and reveals further
                      biologically relevant information when taking into account
                      cellular heterogeneity.},
      keywords     = {Antibodies, Monoclonal (NLM Chemicals) / Antigens, Neoplasm
                      (NLM Chemicals) / CADM1 protein, human (NLM Chemicals) /
                      Cell Adhesion Molecules (NLM Chemicals) / EPCAM protein,
                      human (NLM Chemicals) / Epithelial Cell Adhesion Molecule
                      (NLM Chemicals) / Immunoglobulins (NLM Chemicals)},
      cin          = {D015 / B170},
      ddc          = {610},
      cid          = {I:(DE-He78)D015-20160331 / I:(DE-He78)B170-20160331},
      pnm          = {312 - Functional and structural genomics (POF3-312)},
      pid          = {G:(DE-HGF)POF3-312},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:25967950},
      doi          = {10.1016/j.jim.2015.04.024},
      url          = {https://inrepo02.dkfz.de/record/126676},
}