Methods for Analysis of Nanoparticle Immunosuppressive Properties In Vitro and In Vivo

  • Timothy M. Potter
  • Barry W. Neun
  • Marina A. DobrovolskaiaEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1682)


Adverse drug effects on the immune system function represent a significant concern in the pharmaceutical industry, because 10–20% of the drug withdrawal from the market is accounted to immunotoxicity. Immunosuppression is one such adverse effect. The traditional immune function test used to estimate materials’ immunosuppression is a T-cell-dependent antibody response (TDAR). This method involves a 28 day in vivo study evaluating the animal’s antibody titer to a known antigen (KLH) with and without challenge. Due to the limited quantities of novel drug candidates, an in vitro method called human leukocyte activation (HuLa) assay has been developed to substitute the traditional TDAR assay during early preclinical development. In this test, leukocytes isolated from healthy donors vaccinated with the current year’s flu vaccine are incubated with Fluzone in the presence or absence of a test material. The antigen-specific leukocyte proliferation is then measured by ELISA analyzing incorporation of BrdU into DNA of the proliferating cells. Here, we describe the experimental procedures for investigating immunosuppressive properties of nanoparticles by both TDAR and HuLa assays, discuss the in vitro–in vivo correlation of these methods, and show a case study using the iron oxide nanoparticle formulation, Feraheme.

Key words

Nanoparticles Immunosuppression Leukocyte proliferation Antigen TDAR 



This project has been funded in whole or in part by federal funds from the National Cancer Institute, National Institutes of Health, under contract HHSN261200800001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government.


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Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  • Timothy M. Potter
    • 1
  • Barry W. Neun
    • 1
  • Marina A. Dobrovolskaia
    • 1
    Email author
  1. 1.Cancer Research Technology Program, Nanotechnology Characterization LaboratoryLeidos Biomedical Research, Inc., Frederick National Laboratory for Cancer ResearchFrederickUSA

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