Inert coupling of IRDye800CW and zirconium-89 to monoclonal antibodies for single- or dual-mode fluorescence and PET imaging

Nat Protoc. 2013 May;8(5):1010-8. doi: 10.1038/nprot.2013.054. Epub 2013 Apr 25.


IRDye800CW and zirconium-89 ((89)Zr) have very attractive properties for optical imaging and positron emission tomography (PET) imaging, respectively. Here we describe a procedure for dual labeling of mAbs with IRDye800CW and (89)Zr in a current good manufacturing practice (cGMP)-compliant way. IRDye800CW and (89)Zr are coupled inertly, without impairment of immunoreactivity and pharmacokinetics of the mAb. Organ and whole-body distribution of the final product can be assessed by optical and PET imaging, respectively. For this purpose, a minimal amount of the chelate N-succinyldesferrioxamine (N-sucDf) is first conjugated to the mAb. Next, N-sucDf-mAb is conjugated with IRDye800CW, after which the N-sucDf-mAb-IRDye800CW is labeled with (89)Zr. After each of these three steps, the product is purified by gel filtration. The sequence of this process avoids unnecessary radiation exposure to personnel and takes about 5 h. The process can be scaled up by the production of large batches of premodified mAbs that can be dispensed and stored until they are labeled with (89)Zr.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal / analysis*
  • Benzenesulfonates / chemistry*
  • Indoles / chemistry*
  • Mice
  • Mice, Nude
  • Microscopy, Fluorescence / methods*
  • Positron-Emission Tomography / methods*
  • Staining and Labeling
  • Zirconium / chemistry*


  • Antibodies, Monoclonal
  • Benzenesulfonates
  • IRDye 800CW
  • Indoles
  • Zirconium