Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Protocol
  • Published:

Radiolabeling of DOTA-like conjugated peptides with generator-produced 68Ga and using NaCl-based cationic elution method

Nature Protocols volume 11, pages 1057–1066 (2016)Cite this article

Subjects

Abstract

Gallium-68 (68Ga) is a generator-produced radionuclide with a short half-life (t½ = 68 min) that is particularly well suited for molecular imaging by positron emission tomography (PET). Methods have been developed to synthesize 68Ga-labeled imaging agents possessing certain drawbacks, such as longer synthesis time because of a required final purification step, the use of organic solvents or concentrated hydrochloric acid (HCl). In our manuscript, we provide a detailed protocol for the use of an advantageous sodium chloride (NaCl)-based method for radiolabeling of chelator-modified peptides for molecular imaging. By working in a lead-shielded hot-cell system,68Ga3+ of the generator eluate is trapped on a cation exchanger cartridge (100 mg, ∼8 mm long and 5 mm diameter) and then eluted with acidified 5 M NaCl solution directly into a sodium acetate-buffered solution containing a DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) or DOTA-like chelator-modified peptide. The main advantages of this procedure are the high efficiency and the absence of organic solvents. It can be applied to a variety of peptides, which are stable in 1 M NaCl solution at a pH value of 3–4 during reaction. After labeling, neutralization, sterile filtration and quality control (instant thin-layer chromatography (iTLC), HPLC and pH), the radiopharmaceutical can be directly administered to patients, without determination of organic solvents, which reduces the overall synthesis-to-release time. This procedure has been adapted easily to automated synthesis modules, which leads to a rapid preparation of 68Ga radiopharmaceuticals (12–16 min).

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3: An example of a radio-iTLC chromatogram of [68Ga]-DOTATOC using the iTLC method described in Box 2 (mobile phase consisting of 1 M ammonium acetate/methanol 1:1 vol/vol).
Figure 4
Figure 5: Elution profile of the 68Ge/68Ga generator and formation of 68Ga.
Figure 6: Progress of 68Ga activity during one automated synthesis run for the routine pharmaceutical production of 68Ga-DOTATOC—synthesis time: 14 min, time until application: 20 min (vertical black line), start activity: 1.85 GBq, activity after synthesis: 1.20 GBq, applicable activity for patients: 1.13 GBq (horizontal black line).
Figure 7: Progress of activity of sequentially performed synthesis runs for the routine pharmaceutical production of 68Ga-DOTATOC—first start activity: 1.85 GBq, activity after first synthesis: 1.20 GBq, first applicable activity (20 min, vertical black line): 1.13 GBq (horizontal black line), second start activity: 1.31 GBq, activity after second synthesis: 0.85 GBq, second applicable activity (140 min, vertical black line): 0.80 GBq (horizontal black line).
Figure 8: Decay of 68Ga and formation of 68Zn.

Similar content being viewed by others

References

  1. Menda, Y. et al. Repeatability of gallium-68 DOTATOC positron emission tomographic imaging in neuroendocrine Tumors. Pancreas 42, 937–943 (2013).

    Article  CAS  Google Scholar 

  2. Production of Long Lived Parent Radionuclides for Generators, 68Ge, 92Sr, 90Sr and 188W. (International Atomic Energy Agency, 2010) http://www-pub.iaea.org/MTCD/publications/PDF/Pub1436_web.pdf.

  3. Velikyan, I. Prospective of 68Ga-radiopharmaceutical development. Theranostics 4, 47–80 (2014).

    Article  CAS  Google Scholar 

  4. de Blois, E. et al. Characteristics of SnO2-based 68Ge/68Ga generator and aspects of radiolabelling DOTA-peptides. Appl. Radiat. Isot. 69, 308–315 (2011).

    Article  CAS  Google Scholar 

  5. Meyer, G.J., Mäcke, H., Schuhmacher, J., Knapp, W.H. & Hofmann, M. 68Ga labelled DOTA-derivatised peptide ligands. Eur. J. Nucl. Med. Mol. Imaging 31, 1097–1104 (2004).

    Article  CAS  Google Scholar 

  6. Eder, M. et al. Novel preclinical and radiopharmaceutical aspects of [68Ga]Ga-PSMA-HBED-CC: A new PET tracer for imaging of prostate cancer. Pharmaceuticals 7, 779–796 (2014).

    Article  CAS  Google Scholar 

  7. Zhernosekov, K.P. et al. Processing of generator produced 68Ga for medical application. J. Nucl. Med. 48, 1741–1748 (2007).

    Article  CAS  Google Scholar 

  8. Roesch, F. Post-processing via cation exchange cartridges: versatile options. in Theranostics, Gallium-68 and Other Radionuclides. Recent Results in Cancer Research Vol. 194 (eds. Baum, R.P. and Roesch, F.) 33–42 (Springer-Verlag, 2012).

  9. Eppard, E., Wuttke, M., Nicodemus, P.L. & Rösch, F. Ethanol-based post-processing of generator-derived 68Ga toward kit-type preparation of 68Ga-radiopharmaceuticals. J. Nucl. Med. 55, 1023–1028 (2014).

    Article  CAS  Google Scholar 

  10. Seemann, J., Eppard, E., Waldron, B.P., Ross, T.L. & Roesch, F. Cation exchange-based post-processing of 68Ga-eluate: a comparison of three solvent systems for labelling of DOTATOC, NO2AP(BP) and DATA(m.). Appl. Radiat. Isot. 98, 54–59 (2015).

    Article  CAS  Google Scholar 

  11. Müller, D., Klette, I. & Baum, R.P. The combined cationic-anionic purification of the 68Ge/68Ga generator eluate for the labeling of fragile peptides. World J. Nucl. Med. 10, 77–78 (2011).

    Google Scholar 

  12. Loktionova, N.S. et al. Improved column-based radiochemical processing of the generator produced 68Ga. Appl. Radiat. Isot. 69, 942–946 (2011).

    Article  CAS  Google Scholar 

  13. Mueller, D. et al. Simplified NaCl based 68Ga concentration and labeling procedure for rapid synthesis of 68Ga radiopharmaceuticals in high radiochemical purity. Bioconjug. Chem. 23, 1712–1717 (2012).

    Article  CAS  Google Scholar 

  14. Schultz, M.K., Mueller, D., Baum, R.P., Leonard Watkins, G. & Breeman, W.A.P. A new automated NaCl based robust method for routine production of gallium-68 labeled peptides. Appl. Radiat. Isot. 76, 46–54 (2013).

    Article  CAS  Google Scholar 

  15. Maina, T. et al. Preclinical and first clinical experience with the gastrin-releasing peptide receptor-antagonist [68Ga]SB3 and PET/CT. Eur. J. Nucl. 43, 964–973 (2016).

    Article  CAS  Google Scholar 

  16. Baum, R.P. et al. First-in-human study demonstrating tumor-angiogenesis by PET/CT imaging with Ga-68-NODAGA-THERANOST, a high-affinity peptidomimetic for alpha(v)beta(3) integrin receptor targeting. Cancer Biother. Radiopharm. 30, 152–159 (2015).

    Article  CAS  Google Scholar 

  17. Miller, S.A., Dykes, D.D. & Polesky, H.F. A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acids Res. 16, 1215 (1988).

    Article  CAS  Google Scholar 

  18. Mao, Y.J., Sheng, X.R. & Pan, X.M. The effects of NaCl concentration and pH on the stability of hyperthermophilic protein Ssh10b. BMC Biochem. 8, 28 (2007).

    Article  Google Scholar 

  19. Dominy, B.N., Perl, D., Schmid, F.X. & Brooks, C.L. The effects of ionic strength on protein stability: the cold shock protein family. J. Mol. Biol. 319, 541–554 (2002).

    Article  CAS  Google Scholar 

  20. Lindman, S. et al. Salting the charged surface: pH and salt dependence of protein G B1 stability. Biophys. J. 90, 2911–2921 (2006).

    Article  CAS  Google Scholar 

  21. Šimeček, J., Hermann, P., Wester, H.J. & Notni, J. How is 68Ga labeling of macrocyclic chelators influenced by metal ion contaminants in 68Ge/68Ga generator eluates? ChemMedChem 8, 95–103 (2013).

    Article  Google Scholar 

  22. Chakravarty, R., Chakraborty, S., Dash, A. & Pillai, M.R. Detailed evaluation on the effect of metal ion impurities on complexation of generator eluted 68Ga with different bifunctional chelators. Nucl. Med. Biol. 40, 197–205 (2013).

    Article  CAS  Google Scholar 

  23. '2.4.8 Heavy Metals'. European Pharmacopoeia 8th edn. 128–131 (Council of Europe, 2010).

Download references

Author information

Authors and Affiliations

  1. Department of Nuclear Medicine, University Hospital Halle (Saale), Halle, Germany

    Dirk Mueller, Andreas Odparlik & Manfred Baehre

  2. Department of Nuclear Medicine, Erasmus MC Rotterdam, Rotterdam, the Netherlands

    Wouter A P Breeman

  3. Department of Nuclear Medicine/PET Center, Zentralklinik Bad Berka, Bad Berka, Germany

    Ingo Klette

  4. Friedrich Schiller University of Jena, Institute of Organic and Macromolecular Chemistry, Jena, Germany

    Michael Gottschaldt

  5. Jena Center for Soft Matter (JCSM), Jena, Germany

    Michael Gottschaldt

  6. RadioMedix, Inc., Houston, Texas, USA

    Izabela Tworowska

  7. Departments of Radiology and Radiation Oncology (Free Radical Radiation Biology Program), University of Iowa, Iowa City, Iowa, USA

    Michael K Schultz

Authors
  1. Dirk Mueller
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wouter A P Breeman
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ingo Klette
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Michael Gottschaldt
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Andreas Odparlik
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Manfred Baehre
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Izabela Tworowska
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Michael K Schultz
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

D.M. conceived and developed the original NaCl-based method; W.A.P.B. and I.K. revalidated the procedures and optimized the conditions; M.G. suggested suitable cation-exchange resigns and the NaCl elution from SCX cartriges; M.B. and A.O. evaluated the method from medical point of view; and I.T. and M.K.S. tested the procedure on different generators. All authors wrote and edited the manuscript.

Corresponding author

Correspondence to Dirk Mueller.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Scheme 1

Software interface module diagram of the automated radiosynthesis system (ModularLab PharmTracer, Eckert & Ziegler) used to prepare 68Ga DOTATOC by the NaCl based method.

Supplementary information

Supplementary Figure

Supplementary Figure 1 (PDF 233 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Mueller, D., Breeman, W., Klette, I. et al. Radiolabeling of DOTA-like conjugated peptides with generator-produced 68Ga and using NaCl-based cationic elution method. Nat Protoc 11, 1057–1066 (2016). https://doi.org/10.1038/nprot.2016.060

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nprot.2016.060

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing