Analytical and Bioanalytical Chemistry

, Volume 411, Issue 6, pp 1253–1260 | Cite as

Quantitative imaging of 33P in plant materials using 14C polymer references

  • Maximilian KochEmail author
  • Henning Schiedung
  • Nina Siebers
  • Sean McGovern
  • Diana Hofmann
  • Harry Vereecken
  • Wulf Amelung
Research Paper


Phosphorus (P) research still lacks techniques for rapid imaging of P use and allocation in different soil, sediment, and biological systems in a quantitative manner. In this study, we describe a time-saving and cost-efficient digital autoradiographic method for in situ quantitative imaging of 33P radioisotopes in plant materials. Our method combines autoradiography of the radiotracer applications with additions of commercially available 14C polymer references to obtain 33P activities in a quantitative manner up to 2000 Bq cm−2. Our data show that linear standard regressions for both radioisotopes are obtained, allowing the establishment of photostimulated luminescence equivalence between both radioisotopes with a factor of 9.73. Validating experiments revealed a good agreement between the calculated and applied 33P activity (R2 = 0.96). This finding was also valid for the co-exposure of 14C polymer references and 33P radioisotope specific activities in excised plant leaves for both maize (R2 = 0.99) and wheat (R2 = 0.99). The outlined autoradiographic quantification procedure retrieved 100% ± 12% of the 33P activity in the plant leaves, irrespective of plant tissue density. The simplicity of this methodology opens up new perspectives for fast quantitative imaging of 33P in biological systems and likely, thus, also for other environmental compartments.


33P phosphoric acid Digital autoradiography Radioluminography Bioimaging Maize Wheat 



The authors gratefully acknowledge the German Federal Ministry of Education and Research (BMBF) for funding the BonaRes project InnoSoilPhos [grant number 031A558]. In addition, this study was also conducted under the scope of the NRW-Strategieprojekt BioSC AlgalFertilizer. We thank P. Narf and we also gratefully thank M. Krause and A. Kubica of the Institute of Bio- and Geosciences – Agrosphere (IBG-3) at Forschungszentrum Jülich GmbH for their practical support.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

216_2018_1557_MOESM1_ESM.pdf (495 kb)
ESM 1 (PDF 494 kb)


  1. 1.
    McLaughlin M, Alston A, Martin J. Phosphorus cycling in wheat pasture rotations .II. The role of the microbial biomass in phosphorus cycling. Soil Res. 1988;26(2):333–42. Scholar
  2. 2.
    McLaren TI, McLaughlin MJ, McBeath TM, Simpson RJ, Smernik RJ, Guppy CN, et al. The fate of fertiliser P in soil under pasture and uptake by subterraneum clover – a field study using 33P-labelled single superphosphate. Plant Soil. 2016;401(1):23–38. Scholar
  3. 3.
    McLaughlin M, Alston A, Martin J. Phosphorus cycling in wheat pasture rotations .I. The source of phosphorus taken up by wheat. Soil Res. 1988;26(2):323–31. Scholar
  4. 4.
    Johnston RF, Pickett SC, Barker DL. Autoradiography using storage phosphor technology. Electrophoresis. 1990;11(5):355–60. Scholar
  5. 5.
    Nakajima E. CRC handbook of chromatography: analysis of lipids, Imaging-plate systems for radioluminographic detection of lipids on thin-layer plates. London: CRC Press; 1993. Scholar
  6. 6.
    Amemiya Y, Miyahara J. Imaging plate illuminates many fields. Nature. 1988;336(6194):89–90. Scholar
  7. 7.
    Takahashi K, Miyahara J, Shibahara Y. Photostimulated luminescence (PSL) and color centers in BaFX : Eu2 + (X = Cl , Br , I) phosphors. J Electrochem Soc. 1985;132(6):1492–4. Scholar
  8. 8.
    Shinde KN, Dhoble SJ, Swart HC, Park K. Basic mechanisms of photoluminescence. In: Phosphate phosphors for solid-state lighting. Berlin: Springer; 2012. p. 41–59. Scholar
  9. 9.
    Upham LV, Englert DF. 13 - RADIONUCLIDE IMAGING A2 – L’Annunziata, Michael F. In: Handbook of radioactivity analysis. 2nd ed. San Diego: Academic Press; 2003. p. 1063–127. Scholar
  10. 10.
    Hüve K, Merbach W, Remus R, Lüttschwager D, Wittenmayer L, Hertel K, et al. Transport of phosphorus in leaf veins of Vicia faba L. J Plant Nutr Soil Sci. 2007;170(1):14–23. Scholar
  11. 11.
    Bauke SL, Landl M, Koch M, Hofmann D, Nagel KA, Siebers N, et al. Macropore effects on phosphorus acquisition by wheat roots – a rhizotron study. Plant Soil. 2017:1–16.
  12. 12.
    Cremer CM, Cremer M, Escobar JL, Speckmann E-J, Zilles K. Fast, quantitative in situ hybridization of rare mRNAs using 14C-standards and phosphorus imaging. J Neurosci Methods. 2009;185(1):56–61. Scholar
  13. 13.
    Eakin TJ, Baskin DG, Breininger JF, Stahl WL. Calibration of 14C-plastic standards for quantitative autoradiography with 33P. J Histochem Cytochem. 1994;42(9):1295–8. Scholar
  14. 14.
    Baskin DG, Stahl WL. Fundamentals of quantitative autoradiography by computer densitometry for in situ hybridization, with emphasis on 33P. J Histochem Cytochem. 1993;41(12):1767–76. Scholar
  15. 15.
    Robu E, Giovani C. Gamma-ray self-attenuation corrections in environmental samples. Romanian Rep Phys. 2009;61(2):295–300. Scholar
  16. 16.
    Reichert WL, Stein JE, French B, Goodwin P, Varanasi U. Storage phosphor imaging technique for detection and quantitation of DNA adducts measured by the 32P-postlabeling assay. Carcinogenesis. 1992;13(8):1475–9. Scholar
  17. 17.
    Takahashi K. Phosphor for X-ray and ionizing radiation. In: Yen WM, Shionoya S, Yamamoto H, editors. Practical applications of phosphors: CRC Press; 2006. p. 319–25.Google Scholar
  18. 18.
    Takahashi K, Kohda K, Miyahara J, Kanemitsu Y, Amitani K, Shionoya S. Mechanism of photostimulated luminescence in BaFX:Eu2+ (X=Cl,Br) phosphors. J Lumin. 1984;31–32(Part 1):266–8. Scholar
  19. 19.
    L’Annunziata MF. Chapter 1 - radiation physics and radionuclide decay. In: L’Annunziata MF, editor. Handbook of radioactivity analysis. 3rd ed. Amsterdam: Academic Press; 2012. p. 1–162. Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Maximilian Koch
    • 1
    Email author
  • Henning Schiedung
    • 2
  • Nina Siebers
    • 1
  • Sean McGovern
    • 1
  • Diana Hofmann
    • 1
  • Harry Vereecken
    • 1
  • Wulf Amelung
    • 1
    • 2
  1. 1.Agrosphere, Forschungszentrum Jülich GmbHInstitute for Bio- and Geosciences – IBG-3JülichGermany
  2. 2.Institute of Crop Science and Resource Conservation – Soil Science and Soil EcologyUniversity of BonnBonnGermany

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