Analytical and Bioanalytical Chemistry

, Volume 407, Issue 8, pp 2301–2309 | Cite as

Subcellular-level resolution MALDI-MS imaging of maize leaf metabolites by MALDI-linear ion trap-Orbitrap mass spectrometer

  • Andrew R. Korte
  • Marna D. Yandeau-Nelson
  • Basil J. Nikolau
  • Young Jin LeeEmail author
Research Paper
Part of the following topical collections:
  1. Mass Spectrometry Imaging


A significant limiting factor in achieving high spatial resolution for matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) imaging is the size of the laser spot at the sample surface. Here, we present modifications to the beam-delivery optics of a commercial MALDI-linear ion trap-Orbitrap instrument, incorporating an external Nd:YAG laser, beam-shaping optics, and an aspheric focusing lens, to reduce the minimum laser spot size from ~50 μm for the commercial configuration down to ~9 μm for the modified configuration. This improved system was applied for MALDI-MS imaging of cross sections of juvenile maize leaves at 5-μm spatial resolution using an oversampling method. A variety of different metabolites including amino acids, glycerolipids, and defense-related compounds were imaged at a spatial resolution well below the size of a single cell. Such images provide unprecedented insights into the metabolism associated with the different tissue types of the maize leaf, which is known to asymmetrically distribute the reactions of C4 photosynthesis among the mesophyll and bundle sheath cell types. The metabolite ion images correlate with the optical images that reveal the structures of the different tissues, and previously known and newly revealed asymmetric metabolic features are observed.


Laser optics modification for subcellular-level MS imaging of maize leaf

Open image in new window


Mass spectrometry imaging Metabolite Maize 



This work was supported by the US Department of Energy (DOE), Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences, and Biosciences. MDY-N acknowledges the support of the National Science Foundation under Award No. EEC-0813570 and Award No. IOS-1354799, which co-sponsored the development of the genetic stocks imaged in this study. The Ames Laboratory is operated by Iowa State University under DOE Contract DE-AC02-07CH11358.

Supplementary material

216_2015_8460_MOESM1_ESM.pdf (1.4 mb)
ESM 1 (PDF 1000 kb)


  1. 1.
    Stoeckli M, Staab D, Schweitzer A (2007) Compound and metabolite distribution measured by MALDI mass spectrometric imaging in whole-body tissue sections. Int J Mass Spectrom 260(2–3):195–202. doi: 10.1016/j.ijms.2006.10.007 CrossRefGoogle Scholar
  2. 2.
    Hankin JA, Barkley RM, Murphy RC (2007) Sublimation as a method of matrix application for mass spectrometric imaging. J Am Soc Mass Spectrom 18(9):1646–1652. doi: 10.1016/j.jasms.2007.06.010 CrossRefGoogle Scholar
  3. 3.
    Jun JH, Song Z, Liu Z, Nikolau BJ, Yeung ES, Lee YJ (2010) High-spatial and high-mass resolution imaging of surface metabolites of Arabidopsis thaliana by laser desorption-ionization mass spectrometry using colloidal silver. Anal Chem 82(8):3255–3265. doi: 10.1021/ac902990p CrossRefGoogle Scholar
  4. 4.
    Chen Y, Allegood J, Liu Y, Wang E, Cachon-Gonzalez B, Cox TM, Merrill AH Jr, Sullards MC (2008) Imaging MALDI mass spectrometry using an oscillating capillary nebulizer matrix coating system and its application to analysis of lipids in brain from a mouse model of Tay-Sachs/Sandhoff disease. Anal Chem 80(8):2780–2788. doi: 10.1021/ac702350g CrossRefGoogle Scholar
  5. 5.
    Makarov A, Denisov E (2009) Dynamics of ions of intact proteins in the Orbitrap mass analyzer. J Am Soc Mass Spectrom 20(8):1486–1495. doi: 10.1016/j.jasms.2009.03.024 CrossRefGoogle Scholar
  6. 6.
    Chughtai K, Heeren RM (2010) Mass spectrometric imaging for biomedical tissue analysis. Chem Rev 110(5):3237–3277. doi: 10.1021/cr100012c CrossRefGoogle Scholar
  7. 7.
    Soltwisch J, Goeritz G, Jungmann JH, Kiss A, Smith DF, Ellis SR, Heeren RMA (2014) MALDI mass spectrometry imaging in microscope mode with infrared lasers: bypassing the diffraction limits. Anal Chem 86(1):321–325. doi: 10.1021/ac403421v CrossRefGoogle Scholar
  8. 8.
    Spengler B, Hubert M (2002) Scanning microprobe matrix-assisted laser desorption ionization (SMALDI) mass spectrometry: instrumentation for sub-micrometer resolved LDI and MALDI surface analysis. J Am Soc Mass Spectrom 13(6):735–748. doi: 10.1016/s1044-0305(02)00376-8 CrossRefGoogle Scholar
  9. 9.
    Roempp A, Guenther S, Schober Y, Schulz O, Takats Z, Kummer W, Spengler B (2010) Histology by mass spectrometry: label-free tissue characterization obtained from high-accuracy bioanalytical imaging. Angew Chem Int Ed 49(22):3834–3838. doi: 10.1002/anie.200905559 CrossRefGoogle Scholar
  10. 10.
    Hoelscher D, Shroff R, Knop K, Gottschaldt M, Crecelius A, Schneider B, Heckel DG, Schubert US, Svatos A (2009) Matrix-free UV-laser desorption/ionization (LDI) mass spectrometric imaging at the single-cell level: distribution of secondary metabolites of Arabidopsis thaliana and Hypericum species. Plant J 60(5):907–918. doi: 10.1111/j.1365-313X.2009.04012.x CrossRefGoogle Scholar
  11. 11.
    Zavalin A, Yang J, Caprioli R (2013) Laser beam filtration for high spatial resolution MALDI imaging mass spectrometry. J Am Soc Mass Spectrom 24(7):1153–1156. doi: 10.1007/s13361-013-0638-5 CrossRefGoogle Scholar
  12. 12.
    Zavalin A, Yang J, Haase A, Holle A, Caprioli R (2014) Implementation of a Gaussian beam laser and aspheric optics for high spatial resolution MALDI imaging MS. J Am Soc Mass Spectrom 25(6):1079–1082. doi: 10.1007/s13361-014-0872-5 CrossRefGoogle Scholar
  13. 13.
    Kettling H, Vens-Cappell S, Soltwisch J, Pirkl A, Haier J, Muething J, Dreisewerd K (2014) MALDI mass spectrometry imaging of bioactive lipids in mouse brain with a Synapt G2-S mass spectrometer operated at elevated pressure: improving the analytical sensitivity and the lateral resolution to ten micrometers. Anal Chem 86(15):7798–7805. doi: 10.1021/ac5017248 CrossRefGoogle Scholar
  14. 14.
    Thiery-Lavenant G, Zavalin AI, Caprioli RM (2013) Targeted multiplex imaging mass spectrometry in transmission geometry for subcellular spatial resolution. J Am Soc Mass Spectrom 24(4):609–614. doi: 10.1007/s13361-012-0563-z CrossRefGoogle Scholar
  15. 15.
    Zavalin A, Todd EM, Rawhouser PD, Yang J, Norris JL, Caprioli RM (2012) Direct imaging of single cells and tissue at sub-cellular spatial resolution using transmission geometry MALDI MS. J Mass Spectrom 47(11):1473–1481. doi: 10.1002/jms.3108 CrossRefGoogle Scholar
  16. 16.
    Jurchen JC, Rubakhin SS, Sweedler JV (2005) MALDI-MS imaging of features smaller than the size of the laser beam. J Am Soc Mass Spectrom 16(10):1654–1659. doi: 10.1016/j.jasms.2005.06.006 CrossRefGoogle Scholar
  17. 17.
    Perrot-Rechenmann C, Joannes M, Squalli D, Lebacq P (1989) Detection of phosphoenolpyruvate and ribulose 1,5-bisphosphate carboxylase transcripts in maize leaves by in situ hybridization with sulfonated cDNA probes. J Histochem Cytochem 37(4):423–428CrossRefGoogle Scholar
  18. 18.
    Majeran W, Cai Y, Sun Q, van Wijk KJ (2005) Functional differentiation of bundle sheath and mesophyll maize chloroplasts determined by comparative proteomics. Plant Cell Online 17(11):3111–3140. doi: 10.1105/tpc.105.035519 CrossRefGoogle Scholar
  19. 19.
    Korte AR, Lee YJ (2014) MALDI-MS analysis and imaging of low molecular weight metabolites with 1,5-diaminonaphthalene (DAN). J Mass Spectrom 49(8):737–741CrossRefGoogle Scholar
  20. 20.
    Korte A, Yagnik G, Feenstra A, Lee Y (2015) Multiplex MALDI-MS imaging of plant metabolites using a hybrid MS system. In: He L (ed) Mass spectrometry imaging of small molecules, vol 1203, Methods in Molecular Biology. Springer, New York, pp 49–62. doi: 10.1007/978-1-4939-1357-2_6 CrossRefGoogle Scholar
  21. 21.
    Strupat K, Kovtoun V, Bui H, Viner R, Stafford G, Horning S (2009) MALDI produced ions inspected with a linear ion trap-Orbitrap hybrid mass analyzer. J Am Soc Mass Spectrom 20(8):1451–1463. doi: 10.1016/j.jasms.2009.04.013 CrossRefGoogle Scholar
  22. 22.
    Lundgren MR, Osborne CP, Christin P-A (2014) Deconstructing Kranz anatomy to understand C4 evolution. J Exp Bot. doi: 10.1093/jxb/eru186 Google Scholar
  23. 23.
    Fabre N, Rustan I, de Hoffmann E, Quetin-Leclercq J (2001) Determination of flavone, flavonol, and flavanone aglycones by negative ion liquid chromatography electrospray ion trap mass spectrometry. J Am Soc Mass Spectrom 12(6):707–715. doi: 10.1016/s1044-0305(01)00226-4 CrossRefGoogle Scholar
  24. 24.
    Turgeon R (2006) Phloem loading: how leaves gain their independence. Bioscience 56(1):15–24. doi: 10.1641/0006-3568(2006)056[0015:plhlgt];2 CrossRefGoogle Scholar
  25. 25.
    Fraser CM, Chapple C (2011) The phenylpropanoid pathway in Arabidopsis. Arabidopsis Book 9:e0152. doi: 10.1199/tab.0152 CrossRefGoogle Scholar
  26. 26.
    Vermerris W, Nicholson R (2006) Phenolic compound biochemistry. Springer, DordrechtGoogle Scholar
  27. 27.
    Rector BG, Liang G, Guo Y (2003) Effect of maysin on wild-type, deltamethrin-resistant, and Bt-resistant Helicoverpa armigera (Lepidoptera: Noctuidae). J Econ Entomol 96(3):909–913CrossRefGoogle Scholar
  28. 28.
    Casati P, Walbot V (2005) Differential accumulation of maysin and rhamnosylisoorientin in leaves of high-altitude landraces of maize after UV-B exposure. Plant Cell Environ 28(6):788–799. doi: 10.1111/j.1365-3040.2005.01329.x CrossRefGoogle Scholar
  29. 29.
    Suzuki T, Kim S-J, Yamauchi H, Takigawa S, Honda Y, Mukasa Y (2005) Characterization of a flavonoid 3-O-glucosyltransferase and its activity during cotyledon growth in buckwheat (Fagopyrum esculentum). Plant Sci 169(5):943–948. doi: 10.1016/j.plantsci.2005.06.014 CrossRefGoogle Scholar
  30. 30.
    Sato N (2004) Roles of the acidic lipids sulfoquinovosyl diacylglycerol and phosphatidylglycerol in photosynthesis: their specificity and evolution. J Plant Res 117(6):495–505. doi: 10.1007/s10265-004-0183-1 CrossRefGoogle Scholar
  31. 31.
    Roughan PG (1985) Phosphatidylglycerol and chilling sensitivity in plants. Plant Physiol 77(3):740–746CrossRefGoogle Scholar
  32. 32.
    Kenrick JR, Bishop DG (1986) The fatty acid composition of phosphatidylglycerol and sulfoquinovosyldiacylglycerol of higher plants in relation to chilling sensitivity. Plant Physiol 81(4):946–949CrossRefGoogle Scholar
  33. 33.
    Nishihara M, Yokota K, Kito M (1980) Lipid molecular species composition of thylakoid membranes. Biochim Biophys Acta 617(1):12–19CrossRefGoogle Scholar
  34. 34.
    Glauser G, Marti G, Villard N, Doyen GA, Wolfender J-L, Turlings TCJ, Erb M (2011) Induction and detoxification of maize 1,4-benzoxazin-3-ones by insect herbivores. Plant J 68(5):901–911. doi: 10.1111/j.1365-313X.2011.04740.x CrossRefGoogle Scholar
  35. 35.
    Frey M, Chomet P, Glawischnig E, Stettner C, Grun S, Winklmair A, Eisenreich W, Bacher A, Meeley RB, Briggs SP, Simcox K, Gierl A (1997) Analysis of a chemical plant defense mechanism in grasses. Science 277(5326):696–699. doi: 10.1126/science.277.5326.696 CrossRefGoogle Scholar
  36. 36.
    Meihls LN, Handrick V, Glauser G, Barbier H, Kaur H, Haribal MM, Lipka AE, Gershenzon J, Buckler ES, Erb M, Koellner TG, Jander G (2013) Natural variation in maize aphid resistance is associated with 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside methyltransferase activity. Plant Cell 25(6):2341–2355. doi: 10.1105/tpc.113.112409 CrossRefGoogle Scholar
  37. 37.
    Sicker D, Frey M, Schulz M, Gierl A (2000) Role of natural benzoxazinones in the survival strategy of plants. In: Kwang WJ (ed) International review of cytology, vol 198. Academic Press, p 319–346. doi: 10.1016/S0074-7696(00)98008-2
  38. 38.
    Barber J, Gounaris K (1986) What role does sulpholipid play within the thylakoid membrane? Photosynth Res 9(1–2):239–249. doi: 10.1007/bf00029747 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg (outside the USA) 2015

Authors and Affiliations

  • Andrew R. Korte
    • 1
    • 2
  • Marna D. Yandeau-Nelson
    • 3
  • Basil J. Nikolau
    • 2
    • 4
    • 5
  • Young Jin Lee
    • 1
    • 2
    Email author
  1. 1.Department of ChemistryIowa State UniversityAmesUSA
  2. 2.Ames Laboratory-USDOEAmesUSA
  3. 3.Department of Genetics, Development and Cell BiologyIowa State UniversityAmesUSA
  4. 4.Roy J. Carver Department of Biochemistry, Biophysics and Molecular BiologyIowa State UniversityAmesUSA
  5. 5.Center for Metabolic BiologyIowa State UniversityAmesUSA

Personalised recommendations