The thickness of electron transparent samples can be measured in an electron microscope using several imaging techniques like electron energy loss spectroscopy (EELS) or quantitative scanning transmission electron microscopy (STEM). We extrapolate this method for using a back-scattered electron (BSE) detector in the scanning electron microscope (SEM). This brings the opportunity to measure the thickness not just of the electron transparent samples on TEM mesh grids, but, in addition, also the thickness of thin films on substrates. Nevertheless, the geometry of the microscope and the BSE detector poses a problem with precise calibration of the detector. We present a simple method which can be used for such a type of detector calibration that allows absolute (standardless) measurement of thickness, together with a proof of the method on test samples.

Institute of Scientific Instruments of the Czech Academy of Sciences Kralovopolska 147 CZ 61264 Brno Czech Republic

Zobrazit více v PubMed

Echlin P. Handbook of Sample Preparation for Scanning Electron Microscopy and X-ray Microanalysis. Springer; Boston, MA, USA: 2009. p. 332. DOI

Schatten H., Walocha J., Litwin J., Miodoński A., Sepsenwol S., Naguro T., Nakane H., Inaga S., Walther P., Albrecht R., et al. Scanning Electron Microscopy for the Life Sciences. Cambridge University Press; Cambridge, UK: 2012. p. 261.

Fleck R.A., Humbel B.M. Biological Field Emission Scanning Electron Microscopy. 1st ed. Wiley; Hoboken, NJ, USA: 2018. p. 752.

Brodusch N., Demers H., Gauvin R. Field Emission Scanning Electron Microscopy. 1st ed. Springer; Singapore: 2018. p. 137. Springe Briefs in Applied Sciences and Technology. DOI

Goldstein J., Newbury D.E., Michael J.R., Ritchie N.W.M., Scott J.H.J. Scanning Electron Microscopy and X-ray Microanalysis. 4th ed. Springer; New York, NY, USA: 2018. p. 550.

Assa’d A.M.D., Gomati M.M.E. Backscattering Coefficients for Low Energy Electrons. Scan. Microsc. 1998;12:185–192.

Lin W.R., Chuang Y.J., Lee C.H., Tseng F.G., Chen F.R. Fabrication and characterization of a sensitivity multi-annular backscattered electron detector for desktop SEM. Sensors. 2018;18:3093. doi: 10.3390/s18093093. PubMed DOI PMC

Krzyzanek V., Sporenberg N., Keller U., Guddorf J., Reichelt R., Schönhoff M. Polyelectrolyte multilayer capsules: Nanostructure and visualisation of nanopores in the wall. Soft Matter. 2011;7:7034–7041. doi: 10.1039/c1sm05406f. DOI

Dapor M., Bazzanella N., Toniutti L., Miotello A., Gialanella S. Backscattered electrons from surface films deposited on bulk targets: A comparison between computational and experimental results. Nucl. Instrum. Methods Phys. Res. Sect. B Beam Interact. Mater. Atoms. 2011;269:1672–1674. doi: 10.1016/j.nimb.2010.11.016. DOI

Dapor M., Bazzanella N., Toniutti L., Miotello A., Crivellari M., Gialanella S. Backscattered electrons from gold surface films deposited on silicon substrates: A joint experimental and computational investigation to add new potentiality to electron microscopy. Surf. Interface Anal. 2013;45:677–681. doi: 10.1002/sia.5144. DOI

Assa’d A.M.D. Monte Carlo calculation of the backscattering coefficient of thin films of low on high atomic number materials and the reverse as a function of the incident electron energy and film thickness. Appl. Phys. A. 2018;124:699. doi: 10.1007/s00339-018-2073-8. DOI

Müller E., Gerthsen D. Composition quantification of electron-transparent samples by backscattered electron imaging in scanning electron microscopy. Ultramicroscopy. 2017;173:71–75. doi: 10.1016/j.ultramic.2016.12.003. PubMed DOI

Volkenandt T., Müller E., Hu D.Z., Schaadt D.M., Gerthsen D. Quantification of sample thickness and in-concentration of InGaAs quantum wells by transmission measurements in a scanning electron microscope. Microsc. Microanal. 2010;16:604–613. doi: 10.1017/S1431927610000292. PubMed DOI

Kim H., Negishi T., Kudo M., Takei H., Yasuda K. Quantitative backscattered electron imaging of field emission scanning electron microscopy for discrimination of nano-scale elements with nm-order spatial resolution. J. Electron. Microsc. 2010;59:379–385. doi: 10.1093/jmicro/dfq012. PubMed DOI

Garitagoitia Cid A., Rosenkranz R., Löffler M., Clausner A., Standke Y., Zschech E. Quantitative analysis of backscattered electron (BSE) contrast using low voltage scanning electron microscopy (LVSEM) and its application to AlGaN/GaN layers. Ultramicroscopy. 2018;195:47–52. doi: 10.1016/j.ultramic.2018.08.026. PubMed DOI

Sánchez E., Torres Deluigi M., Castellano G. Mean atomic number quantitative assessment in backscattered electron imaging. Microsc. Microanal. 2012;18:1355–1361. doi: 10.1017/S1431927612013566. PubMed DOI

Reimer L. Scanning Electron Microscopy: Physics of Image Formation and Microanalysis. 2nd ed. Springer; Berlin/Heidelberg, Germany: New York, NY, USA: 1998. p. 538.

Volkenandt T., Müller E., Gerthsen D. Sample thickness determination by scanning transmission electron microscopy at low electron energies. Microsc. Microanal. 2014;20:111–123. doi: 10.1017/S1431927613013913. PubMed DOI

Skoupy R., Nebesarova J., Slouf M., Krzyzanek V. Quantitative STEM imaging of electron beam induced mass loss of epoxy resin sections. Ultramicroscopy. 2019;202:44–50. doi: 10.1016/j.ultramic.2019.03.018. PubMed DOI

Pfaff M., Müller E., Klein M.F.G., Colsmann A., Lemmer U., Krzyzanek V., Reichelt R., Gerthsen D. Low-energy electron scattering in carbon-based materials analyzed by scanning transmission electron microscopy and its application to sample thickness determination. J. Microsc. 2011;243:31–39. doi: 10.1111/j.1365-2818.2010.03475.x. PubMed DOI

Seiter J., Müller E., Blank H., Gehrke H., Marko D., Gerthsen D. Backscattered electron SEM imaging of cells and determination of the information depth. J. Microsc. 2014;254:75–83. doi: 10.1111/jmi.12120. PubMed DOI

Demers H., Poirier-Demers N., Couture A.R., Joly D., Guilmain M., de Jonge N., Drouin D. Three-dimensional electron microscopy simulation with the CASINO Monte Carlo software. Scanning. 2011;33:135–146. doi: 10.1002/sca.20262. PubMed DOI PMC

Salvat F., Jablonski A., Powell C.J. Elsepa—Dirac partial-wave calculation of elastic scattering of electrons and positrons by atoms, positive ions and molecules. Comput. Phys. Commun. 2005;165:157–190. doi: 10.1016/j.cpc.2004.09.006. DOI

Merli P., Morandi V., Corticelli F. Backscattered electron imaging and scanning transmission electron microscopy imaging of multi-layers. Ultramicroscopy. 2003;94:89–98. doi: 10.1016/S0304-3991(02)00217-6. PubMed DOI

Kowoll T., Müller E., Fritsch-Decker S., Hettler S., Störmer H., Weiss C., Gerthsen D. Contrast of Backscattered Electron SEM Images of Nanoparticles on Substrates with Complex Structure. Scanning. 2017;2017:1–12. doi: 10.1155/2017/4907457. PubMed DOI PMC

Leosson K., Ingason A.S., Agnarsson B., Kossoy A., Olafsson S., Gather M.C. Ultra-thin gold films on transparent polymers. Nanophotonics. 2013;2:3–11. doi: 10.1515/nanoph-2012-0030. DOI

Dapor M. Backscattering of low energy electrons from carbon films deposited on aluminum: A Monte Carlo simulation. J. Appl. Phys. 2004;95:718–721. doi: 10.1063/1.1633655. DOI

Assa’d A.M. Monte Carlo computation of the influence of carbon contamination layer on the energy distribution of backscattered electrons emerging from Al and Au. Jordan J. Phys. 2019;12:37–44.

An advanced fast method for the evaluation of multiple immunolabelling using gold nanoparticles based on low-energy STEM

Quantification of STEM Images in High Resolution SEM for Segmented and Pixelated Detectors