Quantitative super-resolution microscopy of nanopipette-deposited fluorescent patterns

Simon Hennig, Sebastian van de Linde, Stephan Bergmann, Thomas Huser, Markus Sauer

Research output: Contribution to journalArticle

  • 7 Citations

Abstract

We describe a method for the deposition of minute amounts of fluorophore-labeled oligonucleotides with high local precision in conductive and transparent solid layers of poly(vinyl alcohol) (PVA) doped with glycerin and cysteamine (PVA-G-C layers). Deposition of negatively charged fluorescent molecules was accomplished with a setup based on a scanning ion conductance microscope (SICM) using nanopipettes with tip diameters of ∼100 nm by using the ion flux flowing between two electrodes through the nanopipette. To investigate the precision of the local deposition process, we performed in situ super-resolution microscopy by direct stochastic optical reconstruction microscopy (dSTORM). Exploiting the single-molecule sensitivity and reliability of dSTORM, we determine the number of fluorescent molecules deposited in single spots. The correlation of applied charge and number of deposited molecules enables the quantification of delivered molecules by measuring the charge during the delivery process. We demonstrate the reproducible deposition of 3-168 fluorescent molecules in single spots and the creation of fluorescent structures. The fluorescent structures are highly stable and can be reused several times.

LanguageEnglish
Pages8122-8130
Number of pages9
JournalACS Nano
Volume9
Issue number8
Early online date14 Jul 2015
DOIs
StatePublished - 25 Aug 2015
Externally publishedYes

Fingerprint

Microscopy
Microscopic examination
microscopy
Molecules
Ions
Cysteamine
molecules
Oligonucleotides
Glycerol
Electrodes
Alcohols
cysteamine
Fluorophores
oligonucleotides
delivery
ions
Microscopes
alcohols
microscopes
Fluxes

Keywords

  • dSTORM
  • nanopipette
  • super-resolution microscopy
  • SICM
  • single-molecule deposition

Cite this

Hennig, Simon ; van de Linde, Sebastian ; Bergmann, Stephan ; Huser, Thomas ; Sauer, Markus. / Quantitative super-resolution microscopy of nanopipette-deposited fluorescent patterns. In: ACS Nano. 2015 ; Vol. 9, No. 8. pp. 8122-8130
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Hennig, S, van de Linde, S, Bergmann, S, Huser, T & Sauer, M 2015, 'Quantitative super-resolution microscopy of nanopipette-deposited fluorescent patterns' ACS Nano, vol. 9, no. 8, pp. 8122-8130. DOI: 10.1021/acsnano.5b02220

Quantitative super-resolution microscopy of nanopipette-deposited fluorescent patterns. / Hennig, Simon; van de Linde, Sebastian; Bergmann, Stephan; Huser, Thomas; Sauer, Markus.

In: ACS Nano, Vol. 9, No. 8, 25.08.2015, p. 8122-8130.

Research output: Contribution to journalArticle

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AB - We describe a method for the deposition of minute amounts of fluorophore-labeled oligonucleotides with high local precision in conductive and transparent solid layers of poly(vinyl alcohol) (PVA) doped with glycerin and cysteamine (PVA-G-C layers). Deposition of negatively charged fluorescent molecules was accomplished with a setup based on a scanning ion conductance microscope (SICM) using nanopipettes with tip diameters of ∼100 nm by using the ion flux flowing between two electrodes through the nanopipette. To investigate the precision of the local deposition process, we performed in situ super-resolution microscopy by direct stochastic optical reconstruction microscopy (dSTORM). Exploiting the single-molecule sensitivity and reliability of dSTORM, we determine the number of fluorescent molecules deposited in single spots. The correlation of applied charge and number of deposited molecules enables the quantification of delivered molecules by measuring the charge during the delivery process. We demonstrate the reproducible deposition of 3-168 fluorescent molecules in single spots and the creation of fluorescent structures. The fluorescent structures are highly stable and can be reused several times.

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Hennig S, van de Linde S, Bergmann S, Huser T, Sauer M. Quantitative super-resolution microscopy of nanopipette-deposited fluorescent patterns. ACS Nano. 2015 Aug 25;9(8):8122-8130. Available from, DOI: 10.1021/acsnano.5b02220