Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Super-resolution Fluorescence Microscopy01:37

Super-resolution Fluorescence Microscopy

Super-resolution fluorescence microscopy (SRFM) provides a better resolution than conventional fluorescence microscopy by reducing the point spread function (PSF). PSF is the light intensity distribution from a point that causes it to appear blurred. Due to PSF, each fluorescing point appears bigger than its actual size, and it is the PSF interference of nearby fluorophores that causes the blurred image. Various approaches to achieving higher resolution through SRFM have recently been developed.

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

PTPN22 regulates T cell synapse formation through PSTPIP1-dependent actin remodeling.

Science signaling·2026
Same author

Enabling the synthesis of multi-payload thio-antibody conjugates through the use of pyridazinediones, <i>p</i>-anisidine derivatives and various click chemistries.

RSC chemical biology·2026
Same author

Mesoscale transport of enveloped viruses.

The Journal of chemical physics·2026
Same author

Triggered "Capture-and-Release" Enables a High-Affinity Rebinding Strategy for Sensitivity Enhancement in Lateral Flow Assays.

ACS sensors·2025
Same author

Bis-amino maleimides: a new class of dual hydrogen bond donor for anion binding and transport.

Organic & biomolecular chemistry·2025
Same author

Mass spectrometric imaging and quantitative analysis of the <i>in vivo</i> biodistribution of trastuzumab using a rhodium(iii) sarcophagine complex.

Inorganic chemistry frontiers·2025

Related Experiment Video

Updated: May 21, 2026

Ground State Depletion Super-resolution Imaging in Mammalian Cells
07:55

Ground State Depletion Super-resolution Imaging in Mammalian Cells

Published on: November 5, 2017

Stoichiometrically Defined Antibody-DNA Conjugates for Quantitative Super-Resolution Imaging.

Luciana P Martinez1, Clíona McMahon2, Cecilia Zaza1

  • 1London Centre for Nanotechnology, University College London, 19 Gordon Street, WC1H 0AH London, United Kingdom.

Nano Letters
|May 20, 2026
PubMed
Summary
This summary is machine-generated.

Accurate protein quantification using DNA-PAINT requires precisely defined antibody-DNA conjugates. Site-selective conjugation methods ensure stoichiometric control, improving quantitative reliability in super-resolution microscopy.

Keywords:
pyridazinedione chemistryquantitative DNA-PAINTsingle-molecule localization microscopysite-selective labelingstoichiometric antibody−DNA conjugates

More Related Videos

Whole-cell Super-Resolution Imaging via DNA-PAINT on a Spinning Disk Confocal with Optical Photon Reassignment
07:12

Whole-cell Super-Resolution Imaging via DNA-PAINT on a Spinning Disk Confocal with Optical Photon Reassignment

Published on: January 6, 2026

Highly Multiplexed, Super-resolution Imaging of T Cells Using madSTORM
08:43

Highly Multiplexed, Super-resolution Imaging of T Cells Using madSTORM

Published on: June 24, 2017

Related Experiment Videos

Last Updated: May 21, 2026

Ground State Depletion Super-resolution Imaging in Mammalian Cells
07:55

Ground State Depletion Super-resolution Imaging in Mammalian Cells

Published on: November 5, 2017

Whole-cell Super-Resolution Imaging via DNA-PAINT on a Spinning Disk Confocal with Optical Photon Reassignment
07:12

Whole-cell Super-Resolution Imaging via DNA-PAINT on a Spinning Disk Confocal with Optical Photon Reassignment

Published on: January 6, 2026

Highly Multiplexed, Super-resolution Imaging of T Cells Using madSTORM
08:43

Highly Multiplexed, Super-resolution Imaging of T Cells Using madSTORM

Published on: June 24, 2017

Area of Science:

  • Biophysics
  • Molecular Imaging
  • Bioconjugation Chemistry

Background:

  • DNA-PAINT super-resolution microscopy allows precise protein copy number determination via qPAINT.
  • Current methods using amine-reactive labeling create heterogeneous antibody-DNA conjugates, limiting quantitative accuracy.
  • Stoichiometric control of DNA docking strands on antibodies is crucial for reliable qPAINT analysis.

Purpose of the Study:

  • To develop a site-selective conjugation method for creating stoichiometrically defined antibody-DNA conjugates.
  • To evaluate the impact of controlled DNA loading on qPAINT accuracy.
  • To enable reliable quantitative super-resolution imaging in complex biological samples.

Main Methods:

  • Site-selective disulfide rebridging using pyridazinedione chemistry for antibody conjugation.
  • Generation of antibody-DNA conjugates with defined stoichiometry: one (OAR1) or four (OAR4) DNA strands per antibody.
  • Application of OAR1 and OAR4 probes to nuclear pore complexes for qPAINT analysis.

Main Results:

  • OAR1 and OAR4 probes produced narrow qPAINT index distributions, indicating stoichiometric control.
  • Quantitative DNA-PAINT analysis with defined conjugates showed accurate protein copy number dependence.
  • Site-selective conjugation enabled discrimination between singly and doubly labeled Nup96 dimers.
  • Conventional lysine-based conjugation resulted in broadened distributions and overestimation of copy numbers.

Conclusions:

  • Site-selective antibody-DNA conjugation is essential for accurate quantitative DNA-PAINT.
  • Pyridazinedione chemistry provides a robust method for generating stoichiometrically defined antibody-DNA conjugates.
  • This approach enhances the reliability of super-resolution microscopy for quantitative biological studies.