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Spooling electrochemiluminescence spectroscopy: development, applications and beyond

Nature Protocols volume 16, pages 2109–2130 (2021)Cite this article

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Abstract

One of the most widely used techniques to generate light through an efficient electron transfer is called electrochemiluminescence, or electrogenerated chemiluminescence (ECL). ECL mechanisms can be explored via ‘spooling spectroscopy’ in which individual ECL spectra showing emitted light are collected continuously during a potentiodynamic course. The obtained spectra are spooled together and plotted along the applied potential axis; because the potential sweep occurs at a defined rate, this axis is directly proportional to time. Any changes in the emission spectra can be correlated to the corresponding potentials and/or times, leading to a deeper understanding of the mechanism for light generation—information that can be used for efficiently maximizing ECL intensities. The formation of intermediates and excited states can also be tracked, which is crucial to interrogating and drawing electron transfer pathways (i.e., understanding the chemical reaction mechanism). Spooling spectroscopy is not limited to ECL; we also include instructions for the use of related methodologies, such as spooling photoluminescence spectroscopy during an electrolysis procedure, which can be easily set up. The total time required to complete the protocol is ~49 h, from making electrodes and an ECL cell, fabricating light-tight housing, to setting up instruments. Preparing the lab for an individual experiment (making an electrolyte solution of a targeted luminophore, cooling down the CCD camera, calibrating the spectrometer and surveying electrochemistry) takes ~1 h 15 min, and performing the spooling ECL spectroscopy experiment itself requires ~10 min.

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Fig. 1: Spooling spectroscopy setup.
Fig. 2: Required components to assemble a light-tight ECL cell.
Fig. 3: Photograph of a typical platinum working electrode.
Fig. 4: A typical ECL cell.
Fig. 5: Fabrication of ECL cell cap.
Fig. 6: Schematic representation of an ECL cell holder.
Fig. 7: Potentiostat and camera synchronization setup details.
Fig. 8: Spooling photoluminescence spectroscopy setup.
Fig. 9: An example cyclic voltammogram.
Fig. 10: An illustration of a home-made polishing wheel.
Fig. 11
Fig. 12: Exemplary spooling ECL spectra.
Fig. 13: In situ spooling photoluminescence spectroscopy.

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Data availability

The authors declare that the main data supporting the findings of this study are available within the article. A simple sample spooling ECL spectroscopy figure in MATLAB for Ru(bpy)32+ in the presence of 5 mM tri-n-propylamine (TPrA) and its user guide (‘Spooling ECL spectroscopy plot of 0.1 mM Ru(bpy)32+ with 5 mM TPrA using MATLAB’ and ‘Manual to use spooling ECL spectroscopy of Ru(bpy) with TPrA’, respectively) are included in Supplementary Data 1. The spooling ECL spectrum source data in ‘acs’ format and other advanced plotting codes in MATLAB and spreadsheet software Igor are available from the corresponding authors upon request. The data relating to this protocol can be found in this paper and the supporting primary research papers. See the Supplementary Data 1 for data relating to Figs. 9 and 12a. The MATLAB code used in this protocol can be found in the Supplementary Information.

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Acknowledgements

Our special thanks go to J. Vanstone and his team in the Chemistry Electronic Shop who patiently helped us to build many custom parts for our lab, especially for our ECL instrumentation. We also thank the Department of Chemistry and ChemBio Store at The University of Western Ontario for quality support and services. We acknowledge the Natural Sciences and Engineering Research Council of Canada (NSERC, DG RGPIN-2013-201697, RGPIN-2018-06556, SPG STPGP-2016-493924 and several Research Tool and Instruments grants since 2002), Canada Foundation of Innovation/Ontario Innovation Trust (CFI/OIT, 9040), and Premier’s Research Excellence Award (PREA, 2003) for financial support of this research. We thank previous and present members of the Ding group for continuously using and developing the spooling ECL spectroscopy as well as the plotting codes. We appreciate the inspiring advice on the protocol submission from Y. Long.

Author information

Authors and Affiliations

  1. Department of Chemistry, The University of Western Ontario, London, Ontario, Canada

    Mahdi Hesari & Zhifeng Ding

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  1. Mahdi Hesari
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Contributions

M.H. and Z.D. conceived the writing plan and designed the research. M.H. took pictures to illustrate instruments and parts, and performed experiments. M.H. organized and summarized the figures. M.H. and Z.D. wrote the manuscript. Z.D. finalized and submitted the manuscript. All the authors were involved in discussions about the manuscript at all stages.

Corresponding authors

Correspondence to Mahdi Hesari or Zhifeng Ding.

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The authors declare no competing interests.

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Peer review information Nature Protocols thanks Joohoon Kim, Jun-Jie Zhu, Gui-Zheng Zou and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Related links

Key references using this protocol

Hesari, M. et al. J. Am. Chem. Soc. 137, 11266–11269 (2015): https://pubs.acs.org/doi/10.1021/jacs.5b07633

Hesari, M., Workentin, M. S. & Ding, Z. ACS Nano 8, 8543–8553 (2014): https://pubs.acs.org/doi/10.1021/nn503176g

Hesari, M., Workentin, M. S. & Ding, Z. Chem. Sci. 5, 3814–3822 (2014): https://pubs.rsc.org/en/content/articlelanding/2014/SC/C4SC01086H#!divAbstract

Swanick, K. N., Hesari, M., Workentin, M. S. & Ding, Z. J. Am. Chem. Soc. 134, 15205–15208 (2012): https://doi.org/10.1021/ja306047u

Key data used in this protocol

Hesari, M., Workentin, M. S. & Ding, Z. ACS Nano. 8, 8543–8553 (2014): https://pubs.acs.org/doi/10.1021/nn503176g

Hesari, M. et al. J. Am. Chem. Soc. 137, 11266–11269 (2015): https://pubs.acs.org/doi/10.1021/jacs.5b07633

Hesari, M., Workentin, M. S. & Ding, Z. Chem. Sci. 5, 3814–3822 (2014): http://pubs.rsc.org/en/content/articlepdf/2014/sc/c4sc01086h

Extended data

Extended Data Fig. 1 Assembly of a platinum electrode inside a soft glass.

A vacuum tube is attached to one end of the soft glass to assist with Pt rod insertion into the other end.

Extended Data Fig. 2 Schematic assembly of ECL cell-focus lens-PMT.

A photograph of an exemplary ECL cell including WE (Pt disc), CE (Pt coil), RE (Pt coil), sample solution, quartz window, focus lens, and a PMT along with a recorder. Note that the represented distances are not scaled.

Extended Data Fig. 3 An illustration of optical filter position.

a, Top view of the spectrograph port where a filter is placed to block the excitation light source to perform a spooling photoluminescence spectroscopy. b, Side view of the filter and spectrograph port.

Extended Data Fig. 4 Spooling spectroscopy software setup.

A representative of an Andor iDus camera acquisition setup (red box), acquisition mode, exposure time and kinetic series length (pink boxes) are set as spooling, time interval between each spectrum, and total number of spectra. As a common example, experimental parameters related to spooling ECL spectroscopy of 0.1 mM [Ru(bpy)3](PF6)2 (Fig. 9) in the presence of 5 mM tri-n-propylyamine (TPrA) are displayed. The spooling ECL spectra are shown in Fig. 12a. In this exemplary experiment, the applied potential was scanned from 0.0 V to +1.5 V and back to 0.0 V vs. SCE where TPrA was oxidized at ~0.8 V to produce TPrA radicals. At the close potential, Ru(bpy)32+ lost one electron at ~1.1 to Ru(bpy)33+. The total time required for the cycle of potential scan is 30 sec. Thus, one can perform a spooling ECL spectroscopy of Ru(bpy)32+ / TPrA by setting up camera acquisition (red box) and selecting “kinetic” for acquisition mode (first pink box at left). Individual spooling spectra can be collected during 1 s of camera exposure time (second pink box at top left). Usually, the number of accumulations needs to be 1. Finally, “kinetic series length” (third pink box at left) represents total time required for an experiment to complete, which is 30 sec in the present example. The spectrum time interval is defined as the density of the collected spectra in time-space. The shorter the time interval, the higher the number of spectra. To set up a trigger, one needs to set the triggering option to “External Start” which means the camera will start to acquire a spectrum (according the setting) as soon as the external signal has been sent through the coaxial cable from the potentiostat.

Extended Data Fig. 5 Spooling spectroscopy software setup.

Before running a spooling spectroscopy experiment, choose the spooling tab (pink tab) in the acquisition setting (red box) and enable spooling. Next, select “Sif” (shown with arrow) as the file type (first green box at top left). One can choose a file name, e.g., “spool” which will be saved with a “.sif” extension. Lastly, select a path where the recorded file will be saved. Note 1: the last two steps can also be done manually. Note 2: all the recorded spooling files with “.sif” extension should be converted to “.acs” in order to be opened with Igor Pro or MATLAB for plotting.

Extended Data Fig. 6 Exemplary electrode junctions.

Photographs of assembled a, counter electrode, and b, reference electrode inside glass capillaries to protect CE and RE from possible side reactions.

Supplementary information

Reporting Summary

Supplementary Data 1

Manual to use spooling spectroscopy of Ru(bpy) with TPrA and sample spooling ECL plot using MATLAB software.

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Hesari, M., Ding, Z. Spooling electrochemiluminescence spectroscopy: development, applications and beyond. Nat Protoc 16, 2109–2130 (2021). https://doi.org/10.1038/s41596-020-00486-x

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