Carbon-coated Grids Research Articles

Correlative atomic force microscopy and scanning electron microscopy of bacteria-diamond-metal nanocomposites

Research investigating the interface between biological organisms and nanomaterials nowadays requires multi-faceted microscopic methods to elucidate the interaction mechanisms and effects. Here we describe a novel approach and methodology correlating data from an atomic force microscope inside a scanning electron microscope (AFM-in-SEM). This approach is demonstrated on bacteria-diamond-metal nanocomposite samples relevant in current life science research. We describe a procedure for preparing such multi-component test samples containing E. coli bacteria and chitosan-coated hydrogenated nanodiamonds decorated with silver nanoparticles on a carbon-coated gold grid. Microscopic topography information (AFM) is combined with chemical, material, and morphological information (SEM using SE and BSE at varied acceleration voltages) from the same region of interest and processed to create 3D correlative probe-electron microscopy (CPEM) images. We also establish a novel 3D RGB color image algorithm for merging multiple SE/BSE data from SEM with the AFM surface topography data which provides additional information about microscopic interaction of the diamond-metal nanocomposite with bacteria, not achievable by individual analyses. The methodology of CPEM data interpretation is independently corroborated by further in-situ (EDS) and ex-situ (micro-Raman) chemical characterization as well as by force volume AFM analysis. We also discuss the broader applicability and benefits of the methodology for life science research.

(Invited) High-Throughput Nanoscale Resolved Electrochemistry to Study Electrochemical Nucleation, Growth and Dissolution

Electrochemical nucleation and growth (EN&G) is the cornerstone for many (nano)material growth routes and the main factor limiting battery durability. At the same time electrochemical dissolution (ED) is the main cause of material degradation in exposed environments (corrosion) or energy conversion and storage devices. The in-depth experimental assessment of both processes is very challenging. The reasons are the random nature of initiation events (nucleation), the heterogeneity of surfaces and the (very) fast kinetics of these processes across several length scales. For all that, our current understanding of the mechanisms involved is inaccurate and incomplete [1].During the last years, we have developed an approach based on using carbon-coated TEM grids as electrodes to combine ex-situ atomic-scale TEM characterization with electron tomography and macroscale electrochemical measurements [2,3]. This approach has brought valuable evidence of non-classical growth pathways such as growth mediated by nanocluster aggregation. Yet, it does not capture the influence of the heterogeneous nature of the surface where EN&G proceeds, nor the dynamics before, during and after nucleation [4,5].In this contribution, we present our recent work in which we combine high-throughput nanoscale resolved electrochemistry by Scanning Electrochemical Cell Microscopy (SECCM), with ex-situ and in-situ high resolution characterization, including electrochemical transmission electron microscopy (EC-TEM), to study the electrochemical nucleation, growth, and dissolution of metal (Cu, Au, Ag and Pt) nanoparticles (NPs) [6,7]. The spatially resolved electrochemical characterization enables a one-to-one correlation between the electrochemical data and the local surface properties, which can be evaluated by different surface analytical tools.Moreover, the confinement of the electrochemical cell to the SECCM meniscus enables us to resolve a diversity of events during the electrochemical dissolution of electrodeposited NPs. EC-TEM experiments advocate that the nature of these events corresponds to the dissolution of individual NPs spanning a wide range of time [6].The combination of SECCM and EC-TEM opens up new opportunities for the rational design of functional nanostructured materials by electrodeposition, and for the evaluation of their durability under electrochemical polarization. The ability to study these taking into account the heterogeneous nature of the supports and the differences within nanomaterial ensembles is essential for applications in electrochemical conversion and storage.

Landmark-based retrieval of inflamed skin vessels enabled by 3D correlative intravital light and volume electron microscopy

The nanometer spatial resolution of electron microscopy imaging remains an advantage over light microscopy, but the restricted field of view that can be inspected and the inability to visualize dynamic cellular events are definitely drawbacks of standard transmission electron microscopy (TEM). Several methods have been developed to overcome these limitations, mainly by correlating the light microscopical image to the electron microscope with correlative light and electron microscopy (CLEM) techniques. Since there is more than one method to obtain the region of interest (ROI), the workflow must be adjusted according to the research question and biological material addressed. Here, we describe in detail the development of a three-dimensional CLEM workflow for mouse skin tissue exposed to an inflammation stimulus and imaged by intravital microscopy (IVM) before fixation. Our aim is to relocate a distinct vessel in the electron microscope, addressing a complex biological question: how do cells interact with each other and the surrounding environment at the ultrastructural level? Retracing the area over several preparation steps did not involve any specific automated instruments but was entirely led by anatomical and artificially introduced landmarks, including blood vessel architecture and carbon-coated grids. Successful retrieval of the ROI by electron microscopy depended on particularly high precision during sample manipulation and extensive documentation. Further modification of the TEM sample preparation protocol for mouse skin tissue even rendered the specimen suitable for serial block-face scanning electron microscopy (SBF-SEM).

First Report of Tomato Brown Rugose Fruit Virus in Tomato in Norway.

In May 2021, tomato (Solanum lycopersicum L.) plants with necrosis and ringspot symptoms were observed in a farm greenhouse at Sundbyfoss, Norway. This greenhouse production focused on ecological growing of several tomato varieties (i.e., "Blush Tiger", "Sailor's Luck", "Evil Oliver", etc.) with compost for local customers. The presence of tomato brown rugose fruit virus (ToBRFV) was suspected due to the symptoms in the diseased plants. Sap inoculation was carried out with extracts from 100 mg symptomatic tomato fresh leaves in 0.03 M PBS (phosphate buffered saline, pH 7.0), and inoculation on three Nicotiana tabacum cv. Xanthi plants at the 4-6 leaf stage grown in an experimental greenhouse. The test plants showed local necrotic lesions after 3-5 days. Two symptomatic leaf samples, one from tomato and the other from a test plant, were analysed by transmission electron microscope (TEM) with the help of negative staining. The samples were treated with 0.1 M PBS (pH 7.3) first and placed on carbon-coated EM grids and stained with 2% uranyl acetate before TEM observation. Rigid rod-shaped viral particles, typical of tobamovirus particls (around 300 nm) were observed. Total RNA was isolated from two tomato leaves showing symptoms using a Norgen Plant/Fungi RNA kit (Norgen Biotek, Canada). One-step reverse-transcription (RT)-PCR with specific primers ToBRFV-F/ToBRFV-R for ToBRFV (Alkowni et al. 2019), which amplified a 560-bp fragment, was performed. The sequence obtained by Sanger sequencing from the amplicon (535 nt) showed 99.8% nt identity with ToBRFV isolate PV-1241 (NCBI accession no. MZ202349) from DSMZ (Leibniz Institute, German) and was deposited in the GenBank database under the accession number OK358628. The infection was also confirmed with duplex real-time RT-PCR test for ToBRFV using CaTa28 and CSP1325 primers and probe (ISF 2020; EPPO, PM7/146 2021). In addition, eight tomato samples from the same farm greenhouse were collected according to cultivars, location in the greenhouse and symptoms before eradication and disinfection: four of the tomato samples were confirmed positive for ToBRFV with one-step RT-PCR as described above. A surveillance program for ToBRFV in commercial greenhouse production has been carried out in 2021 in Norway. Around 4000 tomato plants from 18 commercial tomato growers were tested. No positive ToBRFV samples have been detected. However, unregulated tomato seeds from abroad and self-propagation of tomato by private gardeners and hobby growers is a potential threat to commercial production of tomato in Norway. To our knowledge, this is the first report of ToBRFV associated with tomato in Norway and in the Nordic countries.

Correction: Surprising toxicity and assembly behaviour of amyloid β-protein oxidized to sulfone.

Biochem. J.433 (2), 323–332 https://doi.org/10.1042/BJ20101391A reader has contacted the Biochemical Journal Editorial Board to draw attention to a concern regarding one of the Supplementary Figures in this article. The concern raised is regarding Supplementary Figure 2 and the authors have acknowledged that the same image has been used twice in Supplementary Figure 2 Aβ40-sulfone by mistake. The authors have provided a corrected Supplementary Figure 2, this correction does not impact the main conclusion of the paper.Figure S2 Morphology of native and oxidized A β analoguesAβ analogues were incubated in DMEM at 37°C, and aliquots were spotted on to glow-discharged, carbon-coated grids, stained with 1% uranyl acetate, and examined by electron microscopy. Medium alone also was imaged as a negative control. Scale bars are 100 nm. Each image is representative of three independent experiments.

Surface modification of bare copper grids with charged polymers: A simple alternative for carbon coated copper grids in transmission electron microscopy

We report a methodology to functionalize the bare copper grids for high resolution transmission electron microscopy (TEM), by drop casting a thin polymer film instead of carbon film. The polymer coating actively helped in trapping the sample on the top of the mesh, making it available for the imaging. Gold nanorods (AuNRs), poly(allylamine) hydrochloride (PAH)/dextran sulphate (DS) nanocapsules and PAH/DS nanocapsules incorporated with AuNRs, were used as samples on the bare grids, carbon-coated grids and polymer-coated grids. The effect of polymer charge, polymer concentration and film thickness on imaging quality was studied using different polymers such as chitosan, PAH, polystyrene sulfonate, DS and gellan gum. The polymer-coated copper grids showed a comparable performance with the conventional carbon-coated copper grids. Hence, the methodology provides a simple and economical way to obtain high quality TEM images using the bare copper grids.

The critical role of mobile phase pH in the performance of oligonucleotide ion-pair liquid chromatography-mass spectrometry methods.

The critical role of mobile phase pH in the performance of oligonucleotide ion-pair liquid chromatography-mass spectrometry methods.

Multicomponent perovskite superlattices

Multicomponent perovskite superlattices

Abstract P-29: Cryoem Study of the Inhibition of Bacterial Ribosomes by Madumycin II

Background: The efficiency of widely used antibiotics is limited by continuous improvement of resistance mechanisms. Thus, the research of poorly studied drugs that have not received practical use until now becomes relevant again. Protein translation is one of the major targets for antibiotics. Madumycin II (MADU) is an antibiotic of the streptogramin A class that binds to the peptidyl transferase center of the initiated bacterial 70S ribosome inhibiting the first cycle of peptide bond formation (I.A. Osterman et al. Nucleic Acids Res., 2017). The ability of MADU to interfere with translating ribosome is an open question that we address by investigation of high-resolution cryo-EM structures of MADU bound 70S ribosome complexes from Escherichia coli. Methods: Purified initiated and translating ribosome complexes preincubated with MADU were applied onto freshly glow discharged carbon-coated grids (Quantifoil R 1.2/1.3) and flash-frozen in the liquid ethane pre-cooled by liquid nitrogen in the Vitrobot Mark IV. Frozen grids were transferred into an in-house Titan Krios microscope. Data were collected using EPU software. Movie stacks were preprocessed in Warp software. For image processing, we have used several software packages: Relion 3.1, CryoSPARC, and CisTEM. The model was built in Coot. Results: We have obtained high-resolution cryo-EM structures of two ribosomal complexes with MADU before and after the first cycle of peptide bond formation with an average resolution of 2.3 Å. Preliminary analysis of the structures shows no major differences in the MADU binding mode to the ribosomal complexes under study suggesting that the quantity of amino acid residues attached to the P-site tRNA does not impact MADU bonding. Moreover, in both cases, we observed similar destabilization of the CCA-ends of A- and P-site tRNAs underlining the comparable influence of MADU on the ribosomal complexes. Conclusion: Our results suggest that although MADU binding site is located in the peptidyl transferase center, the presence of the second amino acid residue on the P-site tRNA does not preclude antibiotic binding. We assume that further elongation of the polypeptide chain would not have any impact either. High conformational lability of the CCA-ends of tRNA at the A and P sites upon binding of MADU obviously plays an important role in the inhibition mechanism of the bacterial ribosome. The further structural and biochemical analysis will be necessary to shed more light on the detailed mechanism of MADU action.

Visualization of catalyst dynamics and development of a practical procedure to study complex "cocktail"-type catalytic systems.

The ability to distinguish molecular catalysis from nanoscale catalysis provides a key to success in the field of catalyst development, particularly for the transition to sustainable economies. Complex evolution of catalyst precursors, facilitated by dynamic interconversions and leaching, makes the identification of catalytically active forms an important task, which is sometimes very difficult. We propose a simple method for in situ capturing of nanoparticles with carbon-coated grids directly from reaction mixtures. Application of this method to the Mizoroki-Heck reaction allowed visualization of dynamic changes of the dominant form of palladium particles in the reaction mixtures with homogeneous and heterogeneous catalyst precursors. Changes in the size and shape of the palladium particles reflecting the progress of the catalytic chemical reaction were demonstrated. Detailed computational modeling was carried out to confirm the generality of this approach and its feasibility for different catalytic systems. The computational models revealed strong binding of metal particles to the carbon coating comprising efficient binding sites. The approach was tested for trapping Cr, Co, Ag, Ni, Cu, Pd, Cd, Ir, Ru and Rh nanoparticles from solutions containing micromolar starting concentrations of the metal precursors. The developed approach provides a unique tool for studying intrinsic properties of catalytic systems.

Detection of 8-Hydroxy-2'-Deoxyguanosine Biomarker with a Screen-Printed Electrode Modified with Graphene.

In this work we present the preparation of graphene material by exfoliation of graphite rods via pulses of current in electrolyte, containing a mixture of boric acid (0.05 M) and sodium chloride (0.05 M). The material was morphologically and structurally characterized by SEM/TEM/HR-TEM, XRD and FTIR techniques. TEM investigation of graphene flakes deposited onto carbon-coated grids allowed the visualization of thin and transparent regions, attributed to few-layer graphene (FLG), as well as thick and dark regions attributed to multi-layer graphene (MLG). The mixed composition of the material was additionally confirmed by XRD, which further indicated that the amount of FLG within the sample was around 83%, while MLG was around 17%. The performance of a screen-printed electrode (SPE) modified with graphene (SPE-Gr) was tested for 8-hydroxy-2′-deoxyguanosine detection. The graphene-modified electrode had a higher sensitivity in comparison with that of SPE, both in standard laboratory solutions (phosphate buffered saline—PBS) and in human saliva.

Structural Changes in Tubulin Sheets Caused by Immobilization on Solid Supports.

In the presence of zinc, the protein tubulin assembles into two-dimensional sheets that are a useful model system for the study of both tubulin and microtubule structure. Tubulin sheets present an ideal protein structure for study with atomic force microscopy because they contain a two-dimensional crystalline protein lattice and retain many of the structural features of tubulin and microtubules. However, high-resolution imaging requires nonperturbative immobilization onto an appropriate imaging substrate. In this report, several substrates commonly used for scanning probe microscopy are evaluated for their ability to effectively immobilize tubulin sheets: mica, gold, highly ordered pyrolytic graphite, and carbon-coated electron microscopy grids. We hypothesize that the different intermolecular interactions presented by these substrates will affect the morphology of adsorbed tubulin sheets as well as the amount of other contaminating adsorbates. Tubulin sheets were successfully imaged on all of these substrates and structural characterization is reported. The most consistent results were obtained on carbon-coated electron microscopy grids, which preserved fine structural features of the sheets and had the least amount of contamination from the adsorption of unpolymerized tubulin. Images of tubulin sheets obtained with atomic force microscopy also compare favorably with published electron micrographs of sheets produced using similar procedures. This work demonstrates the importance of assessing substrate effects when studying two-dimensional protein crystals and identifies suitable substrates for immobilizing tubulin sheets.

Effect of proanthocyanidin on ultrastructure and mineralization of dentine collagen

ObjectiveProanthocyanidin (PA) is a natural collagen cross-linker that has been used in dentine matrix biomodification for reparative and preventive therapies. This study evaluated the ultrastructure of collagen after its interaction with PA. Furthermore, the mineralization of PA-biomodified collagen matrix was observed. MethodsTen freshly extracted sound human molars were sectioned into 0.5mm×1.7mm×7mm beams for ultrastructural evaluation of PA and dentine matrix under Field Emission Scanning Electron Microscopy (FESEM) and Transmission Electron Microscopy (TEM). Specimens for TEM were completely demineralized and divided into three groups according to PA treatments: deionized water, 2% PA and 6.5% PA. The specimens were fixed, dehydrated, sectioned and examined using TEM. Specimens for FESEM were lightly conditioned with EDTA and similarly divided into the three groups for observation using FESEM. Type I collagen from calf skin was used to analyse the mineral interaction after treatment with 6.5% PA. Formvar- and carbon-coated 400-mesh Ni grids (EMS, Hatfiels, PA, USA) were placed over a 2mg/mL collagen solution prepared from calf skin-derived Type I collagen to achieve self-assembly of collagen fibrils. Grids were treated with 6.5% PA and divided into two groups. One group was floated over a remineralization solution containing 20mM HEPES, 2.25mM CaCl2-2H2O, 1.35mM KH2PO4, 3.08mM NaN3 and 130mM KCl and the other group was over a CPP-ACP solution (Tooth mousse 1:100 dilution with deionized water). The floating samples were kept in a 37°C and 100% humidity chamber. Grids were taken out at selected time durations (24h, 48h and 72h for mineralization solution/24h for CPP-ACP) and observed under TEM without staining. Selected area electron diffractions (SAEDs) were performed at 110kV. ResultsFollowing treatment of demineralized dentine collagen matrix with PA, the size and number of interfibrillar spaces were reduced. The collagen fibrils aggregated together with a reduction in porosity. A characteristic banding pattern of collagen fibrils was observed under TEM. Treatment of PA-biomodified collagen fibrils with remineralization solution increased mineral aggregation along its long axis, when compared to the control group. Furthermore, treatment of PA-biomodified collagen fibrils with CPP-ACP solution enhanced mineral uptake and deposition as well as initiated apatite formation within 24h. ConclusionProanthocyanidin alters the ultrastructure of demineralized dentine collagen matrix. The PA-biomodified collagen matrix promotes remineralization.

Ultrafast self-assembly of silver nanostructures on carbon-coated copper grids for surface-enhanced Raman scattering detection of trace melamine

Structurally well-defined assemblies of silver nanoparticles, including the dendritic nano-flowers (NFs), planar nano-spheres (NSs) and nano-dendrites (NDs) were obtained by a surfactant-free and ultrafast (≈15min) self-assembly process on as-purchased carbon-coated copper TEM grids. The silver nano-assemblies, especially the NFs modified TEM grids, when serving as surface-enhanced Raman spectroscopy (SERS) substrates for detecting melamine molecules, demonstrated a long-lived limit of detection (LOD) of as low as 10−11M, suggesting the potential of these silver-assemblies modified carbon-coated copper grids as novel potable and cost-effective SERS substrates for trace detection toward various food contaminants like melamine.

The collection of MicroED data for macromolecular crystallography.

The formation of large, well-ordered crystals for crystallographic experiments remains a crucial bottleneck to the structural understanding of many important biological systems. To help alleviate this problem in crystallography, we have developed the MicroED method for the collection of electron diffraction data from 3D microcrystals and nanocrystals of radiation-sensitive biological material. In this approach, liquid solutions containing protein microcrystals are deposited on carbon-coated electron microscopy grids and are vitrified by plunging them into liquid ethane. MicroED data are collected for each selected crystal using cryo-electron microscopy, in which the crystal is diffracted using very few electrons as the stage is continuously rotated. This protocol gives advice on how to identify microcrystals by light microscopy or by negative-stain electron microscopy in samples obtained from standard protein crystallization experiments. The protocol also includes information about custom-designed equipment for controlling crystal rotation and software for recording experimental parameters in diffraction image metadata. Identifying microcrystals, preparing samples and setting up the microscope for diffraction data collection take approximately half an hour for each step. Screening microcrystals for quality diffraction takes roughly an hour, and the collection of a single data set is ∼10 min in duration. Complete data sets and resulting high-resolution structures can be obtained from a single crystal or by merging data from multiple crystals.

Spatial Distributions of Discharged Products of Lithium-Oxygen Batteries Revealed by Synchrotron X-ray Transmission Microscopy.

The discharge products of ether-based Li-O2 cells were grown directly on common carbon-coated TEM grids and observed by oxidation-state-sensitive full field transmission soft X-ray microscopy (TXM). The acquired data have permitted to quantify and localize with spatial resolution the distribution of the oxygen discharge products in these samples (i.e., lithium superoxide, peroxide, and carbonates) and appreciate several compositional, structural, and morphological aspects. Most of the peroxide particles had a toroidal shape, often with a central hole usually open on only one side, and which included significant amounts of superoxide-like phases (LiO2/Li2O2 ratio between 0.2 and 0.5). Smaller particles had smaller or no superoxide content, from which we infer that abundance of soluble LiO2 may have a role in toroid formation. Significant amount of carbonates were found irregularly distributed on the electrode surface, occasionally appearing as small particles and aggregates, and mostly coating lithium peroxide particles. This suggests the formation of a barrier that, similar to the solid electrolyte interface (SEI) critical in Li-ion batteries, requires an appropriate management for a reversible operation.

Study of inelastic mean free path of metal nanostructures using energy filtered transmission electron microscopy imaging.

We report a simple method for measuring the inelastic mean free path of nanostructures of known geometry using energy filtered transmission electron microscopy imaging. The mean free path of inelastic electrons was measured by using systems having known symmetry, such as cylindrical or cubic, combined with Poisson statistics without employing the knowledge of microscope parameters, namely the convergence angle and the collection angle. Having inherent symmetry of such systems, their absolute thickness can be measured from their two-dimensional projection images. We have calculated mean free path of inelastic scattering of electrons in gold, silver and nickel doing case study research by employing gold nanorod, silver nanocube and nickel nanorod lying on a carbon-coated TEM grid at two different electron energies (viz. 200 and 300 keV) following this alternative approach. Results obtained using such alternative approach were verified using microscope parameters.

Precise estimation of HPHT nanodiamond size distribution based on transmission electron microscopy image analysis

Abstract Fluorescent nanodiamonds (NDs) are extremely promising fluorescent probes and sensors in experimental biology. For these applications, NDs obtained from high-pressure, high-temperature (HPHT) synthesis are used. Commercial HPHT NDs are polydisperse and of irregular shape. In this work, we used image analysis of transmission electron microscopy (TEM) micrographs to determine the actual size distribution of an HPHT ND sample with high precision. We developed novel TEM sample preparation procedure, which includes hydrophilization of a carbon-coated TEM grid by UV/ozone treatment and subsequent charge reversal and charge-controlled adsorption of nanoparticles. This method results in NDs spread evenly on the grid and radically improves the quality of TEM samples. Image analysis of TEM micrographs of commercial NDs revealed an unexpectedly high abundance of sub-10 nm NDs in the sample. This method can also be used for preparing TEM samples of other negatively charged nanoparticles, such as citrate-stabilized gold.

A Periodic Pattern of Evolutionarily Conserved Basic and Acidic Residues Constitutes the Binding Interface of Actin-Tropomyosin

Actin filament cytoskeletal and muscle functions are regulated by actin binding proteins using a variety of mechanisms. A universal actin filament regulator is the protein tropomyosin, which binds end-to-end along the length of the filament. The actin-tropomyosin filament structure is unknown, but there are atomic models in different regulatory states based on electron microscopy reconstructions, computational modeling of actin-tropomyosin, and docking of atomic resolution structures of tropomyosin to actin filament models. Here, we have tested models of the actin-tropomyosin interface in the "closed state" where tropomyosin binds to actin in the absence of myosin or troponin. Using mutagenesis coupled with functional analyses, we determined residues of actin and tropomyosin required for complex formation. The sites of mutations in tropomyosin were based on an evolutionary analysis and revealed a pattern of basic and acidic residues in the first halves of the periodic repeats (periods) in tropomyosin. In periods P1, P4, and P6, basic residues are most important for actin affinity, in contrast to periods P2, P3, P5, and P7, where both basic and acidic residues or predominantly acidic residues contribute to actin affinity. Hydrophobic interactions were found to be relatively less important for actin binding. We mutated actin residues in subdomains 1 and 3 (Asp(25)-Glu(334)-Lys(326)-Lys(328)) that are poised to make electrostatic interactions with the residues in the repeating motif on tropomyosin in the models. Tropomyosin failed to bind mutant actin filaments. Our mutagenesis studies provide the first experimental support for the atomic models of the actin-tropomyosin interface.

Direct observation of the electrocrystallization of compound CuInSe2 during the early stages of deposition

Fundamental understanding of the early stages of electrocrystallization proceeding to a compound material deposition is of great importance. In this study, the growth mechanism of chalcopyrite CuInSe2, which is widely applied to thin film solar cells, was investigated. The electrodeposition was performed directly on carbon-coated nickel TEM grids so that the evolution of electrocrystallization can be truly followed using TEM. High-angle annular dark-field scanning transmission electron microscopy (HAADF–STEM) reveals that nanocrystalline cubic structure Cu2−xSe is deposited within the first 4ms. After 48ms, as Cu2+ and H2SeO3 around Cu2−xSe nuclei become depleted, In3+ is then reduced and the tetragonal CuInSe2 nanocrystals start to deposit around the Cu2−xSe nuclei surface to form core–shell structures. The formation of core–shell structures containing two semiconductor materials provides an insight for electrodeposition of compound semiconductors and the composition adjustment for electrodeposited CuInSe2.