The 2012 study by Zhu et al. (DOI: 10.1002/anie.201109089) reporting a hybrid nanosystem composed of carbon and CdSe/ZnS quantum dots functionalized with tris(pyridin-2-ylmethyl)ethane-1,2-diamine (CdSe@C-TPEA) as ratiometric fluorescent sensors for intracellular copper imaging remains highly influential, with 499 citations to date (Web of Science, December 4, 2025). As part of the NanoBubbles replication project, we attempted to reproduce key experimental findings following detailed pre-registered protocols. Despite strict adherence to the reported synthesis procedures, we failed to replicate the central claim of Cu2+-induced fluorescence quenching. Furthermore, our analysis reveals significant data anomalies in the original publication, including spectral overlaps with identical noise patterns inconsistent with independent measurements. These findings raise critical concerns regarding data integrity and reproducibility and cast significant doubt on the reliability of Zhu et al.'s reported results.

The NanoBubbles Reproducibility Initiative investigates the robustness of highly cited articles in nanobioscience. Here, we document the process of setting up the initiative from corpus creation to publishing results. We also present platforms that we adopted and challenges that we faced. Lessons drawn from this study can shape future studies, including experimental designs and reporting standards, and encourage open dialogue and collaboration.

This study presents the results of a replication effort aimed at reproducing key findings fromthe 2012 article "Carbon-Dot-Based Dual-Emission Nanohybrid Produces a Ratiometric Flu-orescent Sensor for In Vivo Imaging of Cellular Copper Ions" by Zhu et al. The original studyclaimed that CQDs functionalized with AE-TPEA (CQDs-TPEA) and their hybrid withCdSe/ZnS quantum dots (CdSe@C-TPEA) exhibited ratiometric fluorescence quenching uponexposure to Cu2+ ions, enabling their application as a dual-emission sensor for intracellular Cu2+imaging. Despite closely following the described synthesis and characterization protocols, wewere unable to reproduce the Cu2+-induced fluorescence quenching, undermining the primaryclaim of the original study and making it impossible to pursue the investigation in live cells.Furthermore, upon close examination of the original data, concerns regarding spectral anoma-lies and potential inconsistencies were raised, prompting us to abort the replication study.

In hundreds of articles published over the past two decades, nanoparticles have been described as probes for sensing and imaging of a variety of intracellular cytosolic targets. However, nanoparticles generally enter cells by endocytosis with only a small fraction reaching the cytosol. Most of those articles do not describe the mechanisms of nanoparticles entry into the cell and therefore the paradox of sensing a target in the cytosol when most particles do not access that compartment remains. To address this paradox, we are initiating a replication project of some of the most influential articles in this field with the aim of confirming their ability to detect their targets and getting additional insights into their intracellular localisation. This article is a pre-registered report for the second replication of this project, namely the replication of Seferos et al ‘Nano-Flares: Probes for Transfection and mRNA Detection in Living Cells’. To achieve our aim, we will carry out both a direct replication of the experiments in the original article, with the addition of some experiments and controls (in particular optical and electron microscopy). We hope that this study will contribute to a better understanding of the intracellular fate of nanoparticles and, with the help of the scientific community, help set standards in the design and reporting of studies in this field.

We aim to foster a discussion of science correction and of how individual researchers can improve the quality and control of scientific production. This is crucial because although the maintenance of rigorous standards and the scrupulous control of research findings and methods are sometimes taken for granted, in practice, we are routinely confronted with articles that contain errors.

Tracking the fate of therapeutic cell types is important for assessing their safety and efficacy. Bioluminescence imaging (BLI) is an effective cell tracking technique, but poor spatial resolution means it has limited ability to precisely map cells in vivo in 3D. This can be overcome by using a bimodal imaging approach that combines BLI with a technique capable of generating high-resolution images. Here we compared the effectiveness of combining either multispectral optoacoustic tomography (MSOT) or micro-computed tomography (micro-CT) with BLI for tracking the fate of luciferase+ human mesenchymal stromal cells (MSCs) labelled with gold nanorods. Following subcutaneous administration in mice, the MSCs could be readily detected with MSOT but not with micro-CT. We conclude that MSOT is more sensitive than micro-CT for tracking gold nanorod-labelled cells in vivo and depending on the route of administration, can be used effectively with BLI to track MSC fate in mice.

In hundreds of articles published over the past two decades, nanoparticles have been described as probes for sensing and imaging of a variety of intracellular cytosolic targets. However, nanoparticles generally enter cells by endocytosis with only a small fraction reaching the cytosol. Most of those articles do not describe the mechanisms of nanoparticles entry into the cell and therefore the paradox of sensing a target in the cytosol when most particles do not access that compartment remains. To address this paradox, we are initiating a replication project of some of the most influential articles in this field with the aim of confirming their ability to detect their targets and getting additional insights into their intracellular localisation. Thus, this article is a pre-registered report for the first replication of this project, namely the replication of Zhu et al ‘Carbon‐Dot‐Based Dual‐Emission Nanohybrid Produces a Ratiometric Fluorescent Sensor for In Vivo Imaging of Cellular Copper Ions’. To achieve our aim, we will carry out both a direct replication of the experiments in the original article, with the addition of some experiments and controls, and a conceptual replication where we test the claim that this ratiometric probe is able to sense intracellular copper levels in a more biologically relevant system. We hope that this study will contribute to a better understanding of the intracellular fate of nanoparticles and, with the help of the scientific community, help set standards in the design and reporting of studies in this field.

Cells expressing firefly luciferase that were labelled with gold nanorods (GNRs) could be visualised with multispectral optoacoustic tomography (MSOT), whereas control cells did not generate an MSOT signal. The white line in the left image indicates the imaging plain in MSOT. By combining these two imaging modalities, we obtain information about cell viability (bioluminescence) whereas MSOT shows the precise location of the cells. For further details please visit the article by Alejandra Hernandez Pichardo, James Littlewood, Arthur Taylor, Bettina Wilm, Raphaël Lévy, and Patricia Murray ( e202300109 ). image

From growing cells in spheroids to arranging them on complex engineered scaffolds, three-dimensional cell culture protocols are rapidly expanding and diversifying. While these systems may often improve the physiological relevance of cell culture models, they come with technical challenges, as many of the analytical methods used to characterize traditional two-dimensional (2D) cells must be modified or replaced to be effective. Here we review the advantages and limitations of quantification methods based either on biochemical measurements or microscopy imaging. We focus on the most basic of parameters that one may want to measure, the number of cells. Precise determination of this number is essential for many analytical techniques where measured quantities are only meaningful when normalized to the number of cells (e.g. cytochrome p450 enzyme activity). Thus, accurate measurement of cell number is often a prerequisite to allowing comparisons across different conditions (culturing conditions or drug and treatment screening) or between cells in different spatial states. We note that this issue is often neglected in the literature with little or no information given regarding how normalization was performed, we highlight the pitfalls and complications of quantification and call for more accurate reporting to improve reproducibility.

Mesenchymal stromal cells (MSCs) injected intravenously are trapped in the capillaries of the lungs and die within the first 24 h. Studying the biodistribution and fate of labelled therapeutic cells in the 3D pulmonary context is important to understand their function in this organ and gain insights into their mechanisms of action. Optical tissue clearing enables volumetric cell tracking at single-cell resolution. Thus, we compared three optical tissue-clearing protocols (Clear, Unobstructed Brain/Body Imaging Cocktails and Computational analysis (CUBIC), modified stabilised 3D imaging of solvent-cleared organs (s-DISCO) and ethyl cinnamate (ECi)) to evaluate their potential to track the biodistribution of human umbilical cord MSCs expressing the tdTomato fluorescence reporter and investigate how they interact with host cells in the mouse lung. The results showed that although CUBIC clearing is the only method that enables direct imaging of fluorescently labelled MSCs, combining s-DISCO or ECi with immunofluorescence or dye labelling allows the interaction of MSCs with endothelial and immune cells to be studied. Overall, this comparative study offers guidance on selecting an optical tissue-clearing method for cell tracking applications.

Elucidating the mechanisms of action and long-term safety of cell therapies is necessary for their clinical translation. Non-invasive imaging technologies such as bioluminescence imaging (BLI), computed tomography (CT) and multispectral optoacoustic tomography (MSOT) have been proposed as tools for longitudinal cell monitoring but their performances have not been compared. Here, we evaluate combinations of these modalities to track the in vivo distribution of gold-labelled mesenchymal stromal cells (MSCs). We found that injected MSCs labelled with gold nanoparticles and expressing the reporter gene firefly luciferase could be detected with BLI and MSOT but not CT. We conclude that the MSCs did not carry enough contrast agent to be tracked by CT, demonstrating that CT tracking of gold-labelled cells is not a practical approach as high amounts of gold, which might impair cell viability, are necessary.

We call for retraction of the 2008 Lancet article on “clinical transplantation of a tissue-engineered airway” by Macchiarini and colleagues.1 The paper received internationalmedia coverage as the “world’s first tissue engineered organ transplant,” and one of the authors, Martin Birchall, told the BBC that “in 20 years’ time, virtually any transplant organ could be made in this way.”2 Reality did not match the hype because nearly all subsequent patientswho received “tissue engineered airways” died.3 The Lancet was informed in May 2018 that the key findings of the article were false. Despite this, and subsequent demands for retraction of thepaper byus andothers, the Lancet has refused to retract, without providing any explanation.

Polyethylene terephthalate (PET) is widely used to elaborate biomaterials and medical devices in particular for long-term implant applications but tuning their surface properties remains challenging. We investigate surface functionalization by grafting poly(sodium 4-styrene sulfonate, PNaSS) with the aim of enhancing protein adhesion and cellular activity. Elucidating the topography and molecular level organization of the modified surfaces is important for understanding and predicting biological activity. In this work, we explore several grafting methods including thermal grafting, thermal grafting in the presence of Mohr's salt, and UV activation. We characterize the different surfaces obtained using atomic force microscopy (AFM), contact angle (CA), and x-ray photoelectron spectroscopy (XPS). We observe an increase in the percentage of sulfur atoms (XPS) that correlates with changes in (CA), and we identify by AFM characteristic features, which we interpret as patches of polymers on the PET surfaces. This work demonstrates tuning of biomaterials surface by functionalization and illustrates the capability of AFM to provide insights into the spatial organization of the grafted polymer.

This data set includes all the raw data collected for the following article: "Comparison between optical tissue clearing methods for detecting administered mesenchymal stromal cells in mouse lungs".

Propagation of small amyloid beta (Aβ) aggregates (or seeds) has been suggested as a potential mechanism of Alzheimer's disease progression. Monitoring the propagation of Aβ seeds in an organism would enable testing of this hypothesis and, if confirmed, provide mechanistic insights. This requires a contrast agent for long-term tracking of the seeds. Gold nanorods combine several attractive features for this challenging task, in particular, their strong absorbance in the infrared (enabling optoacoustic imaging) and the availability of several established protocols for surface functionalisation. In this work, polymer-coated gold nanorods were conjugated with anti-Aβ antibodies and attached to pre-formed Aβ seeds. The resulting complexes were characterised for their optical properties by UV/Vis spectroscopy and multispectral optoacoustic tomography. The complexes retained their biophysical properties, i.e. their ability to seed Aβ fibril formation. They remained stable in biological media for at least 2 days and showed no toxicity to SH-SY5Y neuroblastoma cells up to 1.5 nM and 6 μM of gold nanorods and Aβ seeds, respectively. Taken together, this study describes the first steps in the development of probes for monitoring the spread of Aβ seeds in animal models.

Polycaprolactoneand polyethylene terephthalate are widely used to elaborate biomaterials andmedical devices in particular for long-term implant applications but tuning theirsurface properties remains challenging. We investigate surfacefunctionalization by grafting poly(sodium 4-styrene sulfonate) with the aim ofenhancing protein adhesion and cellular activity. Elucidating the topographyand molecular level organization of the modified surfaces is important forunderstanding and predicting biological activity. In this work, we explore severalgrafting methods including thermal grafting, thermal grafting in the presenceof Mohr's salt, and UV activation. We characterize the different surfacesobtained using atomic force microscopy, contact angle and X-ray photoelectron spectroscopy. The resultsreveal striking differences in the properties of the modified surfaces. This workdemonstrates tuning of biomaterials surface by functionalization and thecapability of atomic force microscopy to provide insights into the conformationand mechanical properties of the grafted polymers.

In this paper co-authored by a philosopher and a scientist, the authors aim to provide some philosophical and scientific insights into the long life of unicorns, such as how deep misconceptions can often persist, and some for a long time, in the scientific literature. We take as an example the belief, often seen in the bio-nano and nanomedicine literature, that nanoparticles have some special abilities at crossing lipid membranes.

Studying and characterising tumour cell migration is critical for understanding disease progression and for assessing drug efficacy. Whilst tumour cell migration occurs fundamentally in 3 spatial dimensions (3D), for practical reasons, most migration studies to date have performed analysis in 2D. Here we imaged live multicellular tumour spheroids with lightsheet fluorescence microscopy to determine cellular migration and invasion in 3D over time (4D). We focused on glioblastoma, which are aggressive brain tumours, where cell invasion into the surrounding normal brain remains a major clinical challenge. We developed a workflow for analysing complex 3D cell movement, taking into account migration within the spheroid as well as invasion into the surrounding matrix. This provided metrics characterising cell motion, which we used to evaluate the efficacy of chemother-apeutics on invasion. These rich datasets open avenues for further studies on drug efficacy, microenvironment composition, as well as collective cell migration and metastatic potential.