Nucleus Size Research Articles

Flow cytometric analysis for Ki67 assessment in formalin-fixed paraffin-embedded breast cancer tissue

BackgroundPathologists commonly employ the Ki67 immunohistochemistry labelling index (LI) when deciding appropriate therapeutic strategies for patients with breast cancer. However, despite several attempts at standardizing the Ki67 LI, inter-observer and inter-laboratory bias remain problematic. We developed a flow cytometric assay that employed tissue dissociation, enzymatic treatment and a gating process to analyse Ki67 in formalin-fixed paraffin-embedded (FFPE) breast cancer tissue.ResultsWe demonstrated that mechanical homogenizations combined with thrombin treatment can be used to recover efficiently intact single-cell nuclei from FFPE breast cancer tissue. Ki67 in the recovered cell nuclei retained reactivity against the MIB-1 antibody, which has been widely used in clinical settings. Additionally, since the method did not alter the nucleoskeletal structure of tissues, the nuclei of cancer cells can be enriched in data analysis based on differences in size and complexity of nuclei of lymphocytes and normal mammary cells. In a clinical study using the developed protocol, Ki67 positivity was correlated with the Ki67 LI obtained by hot spot analysis by a pathologist in Japan (rho = 0.756, P < 0.0001). The number of cancer cell nuclei subjected to the analysis in our assay was more than twice the number routinely checked by pathologists in clinical settings.ConclusionsThe findings of this study showed the application of this new flow cytometry method could potentially be used to standardize Ki67 assessments in breast cancer.

Viscoelastic properties of colorectal liver metastases reflect tumour cell viability

BackgroundColorectal cancer is the third most common tumour entity in the world and up to 50% of the patients develop liver metastases (CRLM) within five years. To improve and personalize therapeutic strategies, new diagnostic tools are urgently needed. For instance, biomechanical tumour properties measured by magnetic resonance elastography (MRE) could be implemented as such a diagnostic tool. We postulate that ex vivo MRE combined with histological and radiological evaluation of CRLM could provide biomechanics-based diagnostic markers for cell viability in tumours.Methods34 CRLM specimens from patients who had undergone hepatic resection were studied using ex vivo MRE in a frequency range from 500 Hz to 5300 Hz with increments of 400 Hz. Single frequency evaluation of shear wave speed and wave penetration rate as proxies for stiffness and viscosity was performed, along with rheological model fitting based on the spring-pot model and powerlaw exponent α, ranging between 0 (complete solid behaviour) and 1 (complete fluid behaviour). For histological analysis, samples were stained with H&E and categorized according to the degree of regression. Quantitative histologic analysis was performed to analyse nucleus size, aspect ratio, and density. Radiological response was assessed according to RECIST-criteria.ResultsFive samples showed major response to chemotherapy, six samples partial response and 23 samples no response. For higher frequencies (> 2100 Hz), shear wave speed correlated significantly with the degree of regression (p ≤ 0.05) indicating stiffer properties with less viable tumour cells. Correspondingly, rheological analysis of α revealed more elastic-solid tissue properties at low cell viability and major response (α = 0.43 IQR 0.36, 0.47) than at higher cell viability and no response (α = 0.51 IQR 0.48, 0.55; p = 0.03). Quantitative histological analysis showed a decreased nuclear area and density as well as a higher nuclear aspect ratio in patients with major response to treatment compared to patients with no response (all p < 0.05).DiscussionOur results suggest that MRE could be useful in the characterization of biomechanical property changes associated with cell viability in CRLM. In the future, MRE could be applied in clinical diagnosis to support individually tailored therapy plans for patients with CRLM.

On-Surface Synthesis of 2D Polymers at the Liquid-Solid Interface

Unraveling the structure and dynamics of the formation of covalent and non-covalent organic-based 2D crystalline materials is key to controlling the quality of these materials, such as the defect density and their size. Understanding these characteristics is important for controlling their properties. This contribution highlights our efforts in using scanning probe microscopy, particularly scanning tunneling microscopy (STM), to visualize the structure and dynamics of substrate-supported metal-organic frameworks (sMOFs) and substrate-supported covalent organic frameworks (sCOFs) at the liquid-solid interface.We will focus on the growth of covalent organic frameworks, primarily based on boroxine chemistry, and reveal both qualitative and quantitative details of the nucleation-elongation processes in real-time and under ambient conditions. Sequential data analysis enables the observation of the amorphous-to-crystalline transition, the time-dependent evolution of nuclei, the existence of 'non-classical' crystallization pathways, and, importantly, the experimental determination of essential crystallization parameters with excellent accuracy, including critical nucleus size, nucleation rate, and growth rate. Other aspects that will be addressed include sCOFs and chirality, as well as the formation of multilayered sCOFs. Additionally, we demonstrate that in specific cases, the electric field between the STM tip and the substrate can induce, on demand, the polymerization or depolymerization process.

Determination of γ/γ' interface free energy for solid state precipitation in Ni-Al alloys from molecular dynamics simulation.

Interface free energy is a fundamental material parameter needed to predict the nucleation and growth of new phases. The high cost of experimentally determining this parameter makes it an ideal target for calculation through a physically informed simulation. Direct determination of interface free energy has many challenges, especially for solid-solid transformations. Indirect determination of the interface free energy from the nucleation data has been done in the case of solidification. However, a slow on molecular dynamics (MD) simulation time scale atomic diffusion makes this method not applicable to the case of nucleation from the solid phase when precipitate composition is different from that in matrix. To address this challenge, we outline the development of a new technique for determining the critical nucleus size from an MD simulation using a recently developed method to accelerate solid-state diffusion. The accuracy of our approach for the Ni-Al system for Ni3Al (γ') precipitates in a Ni-Al (γ) matrix is demonstrated well within experimental accuracy and greatly improves upon previous computational methods [Herrnring et al., Acta Mater. 215(8), 117053 (2021)].

A General Nucleation Model for Semiconductor Nanocrystals.

We introduce a nonclassical model for nanocrystal nucleation in solution which centers on the dynamic interplay of chemical bond breakage and formation coupled with the desolvation of precursor molecules, which we term the molecular chemistry (MC) model. Departing from classical theory, our model employs the bond count as the key variable rather than particle size, thereby redefining the role of supersaturation and its role in determining the so-called critical nucleus size. We apply the model to CdSe nanocrystal formation in nonpolar solvents and showcase its efficacy in predicting solvent dynamics, precursor characteristics, crystal phase, stoichiometry, "magic number" behavior, and transition states. While the coupled-cluster method is used to determine the bond energy, we show that it is possible to derive reaction pathways by reducing the calculations to algebraic approximations for the nucleation energy. This singular set of bond energy parameters allows nanocrystal nucleation and growth to be conceptualized as a straightforward chemical reaction.

Effects of PreOperative radiotherapy in a preclinical glioblastoma model: a paradigm-shift approach.

PreOperative radiotherapy (RT) is commonly used in the treatment of brain metastasis and different cancer types but has never been used in primary glioblastoma (GBM). Here, we aim to establish, describe, and validate the use of PreOperative RT for the treatment of GBM in a preclinical model. Rat brains were locally irradiated with 30-Gy, hypofractionated in five doses 2 weeks before or after the resection of intracranial GBM. Kaplan-Meier analysis determined survival. Hematoxylin-eosin staining was performed, and nuclei size and p21 senescence marker were measured in both resected and recurrent rodent tumors. Immunohistochemistry assessed microglia/macrophage markers, and RNAseq analyzed gene expression changes in recurrent tumors. Akoya Multiplex Staining on two human patients from our ongoing Phase I/IIa trial served as proof of principle. PreOperative RT group median survival was significantly higher than PostOperative RT (p < 0.05). Radiation enlarged cytoplasm and nuclei in PreOperative RT resected tumors (p < 0.001) and induced senescence in PostOperative RT recurrent tumors (p < 0.05). Gene Set Enrichment Analysis (GSEA) suggested a more proliferative profile in PreOperative RT group. PreOperative RT showed lower macrophage/microglia recruitment in recurrent tumors (p < 0.01) compared to PostOperative RT. Akoya Multiplex results indicated TGF-ß accumulation in the cytoplasm of TAMs and CD4 + lymphocyte predominance in PostOperative group. This is the first preclinical study showing feasibility and longer overall survival using neoadjuvant radiotherapy before GBM resection in a mammalian model. This suggests strong superiority for new clinical radiation strategies. Further studies and trials are required to confirm our results.

Role of dislocations on martensitic transformation temperatures and microstructure: A molecular dynamics study

Microstructure and defects strongly affect martensitic transformations in metallic alloys. Significant progress has been made in understanding the atomic-level processes that control the role of grain boundaries and precipitates in these solid-to-solid phase transformations. Yet, the role of dislocations and their structures on martensitic transformation temperature and the resulting microstructure remains unclear. Therefore, we used large-scale molecular dynamics simulations to study the forward and reverse transformation of a martensitic material modeled after Ni63Al37 under cyclic thermal loading. The simulations reveal that dislocations in the austenite phase act as one-dimensional seeds for the martensite phase, which is present at temperatures significantly above the martensite start value. We find a reduction in the dislocation density during cyclic thermal loading, which results in the increase in martensite and austenite transition temperatures, in agreement with experiments. Importantly, we extracted a critical martensitic nuclei size for developing stable domains and found that relatively low dislocation densities are needed to grow independent martensitic variants resulting in a multi-domain structure.

Multi-headed U-Net: an automated nuclei segmentation technique using Tikhonov filter-based unsharp masking

ABSTRACT An automated nuclei segmentation is the key technique for understanding and analyzing cellular properties, which are helpful for disease diagnosis and support computer-aided digital pathology. However, this task is challenging because of the variability in nuclei size and morphology, which results in blurry boundaries and overlapping nuclei. To address such issues, a multi-headed U-Net convolutional neural network (CNN) architecture has been proposed. This architecture has multiple heads to extract multi-resolution features of the source image by using different kernel sizes of the filters. The source images are pre-processed using an unsharp masking approach based on the Tikhonov filter. The Tikhonov filter decomposes the input image into low-frequency and high-frequency band images. The unsharp masking method improves the high-frequency information of the input image by primarily enhancing features such as boundaries, contours, and fine details. We have incorporated intersection over union (IOU) and F1Score as measures along with accuracy for our proposed objective functions. The proposed objective functions are tried to be maximized by the optimization algorithm, and the higher value of the metrics indicates better segmentation performance in the spatial domain during the testing phase. The proposed method attained IOU(JI), Accuracy, Precision, and F1Score values as 0.8299, 0.9642, 0.8918, and 0.9070, respectively. The quantitative and qualitative experimental outcomes indicate that our proposed technique outperforms the state-of-the-art techniques.

Chiral Solvent-Induced Homochiral Hierarchical Molecular Assemblies at the Liquid/Solid Interface.

We herein investigate the formation of homochiral hierarchical self-assembled molecular networks (SAMNs) via chirality induction by the coadsorption of a chiral solvent at the liquid/graphite interface by means of scanning tunneling microscopy (STM). In a mixture of achiral solvents, 1-hexanoic acid, and 1,2,4-trichlorobenzene, an achiral dehydrobenzo[12]annulene (DBA) derivative with three alkoxy and three hydroxy groups in an alternating manner forms chiral hierarchical triangular cluster structures through dynamic self-sorting. Enantiomorphous domains appear in equal probability. On the other hand, in chiral 2-methyl-1-hexanoic acid as a solvent, this molecule produces (i) homochiral small triangular clusters at a low solute concentration, (ii) a chirality-biased hierarchical structure consisting of triangular cluster structures with different cluster sizes at a medium concentration, and (iii) a dense structure with no chirality bias at a high concentration. We attribute the concentration-dependent degree of the chirality transmission to the number of coadsorbed solvent molecules in the SAMNs and to the difference in nucleus structure and size in the initial stage of the SAMN formation.

Cytomorphological Features as a Subtyping Tool of Non-Small-Cell Lung Cancer in Brushing Bronchoscopic Samples.

Nowadays, the separation of adenocarcinomas (ADCs) and squamous cell carcinomas (SCCs) is crucial given that there are new specific targeted therapies. So, the aim of this study was to examine the differences in cytomorphological features between ADC and SCC in bronchoscopic brush samples. The retrospective study was conducted over a 3-year period at Western Balkan University Hospital. All brushing samples were analysed. According to the histopathological report, patients were classified into ADC and SCC groups. The cytomorphological features analysed in 95 samples were presence of necrosis, cell distribution, nuclear atypia, size of nuclei, and visibility of nucleoli. Statistical analysis was performed in JASP, and P values <0.05 were considered significant. The necrotic background was more frequent in SCC samples. Small clusters sized ≤200 µm were found in 17.95% of samples from the SCC group and 53.57% in the ADC group. Large clusters sized >400 µm were found in 43.59% in the SCC group, while in the ADC group, it was found in 5.36%. There were no differences in nuclear atypia between groups. Nuclei that were >5x lymphocyte size were found more often in samples from ADC than in the SCC group (37.50 vs 10.25%). In 89.75% of samples from the SCC group, nuclei were ≤5x lymphocyte sizes, while in the ADC group, the percentage was 63.5%. Nucleoli were more often visible in samples from the ADC group compared to the SCC group (92.86% vs 64.10%, P < 0,05). Small clusters, large nuclei, and visible nucleoli were more frequent in the ADC group (P < 0.05), while large clusters, small nuclei, and invisible nucleoli were more frequent in the SCC group (P < 0.05).

Predictions for Sparse Photometry of Jupiter-family Comet Nuclei in the LSST Era

The Legacy Survey of Space and Time (LSST) at Vera C. Rubin Observatory will deliver high-quality, temporally sparse observations of millions of solar system objects on an unprecedented scale. Such data sets will likely enable the precise estimation of small-body properties on a population-wide basis. In this work, we consider the possible applications of photometric data points from LSST to the characterization of Jupiter-family comet (JFC) nuclei. We simulate sparse-in-time lightcurve points with an LSST-like cadence for the orbit of a JFC between 2024 and 2033. Convex lightcurve inversion is used to assess whether the simulation input parameters can be accurately reproduced for a sample of nucleus rotation periods, pole orientations, activity onsets, shapes, and sizes. We find that the rotation period and pole direction can be reliably constrained across all nucleus variants tested, and that the convex shape models, while limited in their ability to describe complex or bilobed nuclei, are effective for correcting sparse photometry for rotational modulation to improve estimates of nucleus phase functions. Based on this analysis, we anticipate that LSST photometry will significantly enhance our present understanding of the spin state and phase function distributions of JFC nuclei.

Design of iPSC-based cell model to study the functions of the &lt;i&gt;UBE2A&lt;/i&gt; gene

BACKGROUND: The UBE2A protein belongs to the E2 family of ubiquitin-binding enzymes involved in the ubiquitination of substrate proteins. UBE2A mutations lead to congenital X-linked mental retardation syndrome-type Nascimento. How UBE2A participates in the central nervous system development is still unknown. AIM: To establish a cell model based on induced pluripotent stem cells (iPSCs) to study the molecular and cellular functions of UBE2A in neurogenesis. METHODS: Using genomic CRISPR-Cas9 editing and lentiviral transduction, a cell model based on iPSCs from two healthy donors was designed. This cell model includes isogenic iPSCs with knockout and inducible hyperexpression of UBE2A. In addition, iPSCs were obtained by reprogramming peripheral blood mononuclear cells of a patient diagnosed with X-linked mental retardation of Nascimento type, which has a deletion spanning the whole UBE2A locus. RESULTS: The obtained iPSCs demonstrate an ESC-like morphology. They express pluripotent cell markers OCT4, SOX2, SSEA-4, and TRA-1-81 and have normal karyotypes. iPSCs with UBE2A knockout or hyperexpression had significantly increased nuclei size compared with the isogenic control. CONCLUSION: The developed iPSC-based cell model can be used for fundamental studies of the functions of UBE2A in neurogenesis.

Response to Selection of Indigenous Chicken in Rwanda Using Within-Breed Selection Strategy

The study evaluated response to selection from within-breed selection strategy for conventional (&amp;lt;I&amp;gt;CBS&amp;lt;/I&amp;gt;) and genomic (&amp;lt;I&amp;gt;GBS&amp;lt;/I&amp;gt;) breeding schemes. These breeding schemes were evaluated in both high-health environments (nucleus) and smallholder farms (commercial). Breeding goal was to develop a dual-purpose IC for both eggs and meat through selective breeding. Breeding objectives were body weight (BW), egg weight (EW), egg number (EN) and resistance to Newcastle disease (AbR). A deterministic simulation was performed to evaluate rates of genetic gain and inbreeding. Base population in the nucleus was made up of 40 cockerels and 200 pullets. Selection pressure was 4% and 20% in the males and the females, respectively. The impact of nucleus size and selection pressure on rates of genetic gain and inbreeding of the breeding program was investigated through sensitivity analysis. SelAction software was used to predict rates of genetic gain and inbreeding. Results showed that using &amp;lt;I&amp;gt;CBS&amp;lt;/I&amp;gt; in the nucleus, the breeding goal was 340.41\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\$ and 1.13 times higher than that in the commercial flock. Inbreeding rate per generation of selected chicken in the nucleus was 1.45% and lower by 1.32 times compared to their offspring under smallholder farms. Genetic gains per generation in the nucleus for BW and EN traits were 141.10 g and 1.07 eggs and 1.12 and 1.10 times greater than those in smallholder farms, respectively. With &amp;lt;I&amp;gt;GBS&amp;lt;/I&amp;gt;, the breeding goal was increased by 3.00 times whereas inbreeding rate was reduced by 3.15 times. Besides, using &amp;lt;I&amp;gt;GBS&amp;lt;/I&amp;gt;, the selected birds in the nucleus were relatively similar to those in a commercial environment. Finally, the study revealed that the nucleus size and mating ratio influence the rates of genetic gain and inbreeding in both &amp;lt;I&amp;gt;GBS&amp;lt;/I&amp;gt; and &amp;lt;I&amp;gt;CBS.&amp;lt;/I&amp;gt; This study shows that IC in Rwanda have the potential to be improved through within-breed selection strategy using either &amp;lt;I&amp;gt;CBS&amp;lt;/I&amp;gt; or &amp;lt;I&amp;gt;GBS&amp;lt;/I&amp;gt;.

Late gestation fetal hypothyroidism alters cell cycle regulation across multiple organ systems

BackgroundHypothyroidism is a common endocrine disruption observed in utero that adversely affects fetal growth and maturation leading to long-term impacts on health; however, the exact molecular mechanisms by which these deleterious effects occur are unknown. We hypothesize that fetal hypothyroidism during late gestation will disrupt cell cycle regulation in a tissue-specific manner. To evaluate this, eight pregnant gilts were dosed with either methimazole or an equivalent negative control during days 85–106 out of 114 days of gestation (n = 4/group). Following treatment, the gilts were humanely euthanized, and tissue samples of fetal heart, ileum, kidney, lung, liver, muscle, spleen, and thymus taken from two male and two female fetuses (n = 32) from each gilt.ResultsThe relative expression of three cell cycle promoters (CDK1, CDK2, and CDK4), and one cell cycle inhibitor (CDKN1A) was compared in each tissue to determine the effect of hypothyroidism on the developing fetus. All of the eight tissues examined experienced at least one significant up- or downregulation in the expression of the aforementioned genes as a result of treatment with methimazole. Substantial changes were observed in the liver and muscle, with the latter experiencing significant downregulations of CDK1, CDK2, and CDK4 as a result of treatment. In addition, all tissues were examined for changes in protein content, which further elucidated the impact of hypothyroidism on the fetal liver by the observation of a marked increase in protein content in the methimazole-treated group. Finally, the heart and liver were histologically examined for evidence of cellular hyperplasia and hypertrophy by measuring average nuclei density and size in each tissue, with the results showing a significant decrease in average nuclei size in the liver of hypothyroid fetuses.ConclusionsCollectively, these findings indicate the occurrence of organ-specific disruptions in cell cycle progression as a result of in utero hypothyroidism, which may explain the long term and widespread effects of hypothyroidism on fetal development.

Increased vascular smooth muscle cell senescence in aneurysmal Fibulin-4 mutant mice

Aortic aneurysms are dilatations of the aorta that can rupture when left untreated. We used the aneurysmal Fibulin-4R/R mouse model to further unravel the underlying mechanisms of aneurysm formation. RNA sequencing of 3-month-old Fibulin-4R/R aortas revealed significant upregulation of senescence-associated secretory phenotype (SASP) factors and key senescence factors, indicating the involvement of senescence. Analysis of aorta histology and of vascular smooth muscle cells (VSMCs) in vitro confirmed the senescent phenotype of Fibulin-4R/R VSMCs by revealing increased SA-β-gal, p21, and p16 staining, increased IL-6 secretion, increased presence of DNA damage foci and increased nuclei size. Additionally, we found that p21 luminescence was increased in the dilated aorta of Fibulin-4R/R|p21-luciferase mice. Our studies identify a cellular aging cascade in Fibulin-4 aneurysmal disease, by revealing that Fibulin-4R/R aortic VSMCs have a pronounced SASP and a senescent phenotype that may underlie aortic wall degeneration. Additionally, we demonstrated the therapeutic effect of JAK/STAT and TGF-β pathway inhibition, as well as senolytic treatment on Fibulin-4R/R VSMCs in vitro. These findings can contribute to improved therapeutic options for aneurysmal disease aimed at reducing senescent cells.

Fluorescent carbon nanodot-based rapid cell nucleus staining and application for diagnosis.

Cell nucleus status decides the activities of corresponding cells, making its rapid and effective staining important for revealing the actual condition of biological environment in life science and related fields. In this study, fast staining of cell nucleus is realized by fluorescent carbon nanodots (CDs). The staining mechanism is due to the positively charged CD surface-induced cell membrane penetration, which facilitates the CD-nucleus binding via electrostatic attraction. The size of cell nucleus is easily measured with fluorescence imaging technique. In addition, the CD-based cell nucleus stain is applied for discriminating the normal and cancer cells by determining the cell-to-nucleus ratio with fluorescence images.

Robust nuclei segmentation with encoder‐decoder network from the histopathological images

AbstractNuclei segmentation is a prerequisite and an essential step in cancer detection and prognosis. Automatic nuclei segmentation from the histopathological images is challenging due to nuclear overlap, disease types, chromatic stain variability, and cytoplasmic morphology differences. Furthermore, it is demanding to develop a single accurate method for segmenting nuclei of different organs because of the diversity in nuclei size, shape, and appearance across the various organs. To address these challenges, we developed a robust Encoder‐Decoder network for nuclei segmentation from the multi‐organ histopathological images. In this approach, we utilize a pre‐trained EfficientNet‐B4 as an Encoder subnetwork and design a new Decoder subnetwork architecture. Additionally, we have applied morphological operation‐based post‐processing to improve the segmentation results. The performance of our approach has been evaluated on three public datasets, namely, Kumar, TNBC, and CPM‐17 datasets, which contain histopathological images of seven organs, one organ, and four organs, respectively. The proposed method achieved an aggregated Jacquard index of 0.636, 0.611, and 0.706 on Kumar, TNBC, and CPM‐17 datasets, respectively. Our proposed approach also shows superiority over the existing methods.

Integrating Spatial and Morphological Characteristics into Melanoma Prognosis: A Computational Approach.

In this study, the prognostic value of cellular morphology and spatial configurations in melanoma has been examined, aiming to complement traditional prognostic indicators like mitotic activity and tumor thickness. Through a computational pipeline using machine learning and deep learning methods, we quantified nuclei sizes within different spatial regions and analyzed their prognostic significance using univariate and multivariate Cox models. Nuclei sizes in the invasive band demonstrated a significant hazard ratio (HR) of 1.1 (95% CI: 1.03, 1.18). Similarly, the nuclei sizes of tumor cells and Ki67 S100 co-positive cells in the invasive band achieved HRs of 1.07 (95% CI: 1.02, 1.13) and 1.09 (95% CI: 1.04, 1.16), respectively. Our findings reveal that nuclei sizes, particularly in the invasive band, are potentially prognostic factors. Correlation analyses further demonstrated a meaningful relationship between cellular morphology and tumor progression, notably showing that nuclei size within the invasive band correlates substantially with tumor thickness. These results suggest the potential of integrating spatial and morphological analyses into melanoma prognostication.

Guanidine Hydrochloride-Induced Hepatitis B Virus Capsid Disassembly Hysteresis.

Hepatitis B virus (HBV) displays remarkable self-assembly capabilities that interest the scientific community and biotechnological industries as HBV is leading to an annual mortality of up to 1 million people worldwide (especially in Africa and Southeast Asia). When the ionic strength is increased, hepatitis B virus-like particles (VLPs) can assemble from dimers of the first 149 residues of the HBV capsid protein core assembly domain (Cp149). Using solution small-angle X-ray scattering, we investigated the disassembly of the VLPs by titrating guanidine hydrochloride (GuHCl). Measurements were performed with and without 1 M NaCl, added either before or after titrating GuHCl. Fitting the scattering curves to a linear combination of atomic models of Cp149 dimer (the subunit) and T = 3 and T = 4 icosahedral capsids revealed the mass fraction of the dimer in each structure in all the titration points. Based on the mass fractions, the variation in the dimer-dimer association standard free energy was calculated as a function of added GuHCl, showing a linear relation between the interaction strength and GuHCl concentration. Using the data, we estimated the energy barriers for assembly and disassembly and the critical nucleus size for all of the assembly reactions. Extrapolating the standard free energy to [GuHCl] = 0 showed an evident hysteresis in the assembly process, manifested by differences in the dimer-dimer association standard free energy obtained for the disassembly reactions compared with the equivalent assembly reactions. Similar hysteresis was observed in the energy barriers for assembly and disassembly and the critical nucleus size. The results suggest that above 1.5 M, GuHCl disassembled the capsids by attaching to the protein and adding steric repulsion, thereby weakening the hydrophobic attraction.

Recovery of angular scattering profiles through a flexible multimode fiber.

Endoscopic angle-resolved light scattering methods have been developed for early cancer detection but they typically require multi-element coherent fiber optic bundles to recover scattering distributions from tissues. Recent work has focused on using a single multimode fiber (MMF) to measure angle resolved scattering but this approach has practical limitations to overcome before clinical translation. Here we address these limitations by proposing an MMF-based endoscope capable of measuring angular scattering patterns suitable for determining structure. Significantly, this approach implements a spectrally resolved detection scheme to reduce speckle and leverages the azimuthal symmetry of the angular scattering patterns to enable measurements that are robust to fiber bending. This results in a unique method that does not require matrix inversion or machine learning to measure a transmitted scattering distribution. The MMF utilized here is 1000 mm in length with a 200 µm core and is demonstrated to recover angular scattering distributions even with bending displacements of up to 30 cm. This advance has a significant impact on the clinical translation of biomedical endoscopic diagnostic techniques that use angular scattering to determine the size of cell nuclei to detect early cancer.