Search results for: Stijn Luca

Authors: Silvia Gastaldello, Maria Grillo, Luca Tassoni, Claudio Maran, Stefano Balbo

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Antioxidants are nowadays attractive not only for the countless benefits to the human and animal health, but also for the perspective of use as food preservative instead of synthetic chemical molecules. In this study, the radical scavenging activity of six protein extracts from pulse and oleaginous seeds was evaluated. The selected matrices are Pisum sativum (yellow pea from two different origins), Carthamus tinctorius (safflower), Helianthus annuus (sunflower), Lupinus luteus cv Mister (lupin) and Glycine max (soybean), since they are economically interesting for both human and animal nutrition. The seeds were grinded and proteins extracted from 20mg powder with a specific vegetal-extraction kit. Proteins have been quantified through Bradford protocol and scavenging activity was revealed using DPPH assay, based on radical DPPH (2,2-diphenyl-1-picrylhydrazyl) absorbance decrease in the presence of antioxidants molecules. Different concentrations of the protein extract (1, 5, 10, 50, 100, 500 µg/ml) were mixed with DPPH solution (DPPH 0,004% in ethanol 70% v/v). Ascorbic acid was used as a scavenging activity standard reference, at the same six concentrations of protein extracts, while DPPH solution was used as control. Samples and standard were prepared in triplicate and incubated for 30 minutes in dark at room temperature, the absorbance was read at 517nm (ABS30). Average and standard deviation of absorbance values were calculated for each concentration of samples and standard. Statistical analysis using t-students and p-value were performed to assess the statistical significance of the scavenging activity difference between the samples (or standard) and control (ABSctrl). The percentage of antioxidant activity has been calculated using the formula [(ABSctrl-ABS30)/ABSctrl]*100. The obtained results demonstrate that all matrices showed antioxidant activity. Ascorbic acid, used as standard, exhibits a 96% scavenging activity at the concentration of 500 µg/ml. At the same conditions, sunflower, safflower and yellow peas revealed the highest antioxidant performance among the matrices analyzed, with an activity of 74%, 68% and 70% respectively (p < 0.005). Although lupin and soybean exhibit a lower antioxidant activity compared to the other matrices, they showed a percentage of 46 and 36 respectively. All these data suggest the possibility to use undervalued edible matrices as antioxidants source. However, further studies are necessary to investigate a possible synergic effect of several matrices as well as the impact of industrial processes for a large-scale approach.

Keywords: antioxidants, DPPH assay, natural matrices, vegetal proteins

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Authors: Elham Azimzadeh, Hamidollah Hassanlouei, Hadi Nobari, Georgian Badicu, Jorge Pérez-Gómez, Luca Paolo Ardigò

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Children with cerebral palsy (CP) have weak physical abilities and their limitations may have an effect on performing everyday motor activities. One of the most important and common debilitating factors in CP is the malfunction in the upper extremities to perform motor skills and there is strong evidence that task-specific training may lead to improve general upper limb function among this population. However, augmented feedback enhances the acquisition and learning of a motor task. Practice conditions may alter the difficulty, e.g., the reduced frequency of PG could be more challenging for this population to learn a motor task. So, the purpose of this study was to investigate the effect of physical guidance (PG) on learning a tracking task in children with cerebral palsy (CP). Twenty-five independently ambulant children with spastic hemiplegic CP aged 7-15 years were assigned randomly to five groups. After the pre-test, experimental groups participated in an intervention for eight sessions, 12 trials during each session. The 0% PG group received no PG; the 25% PG group received PG for three trials; the 50% PG group received PG for six trials; the 75% PG group received PG for nine trials; and the 100% PG group, received PG for all 12 trials. PG consisted of placing the experimenter's hand around the children's hand, guiding them to stay on track and complete the task. Learning was inferred by acquisition and delayed retention tests. The tests involved two blocks of 12 trials of the tracking task without any PG being performed by all participants. They were asked to make the movement as accurate as possible (i.e., fewer errors) and the number of total touches (errors) in 24 trials was calculated as the scores of the tests. The results showed that the higher frequency of PG led to more accurate performance during the practice phase. However, the group that received 75% PG had significantly better performance compared to the other groups in the retention phase. It is concluded that the optimal frequency of PG played a critical role in learning a tracking task in children with CP and likely this population may benefit from an optimal level of PG to get the appropriate amount of information confirming the challenge point framework (CPF), which state that too much or too little information will retard learning a motor skill. Therefore, an optimum level of PG may help these children to identify appropriate patterns of motor skill using extrinsic information they receive through PG and improve learning by activating the intrinsic feedback mechanisms.

Keywords: cerebral palsy, challenge point framework, motor learning, physical guidance, tracking task

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Authors: Chrislaura Carmo, Luca Deiana, Mafalda Laranjo, Abilio Sobral, Armando Cordova

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Photodynamic therapy (PDT) is currently used for the treatment of malignancies and premalignant tumors. It is based on the capture of a photosensitizing molecule (PS) which, when excited by light at a certain wavelength, reacts with oxygen and generates oxidizing species (radicals, singlet oxygen, triplet species) in target tissues, leading to cell death. BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indaceno) derivatives are emerging as important candidates for photosensitizer in photodynamic therapy of cancer cells due to their high triplet quantum yield. Today these dyes are relevant molecules in photovoltaic materials and fluorescent sensors. In this study, it will be demonstrated the possibility that BODIPY can be covalently linked to thioglycolic acid through the click reaction. Thiol−ene click chemistry has become a powerful synthesis method in materials science and surface modification. The design of biobased allyl-terminated precursors with high renewable carbon content for the construction of the thiol-ene polymer networks is essential for sustainable development and green chemistry. The work aims to synthesize the BODIPY (10-(4-(allyloxy) phenyl)-2,8-diethyl-5,5-difluoro-1,3,7,9-tetramethyl-5H-dipyrrolo[1,2-c:2',1'-f] [1,3,2] diazaborinin-4-ium-5-uide) and to click reaction with Thioglycolic acid. BODIPY was synthesized by the condensation reaction between aldehyde and pyrrole in dichloromethane, followed by in situ complexation with BF3·OEt2 in the presence of the base. Then it was functionalized with allyl bromide to achieve the double bond and thus be able to carry out the click reaction. The thiol−ene click was performed using DMPA (2,2-Dimethoxy-2-phenylacetophenone) as a photo-initiator in the presence of UV light (320–500 nm) in DMF at room temperature for 24 hours. Compounds were characterized by standard analytical techniques, including UV-Vis Spectroscopy, 1H, 13C, 19F NMR and mass spectroscopy. The results of this study will be important to link BODIPY to polymers through the thiol group offering a diversity of applications and functionalization. This new molecule can be tested as third-generation photosensitizers, in which the dye is targeted by antibodies or nanocarriers by cells, mainly in cancer cells, PDT and Photodynamic Antimicrobial Chemotherapy (PACT). According to our studies, it was possible to visualize a click reaction between allyl BODIPY and thioglycolic acid. Our team will also test the reaction with other thiol groups for comparison. Further, we will do the click reaction of BODIPY with a natural polymer linked with a thiol group. The results of the above compounds will be tested in PDT assays on various lung cancer cell lines.

Keywords: bodipy, click reaction, thioglycolic acid, allyl, thiol-ene click

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Authors: Shilpa Sahoo, Michel Louge, Anthony Reeves, Olivier Desjardins, Susan Daniel, Sadik Omowunmi

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The microgravity on the International Space Station (ISS) was exploited to study the imbibition of water into a network of hydrophilic cylindrical capillaries on time and length scales long enough to observe details hitherto inaccessible under Earth gravity. When a drop touches a porous medium, it spreads as if laid on a composite surface. The surface first behaves as a hydrophobic material, as liquid must penetrate pores filled with air. When contact is established, some of the liquid is drawn into pores by a capillarity that is resisted by viscous forces growing with length of the imbibed region. This process always begins with an inertial regime that is complicated by possible contact pinning. To study imbibition on Earth, time and distance must be shrunk to mitigate gravity-induced distortion. These small scales make it impossible to observe the inertial and pinning processes in detail. Instead, in the International Space Station (ISS), astronaut Luca Parmitano slowly extruded water spheres until they touched any of nine capillary plates. The 12mm diameter droplets were large enough for high-speed GX1050C video cameras on top and side to visualize details near individual capillaries, and long enough to observe dynamics of the entire imbibition process. To investigate the role of contact pinning, a text matrix was produced which consisted nine kinds of porous capillary plates made of gold-coated brass treated with Self-Assembled Monolayers (SAM) that fixed advancing and receding contact angles to known values. In the ISS, long-term microgravity allowed unambiguous observations of the role of contact line pinning during the inertial phase of imbibition. The high-speed videos of spreading and imbibition on the porous plates were analyzed using computer vision software to calculate the radius of the droplet contact patch with the plate and height of the droplet vs time. These observations are compared with numerical simulations and with data that we obtained at the ESA ZARM free-fall tower in Bremen with a unique mechanism producing relatively large water spheres and similarity in the results were observed. The data obtained from the ISS can be used as a benchmark for further numerical simulations in the field.

Keywords: droplet imbibition, hydrophilic surface, inertial phase, porous medium

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Authors: Paola Modesto, Floriana Fruscione, Isabella Martini, Simona Perga, Federica Riccardo, Mariateresa Camerino, Davide Giacobino, Cecilia Gola, Luca Licenziato, Elisabetta Razzuoli, Katia Varello, Lorella Maniscalco, Elena Bozzetta, Angelo Ferrari

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Canine and human melanoma, osteosarcoma (OSA), and mammary carcinomas are aggressive tumors with common characteristics making dogs a good model for comparative oncology. Novel therapeutic strategies against these tumors could be useful to both species. In humans, chondroitin sulphate proteoglycan 4 (CSPG4) is a marker involved in tumor progression and could be a candidate target for immunotherapy. The anti-CSPG4 DNA electrovaccination has shown to be an effective approach for canine malignant melanoma (CMM) [1]. An immunohistochemistry evaluation of CSPG4 expression in tumour tissue is generally performed prior to electrovaccination. To assess the possibility to perform a rapid molecular evaluation and in order to validate these spontaneous canine tumors as the model for human studies, we investigate the CSPG4 gene expression by RT qPCR in CMM, OSA, and canine mammary tumors (CMT). The total RNA was extracted from RNAlater stored tissue samples (CMM n=16; OSA n=13; CMT n=6; five paired normal tissues for CMM, five paired normal tissues for OSA and one paired normal tissue for CMT), retro-transcribed and then analyzed by duplex RT-qPCR using two different TaqMan assays for the target gene CSPG4 and the internal reference gene (RG) Ribosomal Protein S19 (RPS19). RPS19 was selected from a panel of 9 candidate RGs, according to NormFinder analysis following the protocol already described [2]. Relative expression was analyzed by CFX Maestro™ Software. Student t-test and ANOVA were performed (significance set at P<0.05). Results showed that gene expression of CSPG4 in OSA tissues is significantly increased by 3-4 folds when compared to controls. In CMT, gene expression of the target was increased from 1.5 to 19.9 folds. In melanoma, although an increasing trend was observed, no significant differences between the two groups were highlighted. Immunohistochemistry analysis of the two cancer types showed that the expression of CSPG4 within CMM is concentrated in isles of cells compared to OSA, where the distribution of positive cells is homogeneous. This evidence could explain the differences in gene expression results.CSPG4 immunohistochemistry evaluation in mammary carcinoma is in progress. The evidence of CSPG4 expression in a different type of canine tumors opens the way to the possibility of extending the CSPG4 immunotherapy marker in CMM, OSA, and CMT and may have an impact to translate this strategy modality to human oncology.

Keywords: canine melanoma, canine mammary carcinomas, canine osteosarcoma, CSPG4, gene expression, immunotherapy

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Authors: Davide Cocchi, Andrea Zucchelli, Luca Raimondi, Maria Brugo Tommaso

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The increasing attention is given to the issues of environmental pollution and climate change is exponentially stimulating the development of electrically propelled vehicles powered by renewable energy, in particular, the solar one. Given the small amount of solar energy that can be stored and subsequently transformed into propulsive energy, it is necessary to develop vehicles with high mechanical, electrical and aerodynamic efficiencies along with reduced masses. The reduction of the masses is of fundamental relevance especially for the unsprung masses, that is the assembly of those elements that do not undergo a variation of their distance from the ground (wheel, suspension system, hub, upright, braking system). Therefore, the reduction of unsprung masses is fundamental in decreasing the rolling inertia and improving the drivability, comfort, and performance of the vehicle. This principle applies even more in solar propelled vehicles, equipped with an electric motor that is connected directly to the wheel hub. In this solution, the electric motor is integrated inside the wheel. Since the electric motor is part of the unsprung masses, the development of compact and lightweight solutions is of fundamental importance. The purpose of this research is the design development and optimization of a CFRP 16 wheel hub motor for solar propulsion vehicles that can carry up to four people. In addition to trying to maximize aspects of primary importance such as mass, strength, and stiffness, other innovative constructive aspects were explored. One of the main objectives has been to achieve a high geometric packing in order to ensure a reduced lateral dimension, without reducing the power exerted by the electric motor. In the final solution, it was possible to realize a wheel hub motor assembly completely comprised inside the rim width, for a total lateral overall dimension of less than 100 mm. This result was achieved by developing an innovative connection system between the wheel and the rotor with a double purpose: centering and transmission of the driving torque. This solution with appropriate interlocking noses allows the transfer of high torques and at the same time guarantees both the centering and the necessary stiffness of the transmission system. Moreover, to avoid delamination in critical areas, evaluated by means of FEM analysis using 3D Hashin damage criteria, electrospun nanofibrous mats have been interleaved between CFRP critical layers. In order to reduce rolling resistance, the rim has been designed to withstand high inflation pressure. Laboratory tests have been performed on the rim using the Digital Image Correlation technique (DIC). The wheel has been tested for fatigue bending according to E/ECE/324 R124e.

Keywords: composite laminate, delamination, DIC, lightweight vehicle, motor hub wheel, nanofiber

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Authors: Fruzsina Luca Kézér, Viktor Jurkovich, Ottó Szenci, János Tőzsér, Levente Kovács

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Positive interaction between people and animals could have a favorable effect on the welfare and production by reducing stress levels. However, to the repeated contact with humans (e.g. farm staff, veterinarians or herdsmen), animals may respond with escape behavior or avoidance, which both have negative effects on the ease of handling, welfare and may lead to the expression of aggressive behaviors. Rough or aversive handling can impair health and the function of the cardiac autonomic activity due to fear and stress, which also can be determined by certain parameters of heart rate variability (HRV). Although the essential relationships between fear from humans and basal tone of the autonomic nervous system were described by the authors previously, several questions remained unclear in terms of the associations between different coping strategies (behavioral syndromes) of the animals and physiological responsiveness to humans. The main goal of this study was to find out whether human behavior and emotions to the animals have an impact on cardiac function and behavior of animals with different coping styles in response situations. Therefore, in the present study, special (fear, approaching, restraint, novel arena, novel object) tests were performed on healthy, 2-year old heifers (n = 104) differing in coping styles [reactive (passive) vs. proactive (active) coping]. Animals were categorized as reactive or proactive based on the following tests: 1) aggressive behavior at the feeding bunk, 2) avoidance from an approaching person, 3) immobility, and 4) daily activity (number of posture changes). Heart rate, the high frequency (HF) component of HRV as a measure of vagal activity and the ratio between the low frequency (LF) and HF components (LF/HF ratio) as a parameter of sympathetic nervous system activity were calculated for all individual during lying posture (baseline) and for response situations in novel object, novel arena, and unfamiliar person tests (both for 5 min), respectively. The differences between baseline and response were compared between groups. Higher sympathetic (higher heart rates and LF/HF ratios) and lower parasympathetic activity (lower HF) was found for proactive animals in response situations than for reactive (passive) animals either during the novel object, the novel arena and the unfamiliar person test. It suggests that animals with different behavioral traits differ in their immediate autonomic adaptation to novelty and people. Based on our preliminary results, it seems, that the analysis of HRV can help to understand the physiological manifestation of responsiveness to novelty and human presence in dairy cattle with different behavioral syndromes.

Keywords: behavioral syndromes, human-cattle interaction, novel arena test, physiological responsiveness, proactive coping, reactive coping

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Authors: Z. Yousefniayejahr, S. Bolognini, A. Bonini, C. Campobasso, N. Poma, F. Vivaldi, M. Di Luca, A. Tavanti, F. Di Francesco

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Pathogenic bacteria represent a threat to healthcare systems and the food industry because their rapid detection remains challenging. Electrochemical biosensors are gaining prominence as a novel technology for the detection of pathogens due to intrinsic features such as low cost, rapid response time, and portability, which make them a valuable alternative to traditional methodologies. These sensors use biorecognition elements that are crucial for the identification of specific bacteria. In this context, bacteriophages are promising tools for their inherent high selectivity towards bacterial hosts, which is of fundamental importance when detecting bacterial pathogens in complex biological samples. In this study, we present the development of a low-cost and portable sensor based on the Zeno phage for the rapid detection of Staphylococcus aureus. Screen-printed gold electrodes functionalized with the Zeno phage were used, and electrochemical impedance spectroscopy was applied to evaluate the change of the charge transfer resistance (Rct) as a result of the interaction with S. aureus MRSA ATCC 43300. The phage-based biosensor showed a linear range from 101 to 104 CFU/mL with a 20-minute response time and a limit of detection (LOD) of 1.2 CFU/mL under physiological conditions. The biosensor’s ability to recognize various strains of staphylococci was also successfully demonstrated in the presence of clinical isolates collected from different geographic areas. Assays using S. epidermidis were also carried out to verify the species-specificity of the phage sensor. We only observed a remarkable change of the Rct in the presence of the target S. aureus bacteria, while no substantial binding to S. epidermidis occurred. This confirmed that the Zeno phage sensor only targets S. aureus species within the genus Staphylococcus. In addition, the biosensor's specificity with respect to other bacterial species, including gram-positive bacteria like Enterococcus faecium and the gram-negative bacterium Pseudomonas aeruginosa, was evaluated, and a non-significant impedimetric signal was observed. Notably, the biosensor successfully identified S. aureus bacterial cells in a complex matrix such as a nasal swab, opening the possibility of its use in a real-case scenario. We diluted different concentrations of S. aureus from 108 to 100 CFU/mL with a ratio of 1:10 in the nasal swap matrices collected from healthy donors. Three different sensors were applied to measure various concentrations of bacteria. Our sensor indicated high selectivity to detect S. aureus in biological matrices compared to time-consuming traditional methods, such as enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR), and radioimmunoassay (RIA), etc. With the aim to study the possibility to use this biosensor to address the challenge associated to pathogen detection, ongoing research is focused on the assessment of the biosensor’s analytical performances in different biological samples and the discovery of new phage bioreceptors.

Keywords: electrochemical impedance spectroscopy, bacteriophage, biosensor, Staphylococcus aureus

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Authors: Giulia Fredi, Andrea Dorigato, Luca Fambri, Alessandro Pegoretti

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Thermal energy storage (TES) is the storage of heat for later use, thus filling the gap between energy request and supply. The most widely used materials for TES are the organic solid-liquid phase change materials (PCMs), such as paraffin. These materials store/release a high amount of latent heat thanks to their high specific melting enthalpy, operate in a narrow temperature range and have a tunable working temperature. However, they suffer from a low thermal conductivity and need to be confined to prevent leakage. These two issues can be tackled by confining PCMs with carbon nanotubes (CNTs). TES applications include the buildings industry, solar thermal energy collection and thermal management of electronics. In most cases, TES systems are an additional component to be added to the main structure, but if weight and volume savings are key issues, it would be advantageous to embed the TES functionality directly in the structure. Such multifunctional materials could be employed in the automotive industry, where the diffusion of lightweight structures could complicate the thermal management of the cockpit environment or of other temperature sensitive components. This work aims to produce epoxy/carbon structural laminates containing CNT-stabilized paraffin. CNTs were added to molten paraffin in a fraction of 10 wt%, as this was the minimum amount at which no leakage was detected above the melting temperature (45°C). The paraffin/CNT blend was cryogenically milled to obtain particles with an average size of 50 µm. They were added in various percentages (20, 30 and 40 wt%) to an epoxy/hardener formulation, which was used as a matrix to produce laminates through a wet layup technique, by stacking five plies of a plain carbon fiber fabric. The samples were characterized microstructurally, thermally and mechanically. Differential scanning calorimetry (DSC) tests showed that the paraffin kept its ability to melt and crystallize also in the laminates, and the melting enthalpy was almost proportional to the paraffin weight fraction. These thermal properties were retained after fifty heating/cooling cycles. Laser flash analysis showed that the thermal conductivity through the thickness increased with an increase of the PCM, due to the presence of CNTs. The ability of the developed laminates to contribute to the thermal management was also assessed by monitoring their cooling rates through a thermal camera. Three-point bending tests showed that the flexural modulus was only slightly impaired by the presence of the paraffin/CNT particles, while a more sensible decrease of the stress and strain at break and the interlaminar shear strength was detected. Optical and scanning electron microscope images revealed that these could be attributed to the preferential location of the PCM in the interlaminar region. These results demonstrated the feasibility of multifunctional structural TES composites and highlighted that the PCM size and distribution affect the mechanical properties. In this perspective, this group is working on the encapsulation of paraffin in a sol-gel derived organosilica shell. Submicron spheres have been produced, and the current activity focuses on the optimization of the synthesis parameters to increase the emulsion efficiency.

Keywords: carbon fibers, carbon nanotubes, lightweight materials, multifunctional composites, thermal energy storage

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Authors: Maddalena Arigoni, Maria Luisa Ratto, Raffaele A. Calogero, Luca Alessandri

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The presence of tumor heterogeneity, where distinct cancer cells exhibit diverse morphological and phenotypic profiles, including gene expression, metabolism, and proliferation, poses challenges for molecular prognostic markers and patient classification for targeted therapies. Understanding the causes and progression of cancer requires research efforts aimed at characterizing heterogeneity, which can be facilitated by evolving single-cell sequencing technologies. However, analyzing single-cell data necessitates computational methods that often lack objective validation. Therefore, the establishment of benchmarking datasets is necessary to provide a controlled environment for validating bioinformatics tools in the field of single-cell oncology. Benchmarking bioinformatics tools for single-cell experiments can be costly due to the high expense involved. Therefore, datasets used for benchmarking are typically sourced from publicly available experiments, which often lack a comprehensive cell annotation. This limitation can affect the accuracy and effectiveness of such experiments as benchmarking tools. To address this issue, we introduce omics benchmark experiments designed to evaluate bioinformatics tools to depict the heterogeneity in single-cell tumor experiments. We conducted single-cell RNA sequencing on six lung cancer tumor cell lines that display resistant clones upon treatment of EGFR mutated tumors and are characterized by driver genes, namely ROS1, ALK, HER2, MET, KRAS, and BRAF. These driver genes are associated with downstream networks controlled by EGFR mutations, such as JAK-STAT, PI3K-AKT-mTOR, and MEK-ERK. The experiment also featured an EGFR-mutated cell line. Using 10XGenomics platform with cellplex technology, we analyzed the seven cell lines together with a pseudo-immunological microenvironment consisting of PBMC cells labeled with the Biolegend TotalSeq™-B Human Universal Cocktail (CITEseq). This technology allowed for independent labeling of each cell line and single-cell analysis of the pooled seven cell lines and the pseudo-microenvironment. The data generated from the aforementioned experiments are available as part of an online tool, which allows users to define cell heterogeneity and generates count tables as an output. The tool provides the cell line derivation for each cell and cell annotations for the pseudo-microenvironment based on CITEseq data by an experienced immunologist. Additionally, we created a range of pseudo-tumor tissues using different ratios of the aforementioned cells embedded in matrigel. These tissues were analyzed using 10XGenomics (FFPE samples) and Curio Bioscience (fresh frozen samples) platforms for spatial transcriptomics, further expanding the scope of our benchmark experiments. The benchmark experiments we conducted provide a unique opportunity to evaluate the performance of bioinformatics tools for detecting and characterizing tumor heterogeneity at the single-cell level. Overall, our experiments provide a controlled and standardized environment for assessing the accuracy and robustness of bioinformatics tools for studying tumor heterogeneity at the single-cell level, which can ultimately lead to more precise and effective cancer diagnosis and treatment.

Keywords: single cell omics, benchmark, spatial transcriptomics, CITEseq

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