40 results on '"Bossi AM"'
Search Results
2. An Efficient Bio-Receptor Layer Combined with a Plasmonic Plastic Optical Fiber Probe for Cortisol Detection in Saliva.
- Author
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Arcadio F, Seggio M, Pitruzzella R, Zeni L, Bossi AM, and Cennamo N
- Subjects
- Humans, Limit of Detection, Plastics, Receptors, Glucocorticoid, Hydrocortisone analysis, Saliva chemistry, Surface Plasmon Resonance, Optical Fibers, Biosensing Techniques
- Abstract
Cortisol is a clinically validated stress biomarker that takes part in many physiological and psychological functions related to the body's response to stress factors. In particular, it has emerged as a pivotal tool for understanding stress levels and overall well-being. Usually, in clinics, cortisol levels are monitored in blood or urine, but significant changes are also registered in sweat and saliva. In this work, a surface plasmon resonance probe based on a D-shaped plastic optical fiber was functionalized with a glucocorticoid receptor exploited as a highly efficient bioreceptor specific to cortisol. The developed plastic optical fiber biosensor was tested for cortisol detection in buffer and artificial saliva. The biosensor response showed very good selectivity towards other hormones and a detection limit of about 59 fM and 96 fM in phosphate saline buffer and artificial saliva, respectively. The obtained detection limit, with a rapid detection time (about 5 min) and a low-cost sensor system, paved the way for determining the cortisol concentration in saliva samples without any extraction process or sample pretreatment via a point-of-care test.
- Published
- 2024
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3. Toward Nano- and Microplastic Sensors: Identification of Nano- and Microplastic Particles via Artificial Intelligence Combined with a Plasmonic Probe Functionalized with an Estrogen Receptor.
- Author
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Seggio M, Arcadio F, Radicchi E, Cennamo N, Zeni L, and Bossi AM
- Abstract
Nano- and microplastic particles are a global and emerging environmental issue that might pose potential threats to human health. The present work exploits artificial intelligence (AI) to identify nano- and microplastics in water by monitoring the interaction of the sample with a sensitive surface. An estrogen receptor (ER) grafted onto a gold surface, realized on a nonexpensive and easy-to-produce plastic optical fiber (POF) platform in order to excite a surface plasmon resonance (SPR) phenomenon, has been developed in order to carry out a "smart" sensitive interface (ER-SPR-POF interface). The ER-SPR-POF interface offers output data useful for exploiting a machine learning-based approach to achieve nano- and microplastic particle sensors. This work developed a proof-of-concept sensor through a training phase carried out by different particles, in terms of materials and size. The experimental results have demonstrated that the proposed "smart" ER-SPR-POF interface combined with AI can be used to identify the kind of particles in terms of the materials (polystyrene; poly(methyl methacrylate)) and size (20 μm; 100 nm) with an accuracy of 90.3%., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)
- Published
- 2024
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4. A plasmonic gold nano-surface functionalized with the estrogen receptor for fast and highly sensitive detection of nanoplastics.
- Author
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Seggio M, Arcadio F, Cennamo N, Zeni L, and Bossi AM
- Subjects
- Humans, Surface Plasmon Resonance, Receptors, Estrogen, Microplastics, Gold, Biosensing Techniques
- Abstract
Nanoplastics are a global emerging environmental problem whose effects might pose potential threats to the human's health. Despite the relevance of the issue, fast, reliable and quantitative in situ analytical approaches to determine nanoplastics are not yet available. The aim of this work was to devise an optical sensor with the goal of direct detecting and quantifying nanoplastics in seawater without sample pre-treatments. To this purpose, a nano-plasmonic biosensor was developed by exploiting an Estrogen Receptor (ER) recognition element grafted onto a polymer-based gold nanograting (GNG) plasmonic platform. The ER-GNG biosensor required just minute sample volumes (2 μL), allowed rapid detection (3 min) and enabled to determine nanoplastics in simulated seawater with a linear dynamic concentrations range of 1-100 ng/mL, thus encompassing the expected environmental loads. The nanostructured grating (GNG) provided remarkable performance enhancements, extending the measurement range across five orders of magnitude, thanks to the both the SPR and the localized SPR phenomena occurring at the GNG chip. At last, the ER-GNG biosensor was tested on real seawater samples collected in the Naples area and the results (∼30 ng/mL) were verified by a conventional approach (filtration and evaporation), confirming the ER-GNG sensor offers a straightforward and highly sensitive method for the direct in-field nanoplastics monitoring., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)
- Published
- 2024
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5. Sensing Approaches Exploiting Molecularly Imprinted Nanoparticles and Lossy Mode Resonance in Polymer Optical Fibers.
- Author
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Arcadio F, Noël L, Del Prete D, Seggio M, Zeni L, Bossi AM, Soppera O, and Cennamo N
- Abstract
In this work, two different lossy mode resonance (LMR) platforms based on plastic optical fibers (POFs) are developed and tested in a biochemical sensing scenario. The LMR platforms are based on the combination of two metal oxides (MOs), i.e., zirconium oxide (ZrO
2 ) and titanium oxide (TiO2 ), and deposited on the exposed core of D-shaped POF chips. More specifically, two experimental sensor configurations were obtained by swapping the mutual position of the Mos films over to the core of the D-shaped POF probe. The POF-LMR sensors were first characterized as refractometers, proving the bulk sensitivities. Then, both the POF-LMR platforms were functionalized using molecularly imprinted nanoparticles (nanoMIPs) specific for human transferrin (HTR) in order to carry out binding tests. The achieved results report a bulk sensitivity equal to about 148 nm/RIU in the best sensor configuration, namely the POF-TiO2 -ZrO2 . In contrast, both optical configurations combined with nanoMIPs showed an ultra-low detection limit (fM), demonstrating excellent efficiency of the used receptor (nanoMIPs) and paving the way to disposable POF-LMR biochemical sensors that are easy-to-use, low-cost, and highly sensitive.- Published
- 2023
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6. Time-Resolved Fluorescence Spectroscopy of Molecularly Imprinted Nanoprobes as an Ultralow Detection Nanosensing Tool for Protein Contaminants.
- Author
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Bossi AM, Marinangeli A, Quaranta A, Pancheri L, and Maniglio D
- Subjects
- Spectrometry, Fluorescence, Limit of Detection, Albumins, Molecular Imprinting methods, Nanoparticles chemistry
- Abstract
Currently, optical sensors based on molecularly imprinted polymers (MIPs) have been attracting significant interest. MIP sensing relies on the combination of the MIP's selective capability, which is conveyed to the polymeric material by a template-assisted synthesis, with optical techniques that offer exquisite sensitivity. In this work, we devised an MIP nanoparticle optical sensor for the ultralow detection of serum albumin through time-resolved fluorescence spectroscopy. The Fluo-nanoMIPs (∅~120 nm) were synthetized using fluorescein-O-methacrylate (0.1×, 1×, 10× mol:mol versus template) as an organic fluorescent reporter. The ability of 0.1× and 1×Fluo-nanoMIPs to bind albumin (15 fM-150 nM) was confirmed by fluorescence intensity analyses and isothermal titration calorimetry. The apparent dissociation constant (K
app ) was 30 pM. Conversely, the 10× fluorophore content did not enable monitoring binding. Then, the time-resolved fluorescence spectroscopy of the nanosensors was studied. The 1×Fluo-nanoMIPs showed a decrease in fluorescence lifetime upon binding to albumin (100 fM-150 nM), Kapp = 28 pM, linear dynamic range 3.0-83.5 pM, limit of detection (LOD) 1.26 pM. Selectivity was confirmed testing 1×Fluo-nanoMIPs against competitor proteins. Finally, as a proof of concept, the nanosensors demonstrated detection of the albumin (1.5 nM) spiked in wine samples, suggesting a possible scaling up of the method in monitoring allergens in wines.- Published
- 2023
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7. Soft molecularly imprinted nanoparticles with simultaneous lossy mode and surface plasmon multi-resonances for femtomolar sensing of serum transferrin protein.
- Author
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Arcadio F, Noël L, Del Prete D, Maniglio D, Seggio M, Soppera O, Cennamo N, Bossi AM, and Zeni L
- Subjects
- Humans, Surface Plasmon Resonance, Blood Proteins, Transferrin, Nanoparticles
- Abstract
The simultaneous interrogation of both lossy mode (LMR) and surface plasmon (SPR) resonances was herein exploited for the first time to devise a sensor in combination with soft molecularly imprinting of nanoparticles (nanoMIPs), specifically entailed of the selectivity towards the protein biomarker human serum transferrin (HTR). Two distinct metal-oxide bilayers, i.e. TiO
2 -ZrO2 and ZrO2 -TiO2 , were used in the SPR-LMR sensing platforms. The responses to binding of the target protein HTR of both sensing configurations (TiO2 -ZrO2 -Au-nanoMIPs, ZrO2 -TiO2 -Au-nanoMIPs) showed femtomolar HTR detection, LODs of tens of fM and KDapp ~ 30 fM. Selectivity for HTR was demonstrated. The SPR interrogation was more efficient for the ZrO2 -TiO2 -Au-nanoMIPs configuration (sensitivity at low concentrations, S = 0.108 nm/fM) than for the TiO2 -ZrO2 -Au-nanoMIPs one (S = 0.061 nm/fM); while LMR was more efficient for TiO2 -ZrO2 -Au-nanoMIPs (S = 0.396 nm/fM) than for ZrO2 -TiO2 -Au-nanoMIPs (S = 0.177 nm/fM). The simultaneous resonance monitoring is advantageous for point of care determinations, both in terms of measurement's redundancy, that enables the cross-control of the measure and the optimization of the detection, by exploiting the individual characteristics of each resonance., (© 2023. The Author(s).)- Published
- 2023
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8. Estradiol Detection for Aquaculture Exploiting Plasmonic Spoon-Shaped Biosensors.
- Author
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Arcadio F, Seggio M, Zeni L, Bossi AM, and Cennamo N
- Subjects
- Estrogen Receptor alpha, Receptors, Estrogen, Surface Plasmon Resonance methods, Estradiol, Biosensing Techniques methods
- Abstract
In this work, a surface plasmon resonance (SPR) biosensor based on a spoon-shaped waveguide combined with an estrogen receptor (ERα) was developed and characterized for the detection and the quantification of estradiol in real water samples. The fabrication process for realizing the SPR platform required a single step consisting of metal deposition on the surface of a polystyrene spoon-shaped waveguide featuring a built-in measuring cell. The biosensor was achieved by functionalizing the bowl sensitive surface with a specific estrogen receptor (ERα) that was able to bind the estradiol. In a first phase, the biosensor tests were performed in a phosphate buffer solution obtaining a limit of detection (LOD) equal to 0.1 pM. Then, in order to evaluate the biosensor's response in different real matrices related to aquaculture, its performances were examined in seawater and freshwater. The experimental results support the possibility of using the ERα-based biosensor for the screening of estradiol in both matrices.
- Published
- 2023
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9. Porous Thermoplastic Molded Regenerated Silk Crosslinked by the Addition of Citric Acid.
- Author
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Bucciarelli A, Vighi N, Bossi AM, Grigolo B, and Maniglio D
- Abstract
Thermoplastic molded regenerated silk fibroin was proposed as a structural material in tissue engineering applications, mainly for application in bone. The protocol allows us to obtain a compact non-porous material with a compression modulus in the order of a Giga Pascal in dry conditions (and in the order of tens of MPa in wet conditions). This material is produced by compressing a lyophilized silk fibroin powder or sponge into a mold temperature higher than the glass transition temperature. The main purpose of the produced resin was the osteofixation and other structural applications in which the lack of porosity was not an issue. In this work, we introduced the use of citric acid in the thermoplastic molding protocol of silk fibroin to obtain porosity inside the structural material. The citric acid powder during the compression acted as a template for the pore formation. The mean pore diameter achieved by the addition of the higher amount of citric acid was around 5 μm. In addition, citric acid could effectively crosslink the silk fibroin chain, improving its mechanical strength. This effect was proved both by evaluating the compression modulus (the highest value recorded was 77 MPa in wet conditions) and by studying the spectra obtained by Fourier transform infrared spectroscopy. This protocol may be applied in the near future to the production of structural bone scaffolds.
- Published
- 2023
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10. Non-Specific Responsive Nanogels and Plasmonics to Design MathMaterial Sensing Interfaces: The Case of a Solvent Sensor.
- Author
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Cennamo N, Arcadio F, Capasso F, Maniglio D, Zeni L, and Bossi AM
- Subjects
- Nanogels, Solvents, Plastics, Polyethylene Glycols, Polyethyleneimine
- Abstract
The combination of non-specific deformable nanogels and plasmonic optical probes provides an innovative solution for specific sensing using a generalistic recognition layer. Soft polyacrylamide nanogels that lack specific selectivity but are characterized by responsive behavior, i.e., shrinking and swelling dependent on the surrounding environment, were grafted to a gold plasmonic D-shaped plastic optical fiber (POF) probe. The nanogel-POF cyclically challenged with water or alcoholic solutions optically reported the reversible solvent-to-phase transitions of the nanomaterial, embodying a primary optical switch. Additionally, the non-specific nanogel-POF interface exhibited more degrees of freedom through which specific sensing was enabled. The real-time monitoring of the refractive index variations due to the time-related volume-to-phase transition effects of the nanogels enabled us to determine the environment's characteristics and broadly classify solvents. Hence the nanogel-POF interface was a descriptor of mathematical functions for substance identification and classification processes. These results epitomize the concept of responsive non-specific nanomaterials to perform a multiparametric description of the environment, offering a specific set of features for the processing stage and particularly suitable for machine and deep learning. Thus, soft MathMaterial interfaces provide the ground to devise devices suitable for the next generation of smart intelligent sensing processes.
- Published
- 2022
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11. Methacrylated Silk Fibroin Additive Manufacturing of Shape Memory Constructs with Possible Application in Bone Regeneration.
- Author
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Bucciarelli A, Petretta M, Grigolo B, Gambari L, Bossi AM, Grassi F, and Maniglio D
- Abstract
Methacrylated silk (Sil-MA) is a chemically modified silk fibroin specifically designed to be crosslinkable under UV light, which makes this material applicable in additive manufacturing techniques and allows the prototyping and development of patient-specific 2D or 3D constructs. In this study, we produced a thin grid structure based on crosslinked Sil-MA that can be withdrawn and ejected and that can recover its shape after rehydration. A complete chemical and physical characterization of Sil-MA was first conducted. Additionally, we tested Sil-MA biocompatibility according to the International Standard Organization protocols (ISO 10993) ensuring the possibility of using it in future trials. Sil-MA was also tested to verify its ability to support osteogenesis. Overall, Sil-MA was shown to be biocompatible and osteoconductive. Finally, two different additive manufacturing technologies, a Digital Light Processing (DLP) UV projector and a pneumatic extrusion technique, were used to develop a Sil-MA grid construct. A proof-of-concept of its shape-memory property was provided. Together, our data support the hypothesis that Sil-MA grid constructs can be injectable and applicable in bone regeneration applications.
- Published
- 2022
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12. Spoon-shaped polymer waveguides to excite multiple plasmonic phenomena: A multisensor based on antibody and molecularly imprinted nanoparticles to detect albumin concentrations over eight orders of magnitude.
- Author
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Cennamo N, Arcadio F, Seggio M, Maniglio D, Zeni L, and Bossi AM
- Subjects
- Antibodies, Gold chemistry, Humans, Polymers chemistry, Serum Albumin, Human, Surface Plasmon Resonance methods, Biosensing Techniques methods, Molecular Imprinting methods, Nanoparticles chemistry
- Abstract
A polymeric multimode waveguide, characterized by a pioneering spoon-shaped geometry, was herein proposed for the first time to devise Surface Plasmon Resonance (SPR) biochemical sensors. The plasmon excitation was enabled by layering a gold nanofilm of ∼60 nm onto the spoon-waveguide. As a consequence of the waveguide's extra-ordinary geometry, two distinct sensing regions were identified: a planar one, located on the spoon's neck, and a concave one on the bowl, with angled surfaces. The bulk sensitivity (S
n ) is correlated both to the way the light was launched in/collected from the sensor (parallel or orthogonal to the main axis of the waveguide) and to the sensing area interrogated (planar-neck or angled-bowl), indicating that the sensor's performance can be conveniently tuned, depending on the chosen measuring configuration. The SPR sensor's characterization showed Sn equal to 750 nm/RIU for the neck and to 950 nm/RIU for the bowl. To further inspect the peculiar sensing-features and assess the application niches, the spoon-shaped waveguide was functionalized with two kinds of receptors, both specific for human serum albumin (HSA): an antibody on the bowl region (high Sn ); molecularly imprinted nanoparticles (nanoMIPs) on the neck region (low Sn ). The experimental results showed a limit of detection (LOD) for the immune-sensor of 280 pM and an LOD for the nanoMIP-sensor of 4.16 fM. The overall response of the HSA multi-sensor encompassed eight orders of magnitude, suggesting that the spoon-shaped waveguide's provides multi-scale detection and holds potential to devise multi-analyte sensing platforms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)- Published
- 2022
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13. A Plasmonic Biosensor Based on Light-Diffusing Fibers Functionalized with Molecularly Imprinted Nanoparticles for Ultralow Sensing of Proteins.
- Author
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Arcadio F, Seggio M, Del Prete D, Buonanno G, Mendes J, Coelho LCC, Jorge PAS, Zeni L, Bossi AM, and Cennamo N
- Abstract
Plasmonic bio/chemical sensing based on optical fibers combined with molecularly imprinted nanoparticles (nanoMIPs), which are polymeric receptors prepared by a template-assisted synthesis, has been demonstrated as a powerful method to attain ultra-low detection limits, particularly when exploiting soft nanoMIPs, which are known to deform upon analyte binding. This work presents the development of a surface plasmon resonance (SPR) sensor in silica light-diffusing fibers (LDFs) functionalized with a specific nanoMIP receptor, entailed for the recognition of the protein human serum transferrin (HTR). Despite their great versatility, to date only SPR-LFDs functionalized with antibodies have been reported. Here, the innovative combination of an SPR-LFD platform and nanoMIPs led to the development of a sensor with an ultra-low limit of detection (LOD), equal to about 4 fM, and selective for its target analyte HTR. It is worth noting that the SPR-LDF-nanoMIP sensor was mounted within a specially designed 3D-printed holder yielding a measurement cell suitable for a rapid and reliable setup, and easy for the scaling up of the measurements. Moreover, the fabrication process to realize the SPR platform is minimal, requiring only a metal deposition step.
- Published
- 2022
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14. BioMIPs: molecularly imprinted silk fibroin nanoparticles to recognize the iron regulating hormone hepcidin.
- Author
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Bossi AM and Maniglio D
- Subjects
- Fibroins chemistry, Hepcidins chemistry, Molecular Imprinting, Nanoparticles chemistry
- Abstract
The possibility to prepare molecularly imprinted nanoparticles from silk fibroin was recently demonstrated starting from methacrylated silk fibroin and choosing a protein as template. Here, we attempted the imprinting of fibroin-based molecularly imprinted polymers (MIPs), called bioMIPs, using as a template hepcidin that is a iron-metabolism regulator-peptide, possessing a hairpin structure. A homogeneous population (PDI < 0.2) of bioMIPs with size ~50 nm was produced. The bioMIPs were selective for the template; the estimated dissociation constant for hepcidin was K
D = 3.6 ± 0.5 10-7 M and the average number of binding sites per bioMIP was equal to 2. The bioMIPs used in a competitive assay for hepcidin in serum showed a detection range of 1.01 10-7 - 6.82 10-7 M and a limit of detection of 3.29 10-8 M., (© 2022. The Author(s).)- Published
- 2022
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15. On the Effect of Soft Molecularly Imprinted Nanoparticles Receptors Combined to Nanoplasmonic Probes for Biomedical Applications.
- Author
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Cennamo N, Bossi AM, Arcadio F, Maniglio D, and Zeni L
- Abstract
Soft, deformable, molecularly imprinted nanoparticles (nanoMIPs) were combined to nano-plasmonic sensor chips realized on poly (methyl methacrylate) (PMMA) substrates to develop highly sensitive bio/chemical sensors. NanoMIPs (d
mean < 50 nm), which are tailor-made nanoreceptors prepared by a template assisted synthesis, were made selective to bind Bovine Serum Albumin (BSA), and were herein used to functionalize gold optical nanostructures placed on a PMMA substrate, this latter acting as a slab waveguide. We compared nanoMIP-functionalized non-optimized gold nanogratings based on periodic nano-stripes to optimized nanogratings with a deposited ultra-thin MIP layer (<100 nm). The sensors performances were tested by the detection of BSA using the same setup, in which both chips were considered as slab waveguides, with the periodic nano-stripes allocated in a longitudinal orientation with respect to the direction of the input light. Result demonstrated the nanoMIP-non optimized nanogratings showed superior performance with respect to the ultra-thin MIP-optimized nanogratings. The peculiar deformable character of the nano-MIPs enabled to significantly enhance the limit of detection (LOD) of the plasmonic bio/sensor, allowing the detection of the low femtomolar concentration of analyte (LOD ∼ 3 fM), thus outpassing of four orders of magnitude the sensitivies achieved so far on optimized nano-patterned plasmonic platforms functionalized with ultra-thin MIP layers. Thus, deformable nanoMIPs onto non-optimized plasmonic probes permit to attain ultralow detections, down to the quasi-single molecule. As a general consideration, the combination of more plasmonic transducers to different kinds of MIP receptors is discussed as a mean to attain the detection range for the selected application field., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Cennamo, Bossi, Arcadio, Maniglio and Zeni.)- Published
- 2021
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16. Molecularly imprinted polymers by epitope imprinting: a journey from molecular interactions to the available bioinformatics resources to scout for epitope templates.
- Author
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Pasquardini L and Bossi AM
- Subjects
- Polymers chemistry, Protein Conformation, Computational Biology methods, Epitopes chemistry, Molecularly Imprinted Polymers chemistry
- Abstract
The molecular imprinting of proteins is the process of forming biomimetics with entailed protein-recognition by means of a template-assisted synthesis. Protein-imprinted polymers (pMIPs) have been successfully employed in separations, assays, sensors, and imaging. From a technical point of view, imprinting a protein is both costly, for protein expression and purification, and challenging, for the preservation of the protein's structural properties. In fact, the imprinting process needs to guarantee the preservation of the same protein three-dimensional conformation that later would be recognized. So far, the captivating idea to imprint just a portion of the protein, i.e., an epitope, instead of the whole, proved successful, offering reduced costs, compatibility with many synthetic conditions (solvents, pH, temperatures), and fine-tuning of the peptide sequence so to target specific physiological and functional conditions of the protein, such as post-translational modifications. Here, protein-protein interactions and the biochemical features of the epitopes are inspected, deriving lessons to prepare more effective pMIPs. Epitopes are categorized in linear or structured, immunogenic or not, located at the protein's surface or buried in its core and the imprinting strategies are discussed. Moreover, attention is given to freely available online bioinformatics resources that might offer key tools to gain further rationale amid the selection process of suitable epitopes templates., (© 2021. The Author(s).)
- Published
- 2021
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17. Molecularly Imprinted Silk Fibroin Nanoparticles.
- Author
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Bossi AM, Bucciarelli A, and Maniglio D
- Subjects
- Materials Testing, Particle Size, Fibroins chemical synthesis, Fibroins chemistry, Molecular Imprinting, Nanoparticles chemistry
- Abstract
Nanosized biomimetics prepared by the strategy of molecular imprinting, that is, the stamping of recognition sites by means of a template-assisted synthesis, are demonstrating potential as plastic antibodies in medicine, proving effective for cell imaging and targeted therapies. Most molecularly imprinted nanoparticles (MIP-NPs) are currently made of soft matter, such as polyacrylamide and derivatives. Yet, MIP-NPs biocompatibility is crucial for their effective translation into clinical uses. Here, we propose the original idea to synthesize fully biocompatible molecularly imprinted nanoparticles starting from the natural polymer silk fibroin (MIP SF-NPs), which is nontoxic and highly biocompatible. The conditions to produce MIP SF-NPs of different sizes ( d
mean ∼ 50 nm; dmean ∼ 100 nm) were set using the response surface method. The stamping of a single, high affinity ( KD = 57 × 10-9 M), and selective recognition site per silk fibroin nanoparticle was demonstrated, together with the confirmation of nontoxicity. Additionally, MIP SF-NPs were used to decorate silk microfibers and silk nanofibers, providing a general means to add entailed biofunctionalities to materials.- Published
- 2021
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18. A Surface Plasmon Resonance Plastic Optical Fiber Biosensor for the Detection of Pancreatic Amylase in Surgically-Placed Drain Effluent.
- Author
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Pasquardini L, Cennamo N, Malleo G, Vanzetti L, Zeni L, Bonamini D, Salvia R, Bassi C, and Bossi AM
- Subjects
- Drainage, Humans, Optical Fibers, Pancreatectomy, Plastics, Amylases analysis, Biosensing Techniques, Pancreas enzymology, Pancreatic Fistula diagnosis, Surface Plasmon Resonance
- Abstract
Postoperative pancreatic fistula (POPF), the major driver of morbidity and mortality following pancreatectomy, is caused by an abnormal communication between the pancreatic ductal epithelium and another epithelial surface containing pancreas-derived, enzyme-rich fluid. There is a strong correlation between the amylase content in surgically-placed drains early in the postoperative course and the development of POPF. A simple and cheap method to determine the amylase content from the drain effluent has been eagerly advocated. Here, we developed an amylase optical biosensor, based on a surface plasmon resonance (SPR) plastic optical fiber (POF), metallized with a 60 nm layer of gold and interrogated with white light. The sensor was made specific by coupling it with an anti-amylase antibody. Each surface derivatization step was optimized and studied by XPS, contact angle, and fluorescence. The POF-biosensor was tested for its response to amylase in diluted drain effluents. The volume of sample required was 50 µL and the measurement time was 8 min. The POF-biosensor showed selectivity for amylase, a calibration curve log-linear in the range of 0.8-25.8 U/L and a limit of detection (LOD) of ~0.5 U/L. In preliminary tests, the POF-biosensor allowed for the measurement of the amylase content of diluted surgically-placed drain effluents with an accuracy of >92% with respect to the gold standard. The POF-biosensor allows for reliable measurement and could be implemented to allow for a rapid bedside assessment of amylase value in drains following pancreatectomy.
- Published
- 2021
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19. The Search for Peptide Epitopes for Molecular Imprinting Through Bioinformatics.
- Author
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Bossi AM and Pasquardini L
- Subjects
- Computational Biology, Epidemiologic Studies, Epitopes, Peptides, Molecular Imprinting
- Abstract
Epitope imprinting is an effective strategy to prepare molecularly imprinted polymers (MIPs) for protein recognition. Indeed, the idea to use as a template just a fragment of the protein of interest, called the epitope, instead of the whole protein, presents some key advantages for the imprinting process, in particular: cutting the costs for MIP production and avoiding protein unfolding during the imprinting process, so to ultimately improve the quality of the stamped binding sites. How to select an epitope for the imprinting is the strategic question. Here, the bioinformatics tools to search for suitable epitopes for the imprinting process and rational tools to select the most suitable epitope are briefly introduced along with protocols for their practical use., (© 2021. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
- Published
- 2021
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20. Tailoring a Dress to Single Protein Molecules: Proteins Can Do It Themselves through Localized Photo-Polymerization and Molecular Imprinting.
- Author
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Bossi AM and Haupt K
- Subjects
- Polymerization, Molecular Imprinting, Nanoparticles chemistry, Polymers chemistry, Proteins chemistry
- Abstract
Molecularly imprinted polymer nanoparticles (MIP NPs) are antibody-like recognition materials prepared by a template-assisted synthesis. MIP NPs able to target biomolecules, like proteins, are under the spotlight for their great potential in medicine, but efficiently imprinting biological templates is still very challenging. Here we propose generating a molecular imprint in single NPs, by photochemically initiating the polymerization from individual protein templates. In this way, each protein molecule tailors itself its own "polymeric dress". For this, the template protein is covalently coupled with a photoinitiator, Eosin Y. Irradiated with light at 533 nm, the Eosin moiety acts as an antenna and transfers energy to a co-initiator (an amine), which generates a radical and initiates polymerization. As a result, a polymer network is forming only around the very template molecule, producing cross-linked NPs of 50 nm, with single binding sites showing high affinity (K
D 10-9 m) for their biological target, and selectivity over other proteins., (© 2020 Wiley-VCH GmbH.)- Published
- 2020
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21. Molecular Imprinted Polymers Coupled to Photonic Structures in Biosensors: The State of Art.
- Author
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Chiappini A, Pasquardini L, and Bossi AM
- Subjects
- Optics and Photonics, Polymers, Surface Plasmon Resonance, Biosensing Techniques, Molecular Imprinting
- Abstract
Optical sensing, taking advantage of the variety of available optical structures, is a rapidly expanding area. Over recent years, whispering gallery mode resonators, photonic crystals, optical waveguides, optical fibers and surface plasmon resonance have been exploited to devise different optical sensing configurations. In the present review, we report on the state of the art of optical sensing devices based on the aforementioned optical structures and on synthetic receptors prepared by means of the molecular imprinting technology. Molecularly imprinted polymers (MIPs) are polymeric receptors, cheap and robust, with high affinity and selectivity, prepared by a template assisted synthesis. The state of the art of the MIP functionalized optical structures is critically discussed, highlighting the key progresses that enabled the achievement of improved sensing performances, the merits and the limits both in MIP synthetic strategies and in MIP coupling.
- Published
- 2020
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22. Does the protein corona take over the selectivity of molecularly imprinted nanoparticles? The biological challenges to recognition.
- Author
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Capriotti A, Piovesana S, Zenezini Chiozzi R, Montone CM, Bossi AM, and Laganà A
- Subjects
- Humans, Polymers, Proteomics, Molecular Imprinting, Nanoparticles, Protein Corona
- Abstract
"Plastic antibodies" are nano-sized biomimetics prepared by the molecular imprinting technology, which have the robustness of polymers, but specificity and selectivity alike natural receptors making them ideal for analytical uses. The current challenge is to translate plastic antibodies to in vivo applications for diagnosis, drug delivery, theranostic, therefore it is crucial to evaluate the effect of the biological sample complexity on the selectivity and the formation of protein corona (PCs), which ultimately dictate the fate of circulating nanoparticles. A set (n = 4) of plastic antibodies (nanoMIPs) against different proteins was prepared. Quantitative (iBAC) shotgun proteomics permitted to define the PC composition of nanoMIPs in human plasma, the relative protein abundances, the correlation between PC and the plasma dilution. NanoMIPs showed >200 proteins PC, while ~150 proteins were found on controls, suggesting the imprinting process influences the nanoparticle's structure hence the protein uptake. NanoMIPs and controls shared the 44% of the PC, but PC iBAQ values on nanoMIPs were 10-100 times higher than controls, suggesting PC/nanoMIPs interactions were far stronger than PC/non imprinted particles. PCs were richer in small proteins and in immunoglobulins, indicating a defensive response, while the selectivity was negatively challenged in the crowded plasma sample. SIGNIFICANCE: The formation and the composition of the protein corona (PC) is key to decide the fate of nanoparticles when in vivo, therefore there is the strong need to study the composition of the PC. To enable and to support the translation of the use of plastic antibodies (nanoMIPs), prepared by means of the molecular imprinting technique, to the clinical practice and to in vivo uses, the present work evaluates the effects of the complexity of the biological sample (plasma) on nanoMIPs composed of highly crosslinked polyacrylamide and acrylamide derivatives. Proteomic study offers an in depth insight of the protein corona formed in plasma on nanoMIPs. A set of nanoMIPs synthesized and raised to recognize either small or large proteins was tested. The selection abilities of the nanoMIPs when placed in plasma at different dilutions was studied. Quantitative shotgun proteomics allowed to define the composition of the formed protein corona (PC) enabling to detail the protein compositions, the relative abundances, its correlation to the biological sample composition and the correlation between PC and nanoMIP's imprinted template. In plasma, all the nanoMIPs gained a PC composed of more than 200 proteins. Type of protein recruited for the corona, molecular weight and abundance in the PC were studied. The PC on the nanoMIPs appeared to be driven by the protein composition of the plasma, while the template protein, towards which a nanoMIP was imprinted and that was proven to have high affinity for, did not influence the PC., (Copyright © 2020 Elsevier B.V. All rights reserved.)
- Published
- 2020
- Full Text
- View/download PDF
23. Deformable molecularly imprinted nanogels permit sensitivity-gain in plasmonic sensing.
- Author
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Cennamo N, Maniglio D, Tatti R, Zeni L, and Bossi AM
- Subjects
- Elastic Modulus, Humans, Limit of Detection, Biosensing Techniques methods, Molecular Imprinting methods, Nanogels chemistry, Transferrin analysis
- Abstract
Soft molecularly imprinted nanogels (nanoMIPs), selective for human transferrin (HTR), were prepared via a template assisted synthesis. Owing to their soft matter, the nanoMIPs were observed to deform at binding to HTR: while no relevant changes were observed in the hydrodynamic sizes of HTR-free compared to HTR-loaded nanoMIPs, the HTR binding resulted in a significant increment of the nanoMIP stiffness, with the mean Young's modulus measured by AFM passing from 17 ± 6 kPa to 56 ± 18 kPa. When coupled to a plastic optical fibre (POF) plasmonic platform, the analyte-induced nanoMIP-deformations amplified the resonance shift, enabling to attain ultra-low sensitivities (LOD = 1.2 fM; linear dynamic range of concentrations from 1.2 fM to 1.8 pM). Therefore, soft molecularly imprinted nanogels that obey to analyte-induced deformation stand as a novel class of sensitivity-gain structures for plasmonic sensing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)
- Published
- 2020
- Full Text
- View/download PDF
24. Molecularly Imprinted Polymers for Cell Recognition.
- Author
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Piletsky S, Canfarotta F, Poma A, Bossi AM, and Piletsky S
- Subjects
- Cell Tracking, Diagnostic Imaging, Drug Delivery Systems, Humans, Molecular Imaging, Tissue Engineering, Epitopes chemistry, Molecular Imprinting methods, Molecularly Imprinted Polymers chemical synthesis, Nanoparticles chemistry
- Abstract
Since their conception 50 years ago, molecularly imprinted polymers (MIPs) have seen extensive development both in terms of synthetic routes and applications. Cells are perhaps the most challenging target for molecular imprinting. Although early work was based almost entirely around microprinting methods, recent developments have shifted towards epitope imprinting to generate MIP nanoparticles (NPs). Simultaneously, the development of techniques such as solid phase MIP synthesis has solved many historic issues of MIP production. This review briefly describes various approaches used in cell imprinting with a focus on applications of the created materials in imaging, drug delivery, diagnostics, and tissue engineering., (Copyright © 2019 Elsevier Ltd. All rights reserved.)
- Published
- 2020
- Full Text
- View/download PDF
25. Plastic antibodies for cancer therapy?
- Author
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Bossi AM
- Subjects
- Animals, Humans, Neoplasms immunology, Polymers administration & dosage, Antibodies chemistry, Antibodies therapeutic use, Nanoparticles chemistry, Neoplasms drug therapy, Polymers chemistry
- Published
- 2020
- Full Text
- View/download PDF
26. D-shaped plastic optical fibre aptasensor for fast thrombin detection in nanomolar range.
- Author
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Cennamo N, Pasquardini L, Arcadio F, Vanzetti LE, Bossi AM, and Zeni L
- Subjects
- Aptamers, Nucleotide chemistry, Aptamers, Nucleotide genetics, Biosensing Techniques economics, Biotin chemistry, Limit of Detection, Polyethylene Glycols chemistry, Thrombin genetics, Biosensing Techniques instrumentation, Optical Fibers, Plastics chemistry, Surface Plasmon Resonance instrumentation, Thrombin analysis
- Abstract
The development of optical biosensors for the rapid and costless determination of clinical biomarkers is of paramount importance in medicine. Here we report a fast and low-cost biosensor based on a plasmonic D-shaped plastic optical fibre (POF) sensor derivatized with an aptamer specific for the recognition of thrombin, the target marker of blood homeostasis and coagulation cascade. In particular, we designed a functional interface based on a Self Assembled Monolayer (SAM) composed of short Poly Ethylene Glycol (PEG) chains and biotin-modified PEG thiol in ratio 8:2 mol:mol, these latter serving as baits for the binding of the aptamer through streptavidin-chemistry. The SAM was studied by X-ray Photoelectron Spectroscopy (XPS) analysis, static contact angle (CA), Surface Plasmon Resonance (SPR) in POFs, and fluorescence microscopy on gold surface. The optimized SAM composition enabled the immobilization of about 112 ng/cm
2 of aptamer. The thrombin detection exploiting POF-Aptasensor occurred in short times (5-10 minutes), the reached Limit of Detection (LOD) was about 1 nM, and the detection range was 1.6-60 nM, indicating the POF-Aptasensor well addresses the needs for a low-cost, simple to use and to realize, rapid, small size and portable diagnostic platform.- Published
- 2019
- Full Text
- View/download PDF
27. Selective PQQPFPQQ Gluten Epitope Chemical Sensor with a Molecularly Imprinted Polymer Recognition Unit and an Extended-Gate Field-Effect Transistor Transduction Unit.
- Author
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Iskierko Z, Sharma PS, Noworyta KR, Borowicz P, Cieplak M, Kutner W, and Bossi AM
- Subjects
- Amino Acid Sequence, Electrodes, Enzyme-Linked Immunosorbent Assay, Epitopes analysis, Epitopes chemistry, Flour analysis, Glutens chemistry, Gold chemistry, Limit of Detection, Glutens analysis, Molecular Imprinting methods, Polymers chemistry, Transistors, Electronic
- Abstract
A molecularly imprinted polymer (MIP) recognition system was devised for selective determination of an immunogenic gluten octamer epitope, PQQPFPQQ. For that, a thin MIP film was devised, guided by density functional theory calculations, and then synthesized to become the chemosensor recognition unit. Bis(bithiophene)-based cross-linking and functional monomers were used for this synthesis. An extended-gate field-effect transistor (EG-FET) was used as the transduction unit. The EG-FET gate surface was coated with the PQQPFPQQ-templated MIP film, by electropolymerization, to result in a complete chemosensor. X-ray photoelectron spectroscopy analysis confirmed the presence of the PQQPFPQQ epitope, and its removal from the MIP film. The chemosensor selectively discriminated between the octamer analyte and another peptide of the same number of amino acids but with two of them mismatched (PQQQFPPQ). The chemosensor was validated with respect to both the PQQPFPQQ analyte and a real gluten extract from semolina flour. It was capable to determine PQQPFPQQ in the concentration range of 0.5-45 ppm with the limit of detection (LOD) = 0.11 ppm. Moreover, it was capable of determining gluten in real samples in the concentration range of 4-25 ppm with LOD = 4 ppm, which is a value sufficient for discriminating between gluten-free and non-gluten-free food products. The gluten content in semolina flour determined with the chemosensor well correlated with that determined with a commercial ELISA gluten kit. The Langmuir, Freundlich, and Langmuir-Freundlich isotherms were fitted to the epitope sorption data. The sorption parameters determined from these isotherms indicated that the imprinted cavities were quite homogeneous and that the epitope analyte was chemisorbed in them.
- Published
- 2019
- Full Text
- View/download PDF
28. Development of an in-house mixed-mode solid-phase extraction for the determination of 16 basic drugs in urine by High Performance Liquid Chromatography-Ion Trap Mass Spectrometry.
- Author
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Musile G, Cenci L, Piletska E, Gottardo R, Bossi AM, and Bortolotti F
- Subjects
- Humans, Chromatography, High Pressure Liquid methods, Mass Spectrometry methods, Pharmaceutical Preparations isolation & purification, Pharmaceutical Preparations urine, Solid Phase Extraction methods
- Abstract
The aim of the present work was to develop a novel in-house mixed-mode SPE sorbent to be used for the HPLC-Ion TrapMS determination of 16 basic drugs in urine. By using a computational modelling, a virtual monomer library was screened identifying three suitable functional monomers, methacrylic acid (MAA), itaconic acid (IA) and 2-acrylamide-2-methylpropane sulfonic acid (AMPSA), respectively. Three different sorbents were then synthetized based on these monomers, and using as cross-linker trimethylolpropane trimethacrylate (TMPTMA). The sorbent characterization analyses brought to the selection of the AMPSA based phase. Using this novel in-house sorbent, a SPE-HPLC-Ion TrapMS method for drug analysis in urine was validated proving to be selective and accurate and showing a sensitivity adequate for toxicological urine analysis. The comparison of the in-house mixed-mode SPE sorbent with two analogous commercial mixed-mode SPE phases showed that the first one was better not only in terms of process efficiency, but also in terms of quality-price rate. To the best of our knowledge, this is the first time in which an in-house SPE procedure has been applied to the toxicological analysis of a complex matrix, such as urine., (Copyright © 2018 Elsevier B.V. All rights reserved.)
- Published
- 2018
- Full Text
- View/download PDF
29. MIRATE: MIps RATional dEsign Science Gateway.
- Author
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Busato M, Distefano R, Bates F, Karim K, Bossi AM, López Vilariño JM, Piletsky S, Bombieri N, and Giorgetti A
- Subjects
- Hepcidins metabolism, Humans, Molecular Dynamics Simulation, Receptors, Artificial metabolism, Troponin I metabolism, Hepcidins chemistry, Molecular Imprinting methods, Polymers chemistry, Receptors, Artificial chemistry, Troponin I chemistry
- Abstract
Molecularly imprinted polymers (MIPs) are high affinity robust synthetic receptors, which can be optimally synthesized and manufactured more economically than their biological equivalents (i.e. antibody). In MIPs production, rational design based on molecular modeling is a commonly employed technique. This mostly aids in (i) virtual screening of functional monomers (FMs), (ii) optimization of monomer-template ratio, and (iii) selectivity analysis. We present MIRATE, an integrated science gateway for the intelligent design of MIPs. By combining and adapting multiple state-of-the-art bioinformatics tools into automated and innovative pipelines, MIRATE guides the user through the entire process of MIPs' design. The platform allows the user to fully customize each stage involved in the MIPs' design, with the main goal to support the synthesis in the wet-laboratory. Availability: MIRATE is freely accessible with no login requirement at http://mirate.di.univr.it/. All major browsers are supported.
- Published
- 2018
- Full Text
- View/download PDF
30. A microRNA signature from serum exosomes of patients with glioma as complementary diagnostic biomarker.
- Author
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Santangelo A, Imbrucè P, Gardenghi B, Belli L, Agushi R, Tamanini A, Munari S, Bossi AM, Scambi I, Benati D, Mariotti R, Di Gennaro G, Sbarbati A, Eccher A, Ricciardi GK, Ciceri EM, Sala F, Pinna G, Lippi G, Cabrini G, and Dechecchi MC
- Subjects
- Adult, Aged, Biomarkers, Tumor, Brain Neoplasms surgery, Diagnosis, Differential, Female, Glioma surgery, Humans, Male, Middle Aged, Sensitivity and Specificity, Brain Neoplasms blood, Brain Neoplasms diagnosis, Exosomes metabolism, Glioma blood, Glioma diagnosis, MicroRNAs blood
- Abstract
Malignant gliomas, the most frequent primary brain tumors, are characterized by a dismal prognosis. Reliable biomarkers complementary to neuroradiology in the differential diagnosis of gliomas and monitoring for post-surgical progression are unmet needs. Altered expression of several microRNAs in tumour tissues from patients with gliomas compared to normal brain tissue have been described, thus supporting the rationale of using microRNA-based biomarkers. Although different circulating microRNAs were proposed in association with gliomas, they have not been introduced into clinical practice so far. Blood samples were collected from patients with high and low grade gliomas, both before and after surgical resection, and the expression of miR-21, miR-222 and miR-124-3p was measured in exosomes isolated from serum. The expression levels of miR-21, miR-222 and miR-124-3p in serum exosomes of patients with high grade gliomas were significantly higher than those of low grade gliomas and healthy controls and were sharply decreased in samples obtained after surgery. The analysis of miR-21, miR-222 and miR-124-3p in serum exosomes of patients affected by gliomas can provide a minimally invasive and innovative tool to help the differential diagnosis of gliomas at their onset in the brain and predict glioma grading and non glial metastases before surgery.
- Published
- 2018
- Full Text
- View/download PDF
31. Micro- versus nano-sized molecularly imprinted polymers in MALDI-TOF mass spectrometry analysis of peptides.
- Author
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Cenci L, Bertolla M, Anesi A, Ambrosi E, Guella G, and Bossi AM
- Subjects
- Crystallization, Humans, Nanostructures chemistry, Peptides analysis, Peptides isolation & purification, Solid Phase Extraction methods, Molecular Imprinting methods, Peptides blood, Polymers chemistry, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods
- Abstract
The integration of molecularly imprinted polymers (MIPs) with MALDI-TOF mass spectrometry (MS) combines MIP selectivity with MS sensitivity. Whether the size of the MIP material-micro versus nano-has an effect on the MS analysis was the object of the study. MIPs, targeting respectively the epitope peptide NR11 of cardiac troponin I and the peptide CK13 of human serum transferrin, were synthesized and characterized. The size-related performance of the MIP materials hyphenated with MALDI-TOF-MS analysis was studied by the incubation of the target peptide with the respective micro- or nano-MIP, followed by rinsing to remove non-specific deposition of the MIP to the MALDI target plate, co-crystallization with the organic matrix, and mass analysis. The quality of the MS analysis was assessed comparing the S/N of the mass peaks of the MIP-bound peptide to that of the same quantity of free peptide. Sweet spots and lower S/N (~ 1 order of magnitude) were observed for micro-MIP materials, while in the case of nano-MIP-bound peptide, the S/N was comparable to that of the free peptide, indicating higher compatibility of the nano-MIPs to MALDI-TOF-MS. The nano-MIP/MALDI-TOF-MS permitted the selective determination of the target peptide in real serum samples. Graphical abstract ᅟ.
- Published
- 2017
- Full Text
- View/download PDF
32. Solvent-Responsive Molecularly Imprinted Nanogels for Targeted Protein Analysis in MALDI-TOF Mass Spectrometry.
- Author
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Bertolla M, Cenci L, Anesi A, Ambrosi E, Tagliaro F, Vanzetti L, Guella G, and Bossi AM
- Subjects
- Humans, Molecular Imprinting, Nanoparticles, Polyethylene Glycols, Polyethyleneimine, Solvents, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
- Abstract
Molecular imprinted poly(acrylamido)-derivative nanogels have shown their selectivity to bind the protein human serum transferrin (HTR) and also showed their capability for instantaneous solvent-induced modification upon the addition of acetonitrile. Integrated to matrix-assisted laser desorption/ionization time-of-flight mass analysis the HTR-imprinted solvent-responsive nanogels permitted the determination of HTR straight from serum and offered novel perspectives in targeted protein analysis.
- Published
- 2017
- Full Text
- View/download PDF
33. Guided folding takes a start from the molecular imprinting of structured epitopes.
- Author
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Cenci L, Guella G, Andreetto E, Ambrosi E, Anesi A, and Bossi AM
- Abstract
A biomimetic route towards assisted folding was explored. Molecularly imprinted polymeric nanoparticles (MIP NPs), i.e. biomimetics with entailed molecular recognition properties made by a template assisted synthesis, were prepared to target a structured epitope: the cystine containing peptide CC9ox, which corresponds to the apical portion of the β-hairpin hormone Hepcidin-25. The structural selection was achieved by the MIP NPs; moreover, the MIP NPs demonstrated favouring the folding of the linear random peptide (CC9red) into the structured one (CC9ox), anticipating the future role of the MIP NPs as in situ nanomachines to counteract folding defects.
- Published
- 2016
- Full Text
- View/download PDF
34. Screening of the binding properties of molecularly imprinted nanoparticles via capillary electrophoresis.
- Author
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Musile G, Cenci L, Andreetto E, Ambrosi E, Tagliaro F, and Bossi AM
- Subjects
- Binding Sites, Chromatography, High Pressure Liquid, Hydrogen-Ion Concentration, Ligands, Temperature, Electrophoresis, Capillary methods, Molecular Imprinting methods, Nanoparticles
- Abstract
In response to the need for straightforward analytical methods to assess the affinity of molecularly imprinted nanoparticles (MIP NPs) for ligands, capillary electrophoresis (CE) was exploited using MIP NPs targeting the iron-regulating hormone hepcidin. In this work, MIP NPs were challenged with their template peptide, i.e., the N-terminal 5-mer of hepcidin, in comparison to unrelated ligand peptides. A CE separation method was developed ex novo achieving, after optimization of the background electrolyte (150 mM sodium phosphate pH 7.4) and of the running temperature (35 °C), the full separation of the free ligand from the complexed MIP NPs. The CE binding isotherm allowed the estimation of a micromolar dissociation constant for the 5-mer template-MIP NPs complex, in agreement with independent measurements. The CE offered the advantages of a direct injection of the MIP NPs/ligand incubation mix, without preliminary fractionation steps, requiring only minimal sample volumes and short analysis times. In conclusion CE proved to be a valid technique for characterizing the interactions of MIP NP libraries for selected target compounds.
- Published
- 2016
- Full Text
- View/download PDF
35. Molecularly imprinted polymers coupled to matrix assisted laser desorption ionization mass spectrometry for femtomoles detection of cardiac troponin I peptides.
- Author
-
Cenci L, Anesi A, Busato M, Guella G, and Bossi AM
- Subjects
- Humans, Limit of Detection, Molecular Imprinting methods, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods, Polymers chemistry, Troponin I analysis
- Abstract
Molecularly imprinted polymers (MIPs) were combined to MALDI-TOF-MS to evaluate a selective enrichment (SE) method for the determination of clinically relevant biomarkers from complex biological samples. The concept was proven with the myocardial injury marker Troponin I (cTnI). In a first part, MIP materials entailed for the recognition of cTnI epitopes (three peptides selected) were prepared and characterized in dimensions (0.7-2μm), dissociation constants (58-817 nM), kinetics of binding (5-60 min), binding capacity (ca. 1.5 µg/mg polymer), imprinting factors (3 > IF > 5) and selectivity for the peptide epitope. Then, the MIPs, incubated with cTnI peptides and spotted on the target with the DHB matrix, were assayed for the desorption of the peptides in MALDI-TOF-MS. The measured detection limit was ca. 300 femtomols. Finally, the MIP-SE MALDI-TOF-MS was tested for its ability to enrich in the cTnI peptides from a complex sample, mimic of serum (i.e. 81 peptides of digested albumin). The MIP-SE MALDI-TOF-MS successfully enriched in cTnI peptides from the complex sample proving the technique could offer a flexible platform to prepare entailed materials suitable for diagnostic purposes., (Copyright © 2015 John Wiley & Sons, Ltd.)
- Published
- 2016
- Full Text
- View/download PDF
36. Surface plasmon resonance based on molecularly imprinted nanoparticles for the picomolar detection of the iron regulating hormone Hepcidin-25.
- Author
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Cenci L, Andreetto E, Vestri A, Bovi M, Barozzi M, Iacob E, Busato M, Castagna A, Girelli D, and Bossi AM
- Subjects
- Hepcidins metabolism, Humans, Iron metabolism, Limit of Detection, Hepcidins blood, Molecular Imprinting, Nanoparticles chemistry, Surface Plasmon Resonance methods
- Abstract
Background: Molecularly imprinted polymer (MIP) technique is a powerful mean to produce tailor made synthetic recognition sites. Here precipitation polymerization was exploited to produce a library of MIP nanoparticles (NPs) targeting the N terminus of the hormone Hepcidin-25, whose serum levels correlate with iron dis-metabolisms and doping. Biotinylated MIP NPs were immobilized to NeutrAvidin™ SPR sensor chip. The response of the MIP NP sensor to Hepcidin-25 was studied., Findings: Morphological analysis showed MIP NPs of 20-50 nm; MIP NP exhibited high affinity and selectivity for the target analyte: low nanomolar Kds for the interaction NP/Hepcidin-25, but none for the NP/non regulative Hepcidin-20. The MIP NP were integrated as recognition element in SPR allowing the detection of Hepcidin-25 in 3 min. Linearity was observed with the logarithm of Hepcidin-25 concentration in the range 7.2-720 pM. LOD was 5 pM. The response for Hepcidin-20 was limited. Hepcidin-25 determination in real serum samples spiked with known analyte concentrations was also attempted., Conclusion: The integration of MIP NP to SPR allowed the determination of Hepcidin-25 at picomolar concentrations in short times outperforming the actual state of art. Optimization is still needed for real sample measurements in view of future clinical applications.
- Published
- 2015
- Full Text
- View/download PDF
37. Murine macrophages response to iron.
- Author
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Polati R, Castagna A, Bossi AM, Alberio T, De Domenico I, Kaplan J, Timperio AM, Zolla L, Gevi F, D'Alessandro A, Brunch R, Olivieri O, and Girelli D
- Subjects
- Animals, Bone Marrow Cells cytology, Bone Marrow Cells immunology, Cells, Cultured, Ferric Compounds pharmacology, Gene Expression Profiling, Gene Expression Regulation drug effects, Iron immunology, Macrophages cytology, Macrophages immunology, Metabolome drug effects, Metabolomics methods, Mice, Proteome drug effects, Proteome immunology, Proteomics methods, Quaternary Ammonium Compounds pharmacology, Bone Marrow Cells metabolism, Gene Expression Regulation physiology, Iron metabolism, Macrophages metabolism, Metabolome physiology, Proteome metabolism
- Abstract
Macrophages play a critical role at the crossroad between iron metabolism and immunity, being able to store and recycle iron derived from the phagocytosis of senescent erythrocytes. The way by which macrophages manage non-heme iron at physiological concentration is still not fully understood. We investigated protein changes in mouse bone marrow macrophages incubated with ferric ammonium citrate (FAC 10 μM iron). Differentially expressed spots were identified by nano RP-HPLC-ESI-MS/MS. Transcriptomic, metabolomics and western immunoblotting analyses complemented the proteomic approach. Pattern analysis was also used for identifying networks of proteins involved in iron homeostasis. FAC treatment resulted in higher abundance of several proteins including ferritins, cytoskeleton related proteins, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) at the membrane level, vimentin, arginase, galectin-3 and macrophage migration inhibitory factor (MIF). Interestingly, GAPDH has been recently proposed to act as an alternative transferrin receptor for iron acquisition through internalization of the GAPDH-transferrin complex into the early endosomes. FAC treatment also induced the up-regulation of oxidative stress-related proteins (PRDX), which was further confirmed at the metabolic level (increase in GSSG, 8-isoprostane and pentose phosphate pathway intermediates) through mass spectrometry-based targeted metabolomics approaches. This study represents an example of the potential usefulness of "integarated omics" in the field of iron biology, especially for the elucidation of the molecular mechanisms controlling iron homeostasis in normal and disease conditions. This article is part of a Special Issue entitled: Integrated omics., (Copyright © 2012 Elsevier B.V. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
38. Fingerprint-imprinted polymer: rational selection of peptide epitope templates for the determination of proteins by molecularly imprinted polymers.
- Author
-
Bossi AM, Sharma PS, Montana L, Zoccatelli G, Laub O, and Levi R
- Subjects
- Amino Acid Sequence, Cardiovascular Diseases diagnosis, Humans, Natriuretic Peptide, Brain isolation & purification, Natriuretic Peptide, Brain metabolism, Peptide Fragments isolation & purification, Peptide Fragments metabolism, Peptides metabolism, Polymers metabolism, Protein Binding, Molecular Imprinting methods, Natriuretic Peptide, Brain blood, Peptide Fragments blood, Peptides chemistry, Polymers chemistry
- Abstract
The pool of peptides composing a protein allows for its distinctive identification in a process named fingerprint (FP) analysis. Here, the FP concept is used to develop a method for the rational preparation of molecularly imprinted polymers (MIPs) for protein recognition. The fingerprint imprinting (FIP) is based on the following: (1) the in silico cleavage of the protein sequence of interest with specific agents; (2) the screening of all the peptide sequences generated against the UniProtKB database in order to allow for the rational selection of distinctive and unique peptides (named as epitopes) of the target protein; (3) the selected epitopes are synthesized and used as templates for the molecular imprinting process. To prove the principle, NT-proBNP, a marker of the risk of cardiovascular events, was chosen as an example. The in silico analysis of the NT-proBNP sequence allowed us to individuate the peptide candidates, which were next used as templates for the preparation of NT-pro-BNP-specific FIPs and tested for their ability to bind the NT-proBNP peptides in complex samples. Results indicated an imprinting factor, IF, of ~10, a binding capacity of 0.5-2 mg/g, and the ability to rebind 40% of the template in a complex sample, composed of the whole digests of NT-proBNP.
- Published
- 2012
- Full Text
- View/download PDF
39. Monocyte/macrophage proteomics: recent findings and biomedical applications.
- Author
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Castagna A, Polati R, Bossi AM, and Girelli D
- Subjects
- Animals, Arteriosclerosis metabolism, Arteriosclerosis pathology, Biomarkers metabolism, Humans, Inflammation metabolism, Iron metabolism, Proteomics, Terminology as Topic, Macrophages metabolism, Monocytes metabolism, Proteome metabolism
- Abstract
Macrophages, originating from the migration and differentiation of circulating monocytes into virtually all tissues, are extremely flexible and plastic cells that play vital homeostatic roles, but also contribute to the pathophysiology of many human diseases. For these reasons, they are intensively studied by different approaches, recently including proteomics. Macrophage cells can be taken from a range of different sources, including blood monocytes and macrophages from tissues. Macrophages can also be generated by in vitro culture from blood monocytes, and cell lines derived from this lineage can be used. Similarly, many different proteomic techniques can be used, ranging from classic approaches based on 2D gel electrophoresis to more recent high-throughput gel-free techniques essentially based on mass spectrometry. Here, we review the application of such techniques to the study of monocytes/macrophages, and summarize some results potentially relevant to two paradigmatic conditions - atherosclerosis and disorders of iron metabolism.
- Published
- 2012
- Full Text
- View/download PDF
40. Proteomic analysis of dopamine and α-synuclein interplay in a cellular model of Parkinson's disease pathogenesis.
- Author
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Alberio T, Bossi AM, Milli A, Parma E, Gariboldi MB, Tosi G, Lopiano L, and Fasano M
- Subjects
- Cell Line, Electrophoresis, Gel, Two-Dimensional, Gene Expression, Humans, In Vitro Techniques, Metabolic Networks and Pathways, Models, Neurological, NF-kappa B metabolism, Parkinson Disease genetics, Proteomics, Tandem Mass Spectrometry, alpha-Synuclein genetics, Dopamine metabolism, Parkinson Disease etiology, Parkinson Disease metabolism, alpha-Synuclein metabolism
- Abstract
Altered dopamine homeostasis is an accepted mechanism in the pathogenesis of Parkinson's disease. α-Synuclein overexpression and impaired disposal contribute to this mechanism. However, biochemical alterations associated with the interplay of cytosolic dopamine and increased α-synuclein are still unclear. Catecholaminergic SH-SY5Y human neuroblastoma cells are a suitable model for investigating dopamine toxicity. In the present study, we report the proteomic pattern of SH-SY5Y cells overexpressing α-synuclein (1.6-fold induction) after dopamine exposure. Dopamine itself is able to upregulate α-synuclein expression. However, the effect is not observed in cells that already overexpress α-synuclein as a consequence of transfection. The proteomic analysis highlights significant changes in 23 proteins linked to specific cellular processes, such as cytoskeleton structure and regulation, mitochondrial function, energetic metabolism, protein synthesis, and neuronal plasticity. A bioinformatic network enrichment procedure generates a significant model encompassing all proteins and allows us to enrich functional categories associated with the combination of factors analyzed in the present study (i.e. dopamine together with α-synuclein). In particular, the model suggests a potential involvement of the nuclear factor kappa B pathway that is experimentally confirmed. Indeed, α-synuclein significantly reduces nuclear factor kappa B activation, which is completely quenched by dopamine treatment., (© 2010 The Authors Journal compilation © 2010 FEBS.)
- Published
- 2010
- Full Text
- View/download PDF
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