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Innovative goods authentication strategies are of fundamental importance considering the increasing counterfeiting levels. Such a task has been effectively addressed with the so-called physical unclonable functions (PUFs), being physical properties of a system that characterize it univocally. PUFs are commonly implemented by exploiting naturally occurring non-idealities in clean-room fabrication processes. The broad availability of classic paradigm PUFs, however, makes them vulnerable. Here, we propose a hybrid plasmonic/photonic multilayered structure working as a three-level strong PUF. Our approach leverages on the combination of a functional nanostructured surface, a resonant response, and a unique chromatic signature all together in one single device. The structure consists of a resonant cavity, where the top mirror is replaced with a layer of plasmonic Ag nanoislands. The naturally random spatial distribution of clusters and nanoparticles formed by this deposition technique constitutes the manufacturer-resistant nanoscale morphological fingerprint of the proposed PUF. The presence of Ag nanoislands allows us to tailor the interplay between the photonic and plasmonic modes to achieve two additional security levels. The first one is constituted by the chromatic response and broad iridescence of our structures, while the second by their rich spectral response, accessible even through a common smartphone light-emitting diode. We demonstrate that the proposed architectures could also be used as an irreversible and quantitative temperature exposure label. The proposed PUFs are inexpensive, chip-to-wafer-size scalable, and can be deposited over a variety of substrates. They also hold a great promise as an encryption framework envisioning morpho-cryptography applications.
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Using a surface forces apparatus and an atomic force microscope, we characterized the adhesive properties of adsorbed layers of two recombinant variants of Perna viridis foot protein 5 (PVFP-5), the main surface-binding protein in the adhesive plaque of the Asian green mussel. In one variant, all tyrosine residues were modified into 3,4-dihydroxy-l-phenylalanine (DOPA) during expression using a residue-specific incorporation strategy. DOPA is a key molecular moiety underlying underwater mussel adhesion. In the other variant, all tyrosine residues were preserved. The layer was adsorbed on a mica substrate and pressed against an uncoated surface. While DOPA produced a stronger adhesion than tyrosine in contact with the nanoscopic Si3N4 probe of the atomic force microscope, the two variants produced comparable adhesion on the curved macroscopic mica surfaces of the surface forces apparatus. These findings show that the presence of DOPA is not a sufficient condition to generate strong underwater adhesion. Surface chemistry and contact geometry affect the strength and abundance of protein-surface bonds created during adsorption and surface contact. Importantly, the adsorbed protein layer has a random and dynamic polymer-network structure that should be optimized to transmit the tensile stress generated during surface separation to DOPA surface bonds rather than other weaker bonds.
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Adhesivos/química , Dihidroxifenilalanina/análisis , Tirosina/análisis , Adsorción , Secuencia de Aminoácidos , Animales , Bivalvos , Microscopía de Fuerza Atómica , Proteínas Recombinantes/química , Propiedades de SuperficieRESUMEN
Keratoconus is the primary cause of corneal transplantation in young adults worldwide. Riboflavin/UV-A corneal cross-linking may effectively halt the progression of keratoconus if an adequate amount of riboflavin enriches the corneal stroma and is photo-oxidated by UV-A light for generating additional cross-linking bonds between stromal proteins and strengthening the biomechanics of the weakened cornea. Here we reported an UV-A theranostic prototype device for performing corneal cross-linking with the ability to assess corneal intrastromal concentration of riboflavin and to estimate treatment efficacy in real time. Seventeen human donor corneas were treated according to the conventional riboflavin/UV-A corneal cross-linking protocol. Ten of these tissues were probed with atomic force microscopy in order to correlate the intrastromal riboflavin concentration recorded during treatment with the increase in elastic modulus of the anterior corneal stroma. The intrastromal riboflavin concentration and its consumption during UV-A irradiation of the cornea were highly significantly correlated (R = 0.79; P = .03) with the treatment-induced stromal stiffening effect. The present study showed an ophthalmic device that provided an innovative, non-invasive, real-time monitoring solution for estimating corneal cross-linking treatment efficacy on a personalized basis.
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Córnea/metabolismo , Dispositivos Ópticos , Riboflavina/metabolismo , Fenómenos Biomecánicos , Fluorescencia , Humanos , Factores de Tiempo , Rayos UltravioletaRESUMEN
Several phenolic compounds bind to proteins and show the ability to interfere with their aggregation process. The impact of the natural polyphenol resveratrol on the stability and heat induced aggregation of human serum albumin (HSA) was investigated by differential scanning calorimetry (DSC), attenuated total reflectance Fourier transform infrared (ATR-FTIR), UV-vis absorbance, ThT fluorescence, atomic force microscopy (AFM) and molecular modeling. The binding of resveratrol to HSA improves the stability of the protein to thermal unfolding, particularly for the energetic domain containing the ligand binding site, as modeled by computational techniques. The thermal unfolding is irreversible and after the melting the protein aggregates, either with or without the ligand. The kinetics of HSA aggregation between 70 and 80°C shows an exponential growth of the absorbance change and it slows down when resveratrol is added. The aggregates have fibril-like morphology and resveratrol attenuates the formation of ß-structured species. The overall results suggest that resveratrol stabilizes the protein structure and modulates the formation of fibrils along the initial stage of the HSA aggregation pathway.
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Agregado de Proteínas , Albúmina Sérica/química , Estilbenos/química , Sitios de Unión , Calor , Humanos , Cinética , Unión Proteica , Dominios Proteicos , Estabilidad Proteica , Estructura Secundaria de Proteína , Desplegamiento Proteico , Resveratrol , TermodinámicaRESUMEN
PURPOSE: To investigate the depth-dependent mechanical anisotropy of the human corneal stroma at the tissue (stroma) and molecular (collagen) level by using atomic force microscopy (AFM). METHODS: Eleven human donor corneas were dissected at different stromal depths by using a microkeratome. Mechanical measurements were performed in 15% dextran on the surface of the exposed stroma of each sample by using a custom-built AFM in force spectroscopy mode using both microspherical (38-µm diameter) and nanoconical (10-nm radius of curvature) indenters at 2-µm/s and 15-µm/s indentation rates. Young's modulus was determined by fitting force curve data using the Hertz and Hertz-Sneddon models for a spherical and a conical indenter, respectively. The depth-dependent anisotropy of stromal elasticity was correlated with images of the corneal stroma acquired by two-photon microscopy. RESULTS: The force curves were obtained at stromal depths ranging from 59 to 218 µm. At the tissue level, Young's modulus (ES) showed a steep decrease at approximately 140-µm stromal depth (from 0.8 MPa to 0.3 MPa; P = 0.03) and then was stable in the posterior stroma. At the molecular level, Young's modulus (EC) was significantly greater than at the tissue level; EC decreased nonlinearly with increasing stromal depth from 3.9 to 2.6 MPa (P = 0.04). The variation of microstructure through the thickness correlated highly with a nonconstant profile of the mechanical properties in the stroma. CONCLUSIONS: The corneal stroma exhibits unique anisotropic elastic behavior at the tissue and molecular levels. This knowledge may benefit modeling of corneal behavior and help in the development of biomimetic materials.
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Sustancia Propia/fisiología , Elasticidad/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Anisotropía , Colágeno/fisiología , Módulo de Elasticidad , Diagnóstico por Imagen de Elasticidad , Femenino , Humanos , Masculino , Microscopía de Fuerza Atómica/métodos , Persona de Mediana Edad , Modelos Teóricos , Adulto JovenRESUMEN
The energetics of amyloid fibrillar aggregation of ß-lactoglobulin (ßLG) following incubation at high temperature and acid pH was studied by differential scanning calorimetry in the presence of Cu(2+) or Fe(3+) cations, and without any metal. Cu(2+) and metal-free protein solutions showed a distinct exothermic response that disappeared almost completely when the Fe(3+) molar concentration was ten times greater than the ßLG concentration. Thioflavin T fluorescence studies in solution and atomic force microscopy analysis of the deposit left on flat mica substrates by heat-incubated ßLG solutions correlated the absence of exothermic response of Fe(3+)-ßLG solutions with a lack of fibril production. In contrast, abundant fibril deposits were observed for Cu(2+)-ßLG solutions, with a rich polymorphism of multistrand fibrillar structures. Electron paramagnetic resonance revealed that Fe(3+) permanently binds to ßLG in the aggregate state whereas Cu(2+) plays a catalytic role without binding to the protein. We propose that Fe(3+) inhibits fibril production after binding to a key region of the protein sequence, possibly interfering with the nucleation step of the fibrillation process and opening a nonfibrillar aggregation pathway. These findings suggest that transition metal ions can be utilized to effectively modulate protein self-assembly into a variety of structures with distinct morphologies at the nanoscale level.
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Amiloide/química , Hierro/química , Lactoglobulinas/química , Amiloide/efectos de los fármacos , Calor , Concentración de Iones de Hidrógeno , Hierro/farmacología , Lactoglobulinas/metabolismo , Unión ProteicaRESUMEN
We present a method to calibrate the light to heat conversion in an aqueous fluid containing nanoparticles. Accurate control of light and heat is of dramatic importance in many fields of science and metal nanoparticles have acquired an increased importance as means to address heat in very small areas when irradiated with an intense light. The proposed method enables to measure the temperature in the environment surrounding nanoparticles, as a function of the exposure time to laser radiation, exploiting the properties of thermochromic cholesteric liquid crystals. This method overcomes the problems of miscibility of nanoparticles in liquid crystals, provides temperature reading at the microscale, since the cholesteric liquid crystal is confined in microdroplets, and it is sensitive to a temperature variation, 28°C-49°C, suitable for biological applications.
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In this paper we report how the confining surfaces and the ionic effects of different concentration of guanosine solution can be used to vary the alignment of liquid crystal phases of guanosine nucleotides. Liquid crystal phases of guanosine 5'-monophosphate ammonium salt and guanosine 5'-monophosphate free acid in pure water, with and without silver sulphate, were studied by polarized optical microscope. A periodic modulation of the texture was observed. This modulation depends on both on the concentration and on the presence of silver ions in the liquid crystal phase. We demonstrate that, according to the surface energy of the alignment layers, it is possible to homeotropically align the guanosine chromonic phase without applying any external magnetic field. Finally, we report the formation of spherical, vesicle-like guanosine 5'-monophosphate aggregates, when the solution was confined between two hydrophobic surfaces containing exposed Si groups.
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Guanosina Monofosfato/química , Cristales Líquidos/química , Plata/química , Interacciones Hidrofóbicas e Hidrofílicas , Microscopía de Polarización , Sulfatos , Propiedades de Superficie , TermodinámicaRESUMEN
A new type of flexible cholesteric liquid crystal mirror is presented. The simple and effective method for the deposition of a cholesteric mixture on a paper substrate and the particular design of the device give a homogeneous alignment of the cholesteric texture providing mirrors with an intense and uniform light reflectance. A desired polarization state for the reflected light, linear or circular, can be easily obtained varying the thickness and optical anisotropy of the polymer cover film. By using non-azobenzene based photosensitive materials a permanent array of RGB mirrors with high reflectivity can be obtained on the same device. Paper like reflective mirrors are versatile and they can find applications in reflective displays, adaptive optics, UV detectors and dosimeters, information recording, medicine and IR converters.
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Colesterol/química , Cristales Líquidos/química , Óptica y Fotónica , Papel , Luz , Polimerizacion , Espectrofotometría UltravioletaRESUMEN
PURPOSE: To analyze the interface quality of the anterior stroma after femtosecond laser flap creation using atomic force microscopy. SETTING: IRCCS Fondazione G.B. Bietti, Rome, Italy. DESIGN: Experimental study. METHODS: A 110 µm depth flap was created in 20 human corneal tissues using a femtosecond laser platform (Intralase iFS). Tissues were divided into 4 groups of various cutting parameters: pulse energy and spot separation of 0.75 µJ and 6 µm (Group 1), 0.65 µJ and 5 µm (Group 2), 0.55 µJ and 4 µm (Group 3), and 0.45 µJ and 4 µm (Group 4). Four additional tissue sections were cut using a motorized microkeratome (Hansatome). Atomic force microscopy (Autoprobe CP) analysis was performed on the stromal bed of each sample. RESULTS: The corneal tissues treated with higher pulse energies and wider spot separations (Groups 1 and 2) showed a rougher stromal bed interface (root mean square [RMS] rough = 0.23 µm ± 0.008 (SD) and 0.24 ± 0.009 µm, respectively) than tissues in Groups 3 and 4 (RMS rough = 0.18 ± 0.006 µm and 0.18 ± 0.008 µm, respectively; P<.001, 1-way analysis of variance). The stromal surface quality of tissues treated with pulse energies of 0.55 µJ or lower and 4 µm spot separation compared favorably with that of tissues cut by the microkeratome (RMS rough = 0.17 ± 0.006 µm; P>.05, Tukey). CONCLUSIONS: The femtosecond stromal interface quality was improved with pulse energy lower and spot separations narrower than those currently used in the clinical setting. The flap interface smoothness created by the femtosecond laser was comparable to that created by the microkeratome. FINANCIAL DISCLOSURE: No author has a financial or proprietary interest in any material or method mentioned.
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Sustancia Propia/patología , Queratomileusis por Láser In Situ , Láseres de Excímeros/uso terapéutico , Colgajos Quirúrgicos/patología , Anciano , Sustancia Propia/cirugía , Humanos , Microscopía de Fuerza Atómica , Modelos Biológicos , Propiedades de Superficie , Donantes de TejidosRESUMEN
PURPOSE: To analyze the effects of femtosecond laser pulses on the posterior human corneal stroma with atomic force microscopy (AFM) and environmental scanning electron microscopy (ESEM). METHODS: A femtosecond laser (IntraLase iFS, Abbott, USA) was programmed to create a full posterior lamellar dissection in 9 human corneal tissues, using 3 different pulse energies (1.00 µJ, 0.75 µJ, and 0.50 µJ). Three corneal tissues were prepared in a similar fashion using a mechanical microkeratome (Moria Evolution 3, Moria, France). Six corneal tissues received an 8.00-mm diameter full cylindrical resection using either the femtosecond laser or the Barron trephine (Katena Products Inc., USA). The posterior corneal lenticules were first examined at AFM (Autoprobe CP, Veeco, USA). Both the posterior lenticules and the trephined corneal samples were scanned by ESEM (FEI Quanta 400, USA). RESULTS: Granules and crater-like features were observed on the stromal interface of all the laser dissected tissues, likely due to a secondary thermal effect of femtosecond laser dissection. Collagen fibers were seen only on samples treated with the 0.50 µJ pulse energy. Images of an even stromal surface were observed on the posterior stroma of mechanically dissected corneal samples. CONCLUSIONS: Mechanical and thermal effects, induced by femtosecond laser pulses on the human corneal stroma, were seen with AFM. Surface regularity of the photodisrupted stroma was inversely and non-linearly related to the pulse energy. The femtosecond laser provided high surface quality for lamellar resection of the posterior stroma comparable to those provided by mechanical devices.
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Sustancia Propia/cirugía , Sustancia Propia/ultraestructura , Terapia por Láser , Láseres de Excímeros/uso terapéutico , Microscopía de Fuerza Atómica , Anciano , Humanos , Microscopía Electrónica de Rastreo , Donantes de TejidosRESUMEN
PURPOSE: To investigate the biomechanics of the anterior human corneal stroma using atomic force microscopy (AFM). METHODS: AFM measurements were performed in liquid on the anterior stroma of human corneas, after gently removing the epithelium, using an atomic force microscope in the force spectroscopy mode. Rectangular silicon cantilevers with tip radius of 10 nm and spring elastic constants of 25- and 33-N/m were used. Each specimen was subjected to increasing loads up to a maximum of 2.7 µN with scan speeds ranging between 3- and 95-µm/s. The anterior stromal hysteresis during the extension-retraction cycle was quantified as a function of the application load and scan rate. The elastic modulus of the anterior stroma was determined by fitting force curve data to the Sneddon model. RESULTS: The anterior stroma exhibited significant viscoelasticity at micrometric level: asymmetry in the curve loading-unloading response with considerable hysteresis dependent both on the application load and scan rate (P < 0.01). The mean elastic modulus ranged between 1.14 and 2.63 MPa and was constant over the range of indentation depths between 1.0 and 2.7 µm in the stroma. CONCLUSIONS: At microscale level, the mechanical response of the most anterior stroma is complex and nonlinear. The microstructure (fibers' packing, number of cross-links, water content) and the combination of elastic (collagen fibers) and viscous (matrix) components of the tissue influence the type of viscoelastic response. Efforts in modeling the biomechanics of human corneal tissue at micrometric level are needed.
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Sustancia Propia/fisiología , Elasticidad/fisiología , Anciano , Fenómenos Biomecánicos , Recuento de Células , Sustancia Propia/citología , Módulo de Elasticidad/fisiología , Endotelio Corneal/citología , Humanos , Microscopía de Fuerza Atómica , Estrés MecánicoRESUMEN
PURPOSE: To investigate and compare the surface roughness and morphology of posterior stromal lenticules created with a femtosecond laser using various pulse energies to that obtained with a mechanical microkeratome. METHODS: A 150 kHz femtosecond laser platform (IntraLase iFS; Abbott Medical Optics) was programmed to create an 8.5-mm-diameter posterior stromal lenticule in 12 human corneal tissues. Specimens were dissected using different pulse energies (1.00, 0.75, 0.65, and 0.50) and fixed 2 µm spot separations. Three additional posterior corneal lenticules were prepared using a mechanical microkeratome (Moria Evolution 3; Moria). After the procedure, each corneal tissue was examined by atomic force microscopy (Autoprobe CP; Veeco). RESULTS: Femtosecond laser-treated tissues revealed similar morphological features, however, with significant differences in surface roughness in relation to the energy pulse used for lamellar dissection (P<0.001). The most regular stromal surface was achieved when using 0.50 µJ pulse energy; on the contrary, the roughest specimens were those dissected using 1.00 µJ pulse energy. No differences in surface roughness were measured between mechanically resected tissues and those treated using 0.50 µJ pulse energy (P>0.05). CONCLUSIONS: Atomic force microscopy submicron analysis of femtosecond-dissected donor tissues provided quantitative demonstration of the relation between pulse energy and stromal surface roughness. Surface quality of posterior corneal lenticules, comparable with that provided by mechanical microkeratome, is significantly improved when setting pulse energy for lamellar dissection of 0.50-µJ and 2-µm spot separations.
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Sustancia Propia/patología , Terapia por Láser , Microscopía de Fuerza Atómica , Anciano , Recuento de Células , Sustancia Propia/cirugía , Queratoplastia Endotelial de la Lámina Limitante Posterior , Disección/métodos , Endotelio Corneal/patología , Humanos , Propiedades de Superficie , Donantes de TejidosRESUMEN
Using electron scanning microscopy, we have studied the protein deposit left on silicon and mica substrates by dried droplets of aqueous solutions of bovine beta-lactoglobulin at low concentration and pH = 2-7. We have observed different self-assembled structures: homogeneous layers, hexagonal platelets and flower-shaped patterns laying flat on the surface, and rods formed by columns. Homogeneous layers covered the largest area of the droplet deposit. The other structures were found in small isolated regions, where the protein solution dried in the form of microdroplets. The presence of hexagonal platelets, flower-shaped patterns and columnar rods shows that beta-lactoglobulin self-assembles at the surface in a hexagonal columnar phase, which has never been observed in solution. A comparison with proteins showing similar aggregates suggests that beta-lactoglobulin structures grow from hexagonal germs composed of discotic nanometric building blocks, possibly possessing an octameric structure. We propose that discotic building blocks of beta-lactoglobulin may be produced by the anisotropic interaction with the solid surface.
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Lactoglobulinas/química , Silicatos de Aluminio , Animales , Bovinos , Concentración de Iones de Hidrógeno , Lactoglobulinas/ultraestructura , Microscopía Electrónica de Rastreo , Unión Proteica , Conformación Proteica , Silicio/químicaRESUMEN
PURPOSE: To analyze intraocular lens (IOL) optic surface adhesiveness using atomic force microscopy (AFM). SETTING: LiCryL Laboratory, University of Calabria, Rende, Italy. METHODS: The surface adhesive properties of poly(methyl methacrylate) (PMMA), silicone, hydrophilic acrylic, and hydrophobic acrylic IOLs were evaluated by AFM. Analysis was performed at room temperature (21 degrees C) in a liquid environment using the force-versus-distance mode of a commercial instrument (NanoScope III). Measurements were acquired with rectangular silicon cantilevers of a nominal elastic constant of 10 Newton/m. The nominal value of the tip's radius of curvature was 1 mum, and the scanning speed during the acquisitions ranged from 10 to 400 nm/s. RESULTS: The adhesion force measurements showed different characteristics for the various types of IOLs (P<.001, analysis of variance). The hydrophobic acrylic IOL had the largest mean adhesive force (283.75 nanoNewton [nN] +/- 0.14 [SD]) followed by the hydrophilic acrylic (84.76 +/- 0.94 nN), PMMA (45.77 +/- 0.47 nN), and silicone (2.10 +/- 0.01 nN) IOLs. CONCLUSIONS: The surface properties of the biomaterials used to manufacture IOLs are important because they can influence the incidence and severity of posterior capsule opacification (PCO). Although further studies are necessary to elucidate the mechanism of PCO development and the interface interactions between the IOL and capsule, the results in this study may bolster the theory of manufacturing more-adhesive materials to prevent PCO.
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Adhesividad , Lentes Intraoculares , Microscopía de Fuerza Atómica , Resinas Acrílicas , Materiales Biocompatibles , Polimetil Metacrilato , Elastómeros de SiliconaRESUMEN
PURPOSE: To analyze the surface optics of 4 currently available intraocular lenses (IOLs) with atomic force microscopy. SETTING: Licryl Laboratory, University of Calabria, Rende, Italy. METHODS: The surface roughness and topography of poly(methyl methacrylate) (PMMA), silicone, hydrophobic, and hydrophilic acrylic IOLs were evaluated with atomic force microscopy in contact mode. The analysis was performed in a liquid environment using cantilevers with a 0.01 Newtonw/meter nominal elastic constant. Measurements were made over areas of 10 microm2 on different locations of the posterior optic surface of the IOL. RESULTS: Atomic force microscopy permitted high-resolution imaging of IOL optic surface characteristics. Surface topography showed different features with respect to the lens biomaterial. The root-mean-square roughness of the IOL optic surface was significantly different between lenses of various materials (P < .001). The hydrophobic acrylic and silicone IOLs had the lowest mean surface roughness, 3.8 nm +/- 0.2 (SD) and 4.0 +/- 0.5 nm, respectively, and the 2 PMMA IOLs had the highest mean surface roughness, 6.6 +/- 0.3 nm and 7.0 +/- 0.6 nm. CONCLUSIONS: Atomic force microscopy was effective and accurate in analyzing IOL optics. The surface topography of IOLs may vary with different manufacturing processes.
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Lentes Intraoculares , Microscopía de Fuerza Atómica , Propiedades de Superficie , Resinas Acrílicas , Polimetil Metacrilato , Elastómeros de SiliconaRESUMEN
Stability of dye doped cholesteric liquid crystal laser emission from several minutes up to two hours and more was achieved by rotating the liquid crystal cell. Significant dependence of stability on surface treatment was observed.
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PURPOSE: To analyze the surface roughness of porcine corneas after excimer laser ablation with and without the smoothing procedure by means of atomic force microscopy. METHODS: Excimer laser photorefractive keratectomy (PRK) was performed on eight porcine corneas. Immediately following the procedure, smoothing was performed on four corneas using a viscous solution of 0.25% sodium hyaluronate. The corneas were examined in balanced salt solution after fixation in 2.5% glutaraldehyde solution using atomic force microscopy. Quantitative analysis of the ablated stromal surface topography was performed using the section analysis module of the atomic force microscopy software. Repeated measurements were made over small areas (< or =50 microm2) near the center of each ablation, with a vertical resolution of <1 nm. RESULTS: Images of the ablated stromal surface showed undulations and granule-like features on the ablated surface of the specimens. The specimens on which the smoothing procedure was performed (root-mean-square [RMS] rough: 0.152 +/- 0.014 microm) were more regular (P < .001) than those on which PRK alone was performed (RMS rough: 0.229 +/- 0.018 microm). CONCLUSIONS: Atomic force microscopy analysis requires a simpler preparation of the specimens with respect to that necessary for scanning electron microscopy; for this reason, atomic force microscopy techniques are more reliable for the study of biological surfaces and prove to be a feasible method to establish the differences when comparing different laser techniques. Our investigations highlight that although the laser cut of scanning-spot excimer laser systems is precise in removing even the smallest amounts of tissue, the smoothing technique may still be useful to reduce post-ablation roughness.