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1.
Sci Rep ; 11(1): 16810, 2021 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-34413368

RESUMO

The aim of this study was to evaluate the nature and mechanisms of interaction between HIV peptide/dendrimer complexes (dendriplex) and artificial lipid membranes, such as large unilayered vesicles (LUV) and lipid monolayers in the air-water interface. Dendriplexes were combined as one of three HIV-derived peptides (Gp160, P24 and Nef) and one of two cationic phosphorus dendrimers (CPD-G3 and CPD-G4). LUVs were formed of 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) or of a mixture of DMPC and dipalmitoyl-phosphatidylglycerol (DPPG). Interactions between dendriplexes and vesicles were characterized by dynamic light scattering (DLS), fluorescence anisotropy, differential scanning calorimetry (DSC) and Langmuir-Blodgett methods. The morphology of formed systems was examined by transmission electron microscopy (TEM). The results suggest that dendriplexes interact with both hydrophobic and hydrophilic regions of lipid bilayers. The interactions between dendriplexes and negatively charged lipids (DMPC-DPPG) were stronger than those between dendriplexes and liposomes composed of zwitterionic lipids (DMPC). The former were primarily of electrostatic nature due to the positive charge of dendriplexes and the negative charge of the membrane, whereas the latter can be attributed to disturbances in the hydrophobic domain of the membrane. Obtained results provide new information about mechanisms of interaction between lipid membranes and nanocomplexes formed with HIV-derived peptides and phosphorus dendrimers. These data could be important for the choosing the appropriate antigen delivery vehicle in the new vaccines against HIV infection.

2.
Sensors (Basel) ; 21(16)2021 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-34451028

RESUMO

A multiharmonic quartz crystal microbalance (QCM) has been applied to study the viscoelastic properties of the aptamer-based sensing layers at the surface of a QCM transducer covered by neutravidin following interaction with bacteria Listeria innocua. Addition of bacteria in the concentration range 5 × 103-106 CFU/mL resulted in a decrease of resonant frequency and in an increase of dissipation. The frequency decrease has been lower than one would expect considering the dimension of the bacteria. This can be caused by lower penetration depth of the acoustics wave (approximately 120 nm) in comparison with the thickness of the bacterial layer (approximately 500 nm). Addition of E. coli at the surface of neutravidin as well as aptamer layers did not result in significant changes in frequency and dissipation. Using the Kelvin-Voight model the analysis of the viscoelastic properties of the sensing layers was performed and several parameters such as penetration depth, Γ, viscosity coefficient, η, and shear modulus, µ, were determined following various modifications of QCM transducer. The penetration depth decreased following adsorption of the neutravidin layer, which is evidence of the formation of a rigid protein structure. This value did not change significantly following adsorption of aptamers and Listeria innocua. Viscosity coefficient was higher for the neutravidin layer in comparison with the naked QCM transducer in a buffer. However, a further increase of viscosity coefficient took place following attachment of aptamers suggesting their softer structure. The interaction of Listeria innocua with the aptamer layer resulted in slight decrease of viscosity coefficient. The shearing modulus increased for the neutravidin layer and decreased following aptamer adsorption, while a slight increase of µ was observed after the addition of Listeria innocua.


Assuntos
Escherichia coli , Técnicas de Microbalança de Cristal de Quartzo , Adsorção , Listeria , Propriedades de Superfície , Viscosidade
3.
Biosensors (Basel) ; 11(6)2021 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-34073054

RESUMO

Early diagnostics of leukemia is crucial for successful therapy of this disease. Therefore, development of rapid, sensitive, and easy-to-use methods for detection of this disease is of increased interest. Biosensor technology is challenged for this purpose. This review includes a brief description of the methods used in current clinical diagnostics of leukemia and provides recent achievements in sensor technology based on immuno- and DNA aptamer-based electrochemical and acoustic biosensors. The comparative analysis of immuno- and aptamer-based sensors shows a significant advantage of DNA aptasensors over immunosensors in the detection of cancer cells. The acoustic technique is of comparable sensitivity with those based on electrochemical methods; moreover, it is label-free and provides straightforward evaluation of the signal. Several examples of sensor development are provided and discussed.


Assuntos
Técnicas Biossensoriais , Leucemia/diagnóstico , Acústica , Aptâmeros de Nucleotídeos , Técnicas Eletroquímicas , Humanos , Imunoensaio
4.
Int J Mol Sci ; 22(5)2021 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-33800897

RESUMO

Neutrophil-mediated innate host defense mechanisms include pathogen elimination through bacterial phagocytosis, which activates the 5-lipoxygenase (5-LOX) product synthesis. Here, we studied the effect of synthetic oligodeoxyribonucleotides (ODNs), which mimic the receptor-recognized sites of bacterial (CpG-ODNs) and genomic (G-rich ODNs) DNAs released from the inflammatory area, on the neutrophil functions after cell stimulation with Salmonella typhimurium. A possible mechanism for ODN recognition by Toll-like receptor 9 (TLR9) and RAGE receptor has been proposed. We found for the first time that the combination of the magic peptide LRR11 from the leucine-rich repeat (LRR) of TLR9 with the CpG-ODNs modulates the uptake and signaling from ODNs, in particular, dramatically stimulates 5-LOX pathway. Using thickness shear mode acoustic method, we confirmed the specific binding of CpG-ODNs, but not G-rich ODN, to LRR11. The RAGE receptor has been shown to play an important role in promoting ODN uptake. Thus, FPS-ZM1, a high-affinity RAGE inhibitor, suppresses the synthesis of 5-LOX products and reduces the uptake of ODNs by neutrophils; the inhibitor effect being abolished by the addition of LRR11. The results obtained revealed that the studied peptide-ODN complexes possess high biological activity and can be promising for the development of effective vaccine adjuvants and antimicrobial therapeutics.


Assuntos
Araquidonato 5-Lipoxigenase/metabolismo , Leucotrienos/biossíntese , Neutrófilos/metabolismo , Fragmentos de Peptídeos/farmacologia , Receptor Toll-Like 9/fisiologia , Benzamidas/farmacologia , Cálcio/metabolismo , Ilhas de CpG , Ativação Enzimática/efeitos dos fármacos , Humanos , Cinética , Neutrófilos/efeitos dos fármacos , Oligodesoxirribonucleotídeos/síntese química , Oligodesoxirribonucleotídeos/farmacologia , Proteínas Opsonizantes , Espécies Reativas de Oxigênio/metabolismo , Receptor para Produtos Finais de Glicação Avançada/antagonistas & inibidores , Salmonella typhimurium
5.
Biosensors (Basel) ; 11(4)2021 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-33920444

RESUMO

The determination of protease activity is very important for disease diagnosis, drug development, and quality and safety assurance for dairy products. Therefore, the development of low-cost and sensitive methods for assessing protease activity is crucial. We report two approaches for monitoring protease activity: in a volume and at surface, via colorimetric and acoustic wave-based biosensors operated in the thickness-shear mode (TSM), respectively. The TSM sensor was based on a ß-casein substrate immobilized on a piezoelectric quartz crystal transducer. After an enzymatic reaction with trypsin, it cleaved the surface-bound ß-casein, which increased the resonant frequency of the crystal. The limit of detection (LOD) was 0.48 ± 0.08 nM. A label-free colorimetric assay for trypsin detection has also been performed using ß-casein and 6-mercaptohexanol (MCH) functionalized gold nanoparticles (AuNPs/MCH-ß-casein). Due to the trypsin cleavage of ß-casein, the gold nanoparticles lost shelter, and MCH increased the attractive force between the modified AuNPs. Consequently, AuNPs aggregated, and the red shift of the absorption spectra was observed. Spectrophotometric assay enabled an LOD of 0.42 ± 0.03 nM. The Michaelis-Menten constant, KM, for reverse enzyme reaction has also been estimated by both methods. This value for the colorimetric assay (0.56 ± 0.10 nM) is lower in comparison with those for the TSM sensor (0.92 ± 0.44 nM). This is likely due to the better access of the trypsin to the ß-casein substrate at the surface of AuNPs in comparison with those at the TSM transducer.


Assuntos
Técnicas Biossensoriais , Tripsina/análise , Acústica , Caseínas , Colorimetria , Ouro , Limite de Detecção , Nanopartículas Metálicas , Espectrofotometria
6.
Food Chem ; 353: 129373, 2021 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-33730667

RESUMO

High-resolution ultrasonic spectroscopy (HR-US) was applied for precise detection of plasmin activity towards ß-casein in buffer at pH 7.8 and 37 °C. The evolution of ultrasonic velocity and ultrasonic attenuation measured at 15.5 MHz is related to the concentration of peptide bonds hydrolyzed and loss of ß-casein aggregates, respectively. The ultrasonic assay presents sensitive and direct activity-based quantification of plasmin levels in milk. The variation in plasmin concentration between HR-US and ELISA method owed to the differing detection principles. The real-time ultrasonic profiles of hydrolysis were utilized to describe the kinetic aspect of plasmin activity. The non-linear activity curve was fitted with classic and inverse Michaelis-Menten type models. Within 1-8.6 mg·mL-1 ß-casein, the Vmax and KM obtained were (6.30 ± 2.21) × 10-5 mol.kg-1·min-1 and 10.33 ± 3.50 mg·mL-1, respectively. The maximum peptide bond cleaved was 5-6 (2.7% degree of hydrolysis) achieved at 1 mg·mL-1 ß-casein.


Assuntos
Caseínas/análise , Fibrinolisina/análise , Análise Espectral/métodos , Ultrassom/métodos , Animais , Ensaio de Imunoadsorção Enzimática , Limite de Detecção , Leite/química , Proteólise , Reprodutibilidade dos Testes
7.
Biosensors (Basel) ; 11(3)2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33652946

RESUMO

Chymotrypsin is an important proteolytic enzyme in the human digestive system that cleaves milk proteins through the hydrolysis reaction, making it an interesting subject to study the activity of milk proteases. In this work, we compared detection of chymotrypsin by spectrophotometric dynamic light scattering (DLS) and quartz crystal microbalance (QCM) methods and determined the limit of chymotrypsin detection (LOD), 0.15 ± 0.01 nM for spectrophotometric, 0.67 ± 0.05 nM for DLS and 1.40 ± 0.30 nM for QCM methods, respectively. The sensors are relatively cheap and are able to detect chymotrypsin in 3035 min. While the optical detection methods are simple to implement, the QCM method is more robust for sample preparation, and allows detection of chymotrypsin in non-transparent samples. We give an overview on methods and instruments for detection of chymotrypsin and other milk proteases.


Assuntos
Técnicas Biossensoriais , Quimotripsina/análise , Acústica , Humanos , Técnicas de Microbalança de Cristal de Quartzo
8.
Methods ; 2021 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-33677061

RESUMO

Biosensing atomic force microscopy (AFM) offers the unique feature to determine the energy landscape of a bimolecular interaction at the real single molecule level. Furthermore, simultaneous and label-free mapping of molecular recognition and the determination of sample topography at the nanoscale gets possible. A prerequisite and one of the major parts in biosensing AFM are the bio-functionalized AFM tips. In the past decades, different approaches for tip functionalization have been developed. Using these functionalization strategies, several biological highly relevant interactions at the single molecule level have been explored. For the most common approach, the use of a heterobifunctional poly(ethylenglycol) crosslinker, a broad range of linkers for different chemical coupling strategies is available. Nonetheless, the time consuming functionalization protocol as well as the broad distribution of rupture length reduces the possibility of automation and may reduce the accuracy of the results. Here we present a stable and fast forward approach based on tetra-functional DNA tetrahedra. A fast functionalization and a sharp defined distribution of rupture length gets possible with low effort and high success rate. We tested the performance on the classical avidin biotin system by using tetrahedra with three disulfide legs for stable and site directed coupling to gold coated tips and a biotinylated end at the fourth vertex. A special advantage appears when working with a DNA aptamer as sensing molecule. In this case, the fourth strand can be extended by a certain DNA sequence complementary to the linkage part of an aptamer. This AFM tip functionalization protocol was applied on thrombin using DNA aptamers directed against the fibrinogen binding side of human thrombin.

9.
Langmuir ; 37(4): 1542-1550, 2021 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-33475368

RESUMO

The unavailability of effective and safe human immunodeficiency virus (HIV) vaccines incites several approaches for development of the efficient antigen/adjuvant vaccination composite. In this study, three different dendronized gold nanoparticles (AuNPs 13-15) were investigated for a complexation ability with gp160 synthetic peptides derived from an HIV envelope. It has been shown that HIV peptides interacted with nanoparticles as evident from the changes in their secondary structures, restricted the mobility of the attached fluorescence dye, and enhanced peptide helicity confirmed by the fluorescence polarization and circular dichroism results. Transmission electron microscopy visualized complexes as cloud-like structures with attached nanoparticles. AuNP 13-15 nanoparticles bind negatively charged peptides depending on the number of functional groups; the fastest saturation and peptide retardation were observed for the most dendronized nanoparticle as indicated from dynamic light scattering, laser Doppler velocimetry, and agarose gel electrophoresis experiments. Dendronized gold nanoparticles can be considered one of the potential HIV peptide-based vaccination platforms.


Assuntos
HIV-1 , Nanopartículas Metálicas , Ouro , Proteína gp160 do Envelope de HIV , Humanos , Microscopia Eletrônica de Transmissão , Peptídeos
10.
Polymers (Basel) ; 13(3)2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33494545

RESUMO

Polymer nanoparticles and nano/micromotors are novel nanostructures that are of increased interest especially in the diagnosis and therapy of cancer. These structures are modified by antibodies or nucleic acid aptamers and can recognize the cancer markers at the membrane of the cancer cells or in the intracellular side. They can serve as a cargo for targeted transport of drugs or nucleic acids in chemo- immuno- or gene therapy. The various mechanisms, such as enzyme, ultrasound, magnetic, electrical, or light, served as a driving force for nano/micromotors, allowing their transport into the cells. This review is focused on the recent achievements in the development of polymer nanoparticles and nano/micromotors modified by antibodies and nucleic acid aptamers. The methods of preparation of polymer nanoparticles, their structure and properties are provided together with those for synthesis and the application of nano/micromotors. The various mechanisms of the driving of nano/micromotors such as chemical, light, ultrasound, electric and magnetic fields are explained. The targeting drug delivery is based on the modification of nanostructures by receptors such as nucleic acid aptamers and antibodies. Special focus is therefore on the method of selection aptamers for recognition cancer markers as well as on the comparison of the properties of nucleic acid aptamers and antibodies. The methods of immobilization of aptamers at the nanoparticles and nano/micromotors are provided. Examples of applications of polymer nanoparticles and nano/micromotors in targeted delivery and in controlled drug release are presented. The future perspectives of biomimetic nanostructures in personalized nanomedicine are also discussed.

11.
Food Chem ; 337: 127759, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-32777568

RESUMO

High-resolution ultrasonic spectroscopy (HR-US) was applied for real-time monitoring of ß-casein hydrolysis by trypsin at various conditions for the first time. The technique is based on the precision measurement of hydration changes proportional to the number of peptide bond hydrolyzed. As HR-US exhibits ultrasonic transparency for most solution, the analysis did not require optical transparency like for 2,4,6-trinitrobenzenesulfonic acid (TNBS) assay. Appropriate enzymatic models were fitted with degree of hydrolysis (dh) profiles to provide kinetic and mechanistic description of proteolysis in terms of initial hydrolysis rate, r0, and rate constant of hydrolysis, kh, and enzyme inactivation, kd. Maximal r0 and dh were obtained at 45 °C and pH 8. The exponential dependence of kinetic parameters allowed determination of the activation (EA = 50.3 ± 7 kJ/mol) and deactivation (ED = 62.23 ± 3 kJ/mol) energies of hydrolysis. The ultrasonic assay provided rapid detection of trypsin activity even at sub-nanomolar concentration.


Assuntos
Caseínas/metabolismo , Ensaios Enzimáticos/métodos , Análise Espectral , Tripsina/metabolismo , Ondas Ultrassônicas , Ativação Enzimática , Hidrólise , Cinética , Proteólise , Soluções
12.
Sensors (Basel) ; 20(23)2020 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-33291498

RESUMO

Metal-organic frameworks (MOFs) offer a unique variety of properties and morphology of the structure that make it possible to extend the performance of existing and design new electrochemical biosensors. High porosity, variable size and morphology, compatibility with common components of electrochemical sensors, and easy combination with bioreceptors make MOFs very attractive for application in the assembly of electrochemical aptasensors. In this review, the progress in the synthesis and application of the MOFs in electrochemical aptasensors are considered with an emphasis on the role of the MOF materials in aptamer immobilization and signal generation. The literature information of the use of MOFs in electrochemical aptasensors is classified in accordance with the nature and role of MOFs and a signal mode. In conclusion, future trends in the application of MOFs in electrochemical aptasensors are briefly discussed.

13.
Sensors (Basel) ; 20(19)2020 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-33003538

RESUMO

This review is focused on the application of surface and volume-sensitive acoustic methods for the detection of milk proteases such as trypsin and plasmin. While trypsin is an important protein of human milk, plasmin is a protease that plays an important role in the quality of bovine, sheep and goat milks. The increased activity of plasmin can cause an extensive cleavage of ß-casein and, thus, affect the milk gelation and taste. The basic principles of surface-sensitive acoustic methods, as well as high-resolution ultrasonic spectroscopy (HR-US), are presented. The current state-of-the-art examples of the application of acoustic sensors for protease detection in real time are discussed. The application of the HR-US method for studying the kinetics of the enzyme reaction is demonstrated. The sensitivity of the acoustics biosensors and HR-US methods for protease detection are compared.


Assuntos
Caseínas , Leite/enzimologia , Peptídeo Hidrolases/metabolismo , Acústica , Animais , Bovinos , Fibrinolisina/metabolismo , Cabras , Ovinos
14.
Sensors (Basel) ; 20(17)2020 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-32839389

RESUMO

The determination of antibiotics in food is important due to their negative effect on human health related to antimicrobial resistance problem, renal toxicity, and allergic effects. We propose an impedimetric aptasensor for the determination of kanamycin A (KANA), which was assembled on the glassy carbon electrode by the deposition of carbon black in a chitosan matrix followed by carbodiimide binding of aminated aptamer mixed with oligolactide derivative of thiacalix[4]arene in a cone configuration. The assembling was monitored by cyclic voltammetry, electrochemical impedance spectroscopy, and scanning electron microscopy. In the presence of the KANA, the charge transfer resistance of the inner interface surprisingly decreased with the analyte concentration within 0.7 and 50 nM (limit of detection 0.3 nM). This was attributed to the partial shielding of the negative charge of the aptamer and of its support, a highly porous 3D structure of the surface layer caused by a macrocyclic core of the carrier. The use of electrostatic assembling in the presence of cationic polyelectrolyte decreased tenfold the detectable concentration of KANA. The aptasensor was successfully tested in the determination of KANA in spiked milk and yogurt with recoveries within 95% and 115%.

15.
Biosensors (Basel) ; 10(3)2020 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-32138172

RESUMO

This paper describes the anti-fouling capability of the novel monolayer-forming surface linker 3-(3-(trichlorosilylpropyloxy) propanoyl chloride (MEG-Cl). This compound was successfully attached to quartz crystal surfaces which are employed in an electromagnetic piezoelectric acoustic sensor (EMPAS) configuration. The MEG-Cl coated surface was both employed with Ni-NTA for the binding of recombinant proteins and for the tandem property of the avoidance of fouling from serum and milk. The MEG-Cl coated surfaces were found to provide a large degree of anti-fouling on the EMPAS device, and were comparable to previously studied MEG-OH surfaces. Importantly, the monolayer continued to provide anti-fouling capability to the biosensor following extension with Ni-NTA in place. Accordingly, this surface linker provides an attractive system for use in biosensor technology in terms of both its anti-fouling and linking properties.


Assuntos
Técnicas Biossensoriais/métodos , Sondas Moleculares/uso terapêutico , Humanos
16.
Sensors (Basel) ; 19(22)2019 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-31739501

RESUMO

DNA sensors were assembled by consecutive deposition of thiacalix[4]arenes bearing oligolactic fragments, poly(ethylene imine), and DNA onto the glassy carbon electrode. The assembling of the layers was monitored with scanning electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy. The configuration of the thiacalix[4]arene core determined self-assembling of the polymeric species to the nano/micro particles with a size of 70-350 nm. Depending on the granulation, the coatings show the accumulation of a variety of DNA quantities, charges, and internal pore volumes. These parameters were used to optimize the DNA sensors based on these coatings. Thus, doxorubicin was determined to have limits of detection of 0.01 nM (cone configuration), 0.05 nM (partial cone configuration), and 0.10 nM (1,3-alternate configuration of the macrocycle core). Substitution of native DNA with aptamer specific to aflatoxin M1 resulted in the detection of the toxin in the range of 20 to 200 ng/L (limit of detection 5 ng/L). The aptasensor was tested in spiked milk samples and showed a recovery of 80 and 85% for 20 and 50 ng/L of the aflatoxin M1, respectively.


Assuntos
Aflatoxina M1/isolamento & purificação , Técnicas Biossensoriais , DNA/química , Poliésteres/química , Aflatoxina M1/química , Animais , Aptâmeros de Nucleotídeos/química , Carbono/química , Bovinos , Espectroscopia Dielétrica , Ouro/química , Humanos , Microscopia Eletrônica de Varredura , Leite/química , Leite/microbiologia
17.
Chemphyschem ; 20(23): 3177-3180, 2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31639248

RESUMO

Herein, we report ultrasound-propelled graphene-oxide coated gold nanowire motors, functionalized with fluorescein-labeled DNA aptamers (FAM-AIB1-apt), for qualitative detection of overexpressed AIB1 oncoproteins in MCF-7 breast cancer cells. The movement of nanomotors under the ultrasound field facilitated intracellular uptake and resulted in a faster aptamer binding with the target protein and thus faster fluorescence recovery. The propulsion behavior of the aptamer functionalized nanomotors greatly enhanced the fluorescence intensity compared to static conditions. The new aptamer@nanomotor-based strategy offers considerable potential for further development of sensing methodologies towards diagnosis of breast cancer.


Assuntos
Aptâmeros de Nucleotídeos/metabolismo , Fluoresceína/química , Aptâmeros de Nucleotídeos/química , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Feminino , Ouro/química , Humanos , Células MCF-7 , Microscopia Eletrônica de Varredura , Nanotecnologia/instrumentação , Nanotecnologia/métodos , Nanofios/química , Sonicação
18.
Biosensors (Basel) ; 9(2)2019 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-31137893

RESUMO

Detection of the breast cancer cells is important for early diagnosis of the cancer. We applied thickness shear mode acoustics method (TSM) for detection of SK-BR-3 breast cancer cells using DNA aptamers specific to HER2 positive membrane receptors. The biotinylated aptamers were immobilized at the neutravidin layer chemisorbed at gold surface of TSM transducer. Addition of the cells resulted in decrease of resonant frequency, fs, and in increase of motional resistance, Rm. Using gold nanoparticles (AuNPs), modified by aptamers it was possible improving the limit of detection (LOD) that reached 550 cells/mL, while without amplification the sensitivity of the detection of SK-BR-3 cells was 1574 cells/mL. HER2 negative cell line MDA-MB-231 did not resulted in significant changes of fs. The viability studies demonstrated that cells are stable at experimental conditions used during at least 8 h. AuNPs were not toxic on the cells up to concentration of 1 µg/mL.


Assuntos
Acústica/instrumentação , Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Neoplasias da Mama/patologia , Receptor ErbB-2/análise , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Feminino , Ouro/química , Humanos , Nanopartículas Metálicas/química , Técnicas de Diagnóstico Molecular/métodos , Receptor ErbB-2/metabolismo
19.
Sensors (Basel) ; 19(3)2019 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-30678376

RESUMO

A DNA sensor has been proposed on the platform of glassy carbon electrode modified with native DNA implemented between two electropolymerized layers of polyaniline. The surface layer was assembled by consecutive stages of potentiodynamic electrolysis, DNA drop casting, and second electrolysis, which was required for capsulation of the DNA molecules and prevented their leaching into the solution. Surface layer assembling was controlled by cyclic voltammetry, electrochemical impedance spectroscopy, atomic force, and scanning electron microscopy. For doxorubicin measurement, the DNA sensor was first incubated in the Methylene blue solution that amplified signal due to DNA intercalation and competition with the doxorubicin molecules for the DNA binding sites. The charge transfer resistance of the inner layer interface decreased with the doxorubicin concentration in the range from 1.0 pM to 0.1 µM (LOD 0.6 pM). The DNA sensor was tested for the analysis of spiked artificial urine samples and showed satisfactory recovery in concentration range of 0.05⁻10 µM. The DNA sensor developed can find application in testing of antitumor drugs and some other DNA damaging factors.


Assuntos
DNA/química , Doxorrubicina/química , Técnicas Eletroquímicas/métodos , Compostos de Anilina/química , Técnicas Biossensoriais/métodos , Espectroscopia Dielétrica/métodos
20.
Chemphyschem ; 20(4): 545-554, 2019 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-30552789

RESUMO

By using the thickness shear mode acoustics method (TSM) and single-molecule force spectroscopy (SMFS) we studied the interactions between DNA aptamers (sgc8c) specific to the protein tyrosine kinase 7 (PTK7), which is localized in the membranes of leukemia lymphoblastics (MOLT-4), and lymphocyte (Jurkat) cell lines, as well with PTK7-negative U266 myeloid leukemia cells. The TSM method allowed the development of a highly sensitive, label-free biosensor for the detection leukemia cells with a limit of detection of (195±20) cells/mL. SMFS approved the high selectivity of the sgc8c aptamers to the PTK7 receptors at the cell surface and allowed determining the binding probability of the aptamers to the PTK7 receptors at different cell lines.


Assuntos
Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , DNA/química , Leucemia/diagnóstico , Acústica , Moléculas de Adesão Celular/química , Linhagem Celular Tumoral , Humanos , Leucemia/sangue , Limite de Detecção , Receptores Proteína Tirosina Quinases/química
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