RESUMO
An innovative supramolecular architecture is reported for bienzymatic glucose biosensing based on the use of a nanohybrid made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with a Schiff base modified with two phenylboronic acid residues (SB-dBA) as platform for the site-specific immobilization of the glycoproteins glucose oxidase (GOx) and horseradish peroxidase (HRP). The analytical signal was obtained from amperometric experiments at - 0.050 V in the presence of 5.0 × 10-4 M hydroquinone as redox mediator. The concentration of GOx and HRP and the interaction time between the enzymes and the nanohybrid MWCNT-SB-dBA deposited at glassy carbon electrodes (GCEs) were optimized through a central composite design (CCD)/response surface methodology (RSM). The optimal concentrations of GOx and HRP were 3.0 mg mL-1 and 1.50 mg mL-1, respectively, while the optimum interaction time was 3.0 min. The bienzymatic biosensor presented a sensitivity of (24 ± 2) × 102 µA dL mg-1 ((44 ± 4) × 102 µA M-1), a linear range between 0.06 mg dL-1 and 21.6 mg dL-1 (3.1 µM-1.2 mM) (R2 = 0.9991), and detection and quantification limits of 0.02 mg dL-1 (1.0 µM) and 0.06 mg dL-1 (3.1 µM), respectively. The reproducibility for five sensors prepared with the same MWCNT-SB-dBA nanohybrid was 6.3%, while the reproducibility for sensors prepared with five different nanohybrids and five electrodes each was 7.9%. The GCE/MWCNT-SB-dBA/GOx-HRP was successfully used for the quantification of glucose in artificial human urine and commercial human serum samples.
Assuntos
Técnicas Biossensoriais , Ácidos Borônicos , Enzimas Imobilizadas , Glucose Oxidase , Peroxidase do Rábano Silvestre , Nanotubos de Carbono , Bases de Schiff , Nanotubos de Carbono/química , Bases de Schiff/química , Técnicas Biossensoriais/métodos , Ácidos Borônicos/química , Glucose Oxidase/química , Glucose Oxidase/metabolismo , Peroxidase do Rábano Silvestre/química , Peroxidase do Rábano Silvestre/metabolismo , Humanos , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Glucose/análise , Eletrodos , Limite de Detecção , Técnicas Eletroquímicas/métodos , Glicemia/análiseRESUMO
An innovative strategy is proposed to simultaneously exfoliate multi-walled carbon nanotubes (MWCNTs) and generate MWCNTs with immunoaffinity properties. This strategy was based on the non-covalent functionalization of MWCNTs with human immunoglobulin G (IgG) by sonicating 2.5 mg mL-1 MWCNTs in 2.0 mg mL-1 IgG for 15 min with sonicator bath. Impedimetric experiments performed at glassy carbon electrodes (GCE) modified with the resulting MWCNT-IgG nanohybrid in the presence of anti-human immunoglobulin G antibody (Anti-IgG) demonstrated that the immunoglobulin retains their biorecognition properties even after the treatment during the MWCNT functionalization. We proposed, as proof-of-concept, two model electrochemical sensors, a voltammetric one for uric acid quantification by taking advantages of the exfoliated MWCNTs electroactivity (linear range, 5.0 × 10-7 M - 5.0 × 10-6 M; detection limit, 165 nM) and an impedimetric immunosensor for the detection of Anti-IgG through the use of the bioaffinity properties of the IgG present in the nanohybrid (linear range, 5-50 µg mL-1; detection limit, 2 µg mL-1).
Assuntos
Técnicas Biossensoriais , Nanotubos de Carbono , Humanos , Técnicas Biossensoriais/métodos , Nanotubos de Carbono/química , Imunoensaio , Imunoglobulina G , EletrodosRESUMO
This work is focused on the development of a genosensor for microRNA-21 quantification using surface plasmon resonance (SPR) to transduce the hybridization event. The biosensing platform was built by self-assembling two bilayers of poly(diallyldimethylammonium chloride) (PDDA) and graphene oxide (GO) at a gold surface modified with 3-mercaptopropane sulfonate (MPS), followed by the covalent attachment of the DNA probe. GO was used in two directions, to allow the anchoring of the probe DNA and to increase the sensitivity of the biosensing event due to its field enhancer effect. The new bioanalytical platform represents an interesting alternative for the label-free biosensing of microRNA-21, with a linear range between 1.0 fM and 10 nM, a sensitivity of 5.1 ± 0.1 moM-1 and a detection limit of 0.3fM. The proposed sensing strategy was successfully used for the quantification of microRNA-21 in enriched urine samples. Graphical abstract.
Assuntos
Grafite/química , MicroRNAs/urina , Ressonância de Plasmônio de Superfície/métodos , Sondas de DNA/química , Ouro/química , Humanos , Limite de Detecção , MicroRNAs/análise , Polietilenos/química , Compostos de Amônio Quaternário/químicaRESUMO
L-Lactate is an important bioanalyte in the food industry, biotechnology, and human healthcare. In this work, we report the development of a new L-lactate electrochemical biosensor based on the use of multiwalled carbon nanotubes non-covalently functionalized with avidin (MWCNT-Av) deposited at glassy carbon electrodes (GCEs) as anchoring sites for the bioaffinity-based immobilization of a new recombinant biotinylated lactate oxidase (bLOx) produced in Escherichia coli through in vivo biotinylation. The specific binding of MWCNT-Av to bLOx was characterized by amperometry, surface plasmon resonance (SPR), and electrochemical impedance spectroscopy (EIS). The amperometric detection of L-lactate was performed at -0.100 V, with a linear range between 100 and 700 µM, a detection limit of 33 µM, and a quantification limit of 100 µM. The proposed biosensor (GCE/MWCNT-Av/bLOx) showed a reproducibility of 6.0% and it was successfully used for determining L-lactate in food and enriched serum samples.
Assuntos
Avidina , Técnicas Biossensoriais , Ácido Láctico , Oxigenases de Função Mista , Nanotubos de Carbono , Nanotubos de Carbono/química , Oxigenases de Função Mista/química , Avidina/química , Técnicas Eletroquímicas , Ressonância de Plasmônio de Superfície , Enzimas Imobilizadas/química , Escherichia coli , Biotinilação , Eletrodos , Espectroscopia Dielétrica , Limite de DetecçãoRESUMO
The COVID-19 pandemic had devastating effects throughout the world, producing a severe crisis in the health systems and in the economy of a long list of countries, even developed ones. Therefore, highly sensitive and selective analytical bioplatforms that allow the descentralized and fast detection of the severe acute respiratory síndrome coronavirus 2 (SARS-CoV-2), are extremely necessary. Since 2020, several reviews have been published, most of them focused on the different strategies to detect the SARS-CoV-2, either from RNA, viral proteins or host antibodies produced due to the presence of the virus. In this review, the most relevant biosensors for the detection of SARS-CoV-2 RNA are particularly addressed, with special emphasis on the discussion of the biorecognition layers and the different schemes for transducing the hybridization event.
Assuntos
Técnicas Biossensoriais , COVID-19 , Ácidos Nucleicos , Humanos , SARS-CoV-2 , COVID-19/diagnóstico , RNA Viral/genética , PandemiasRESUMO
We report two novel genosensors for the quantification of SARS-CoV-2 nucleic acid using glassy carbon electrodes modified with a biocapture nanoplatform made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with avidin (Av) as a support of the biotinylated-DNA probes. One of the genosensors was based on impedimetric transduction offering a non-labelled and non-amplified detection of SARS-CoV-2 nucleic acid through the increment of [Fe(CN)6]3-/4- charge transfer resistance. This biosensor presented an excellent analytical performance, with a linear range of 1.0 × 10-18 M - 1.0 × 10-11 M, a sensitivity of (5.8 ± 0.6) x 102 Ω M-1 (r2 = 0.994), detection and quantification limits of 0.33 aM and 1.0 aM, respectively; and reproducibilities of 5.4% for 1.0 × 10-15 M target using the same MWCNTs-Av-bDNAp nanoplatform, and 6.9% for 1.0 × 10-15 M target using 3 different nanoplatforms. The other genosensor was based on a sandwich hybridization scheme and amperometric transduction using the streptavidin(Strep)-biotinylated horseradish peroxidase (bHRP)/hydrogen peroxide/hydroquinone (HQ) system. This genosensor allowed an extremely sensitive quantification of the SARS-CoV-2 nucleic acid, with a linear range of 1.0 × 10-20 M - 1.0 × 10-17 M, detection limit at zM level, and a reproducibility of 11% for genosensors prepared with the same MWCNTs-Av-bDNAp1 nanoplatform. As a proof-of-concept, and considering the extremely high sensitivity, the genosensor was challenged with highly diluted samples obtained from SARS-CoV-2 RNA PCR amplification.
RESUMO
MicroRNAs (miRNAs) are non-coding regulatory RNAs that play an important role in RNA silencing and post-transcriptional gene expression regulation. Since their dysregulation has been associated with Alzheimer disease, cardiovascular diseases and different types of cancer, among others, miRNAs can be used as biomarkers for early diagnosis and prognosis of these diseases. The methods commonly used to quantify miRNAs are, in general, complex, costly, with limited application for point-of-care devices or resource-limited facilities. Electrochemical biosensors, mainly those based on nanomaterials, have emerged as a promising alternative to the conventional miRNA detection methods and have paved the way to the development of sensitive, fast, and low-cost detection systems. This review is focused on the most relevant contributions performed in the field of electrochemical miRNAs biosensors between 2017 and the beginning of 2020. The main contribution of this article is the critical discussion of the different amplification strategies and the comparative analysis between amplified and non-amplified miRNA electrochemical biosensing and between the different amplification schemes. Particular emphasis was given to the importance of the nanostructures, enzymes, labelling molecules, and special sequences of nucleic acids or analogues on the organization of the different bioanalytical platforms, the transduction of the hybridization event and the generation the analytical signal.