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1.
Food Chem ; 398: 133846, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35961172

RESUMO

The unregulated usage of Cephalexin (CFX) in animal source food products has led to antimicrobial resistance (AMR) in humans. Graphene quantum dots (GQD) are zero-dimensional nanomaterials possessing both unique optical and electrical propertiesbased on their tuneable size that serves as an excellent signal enhancer. The fluorescence quenching and conductive properties of GQD were exploited for the detection of CFX. In this study, a zero-length conjugation approach was utilized to develop Cephalexin-Bovine Serum Albumin (CFX-BSA) conjugate and used to develop antibodies (Ab). Conjugated CFX-BSA Abs with GQD enhanced the electrochemical response of the sensor for sensitive detection of CFX. The fabricated electrode was optimised by Electrochemical Impedance Spectroscopy (EIS). The limit of detection for CFX was found to be 0.53 fM in standard buffer with negligible cross-reactivity against other ß-lactam antibiotics. The biofunctionalized electrode based on GQD-antibody may potentially be miniaturised for on-site detection of other antibiotics in food samples.


Assuntos
Técnicas Biossensoriais , Grafite , Pontos Quânticos , Antibacterianos , Anticorpos , Técnicas Biossensoriais/métodos , Cefalexina , Técnicas Eletroquímicas/métodos , Grafite/química , Humanos , Imunoensaio/métodos , Limite de Detecção , Pontos Quânticos/química
2.
Food Chem ; 398: 133848, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35964572

RESUMO

Aflatoxin M1 (AFM1) is an important risk factor threatening the safety of milk and dairy products due to its carcinogenic and teratogenic effects on humans. To prevent AFM1 from causing damage to human health, developing reliable methods to monitor its levels in milk and dairy products is of great importance. Biosensors built with recognition and detection systems have attracted extensive attention for their simplicity, portability, sensitivity, and selectivity. The commonly developed biosensors for detecting AFM1 are antibody-based sensors (immunosensors) and aptamer-based sensors (aptasensors). This review focused on the advances of immunosensors, aptasensors, and other emerging receptors-based sensors for the detection of AFM1 in milk and dairy products in the past five years. These biosensors were reviewed with representative examples according to their signal transduction systems, mainly including colorimetry, fluorescence, electrochemistry, and others. The unique advantages and drawbacks of immunosensor and aptasensor, and the future opportunities and challenges were also discussed.


Assuntos
Aflatoxina M1 , Técnicas Biossensoriais , Aflatoxina M1/análise , Animais , Técnicas Biossensoriais/métodos , Laticínios/análise , Eletroquímica , Contaminação de Alimentos/análise , Humanos , Imunoensaio , Leite/química
3.
Food Chem ; 398: 133861, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35973297

RESUMO

Bisacodyl, sodium picosulfate and their metabolite bis-(p-hydroxyphenyl)-pyridyl-2-methane (BHPM) are clinically used to treat constipation. In this study, with the rational hapten design, a broad-specific and highly sensitive monoclonal antibody (mAb) was obtained with half of inhibitory concentrations of 0.16, 0.18 and 0.65 ng/mL for bisacodyl, sodium picosulfate and BHPM, respectively. Based on this mAb, a rapid and sensitive lateral flow immunochromatographic assay toward bisacodyl, sodium picosulfate and BHPM in slimming foods was developed. This method can qualitatively and quantitatively screen three stimulant laxatives with cut-off values of 3-6 ng/mL by naked eye and quantitative detection limits of 0.14-0.41 ng/mL by reader. The acceptable recoveries of spiked samples (78.00 %-120.12 %) and consistent results with liquid chromatography with tandem mass spectrometry (LC-MS/MS) in real sample detection confirmed the accuracy of the method. The established method provides a technique for multiplex, sensitive, fast, and on-site detection of three stimulant laxatives in slimming food.


Assuntos
Bisacodil , Laxantes , Anticorpos Monoclonais/química , Cromatografia de Afinidade/métodos , Cromatografia Líquida , Imunoensaio/métodos , Limite de Detecção , Espectrometria de Massas em Tandem
4.
Food Chem ; 398: 133882, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35986996

RESUMO

Herein, based on an artificial clickase-catalyzed bio-conjugation strategy, we established a sensitive fluorescent clickase-linked immunosorbent assay (FCLISA) platform using an oligonucleotide-molecular beacon (Oligo-MB) hairpin structure as a fluorescence switch for detection of food allergenic protein. Firstly, a highly stable Cu(I)-containing nanocube was prepared for usage as an artificial clickase, which could catalyze the bio-conjugation of two short oligonucleotides (i.e., Oligo-A and Oligo-B labeled by a 5'-alkyne and a 3'-azide group, respectively) through clickase-catalyzed azide/alkyne cycloaddition reaction. Subsequently, the formed long-chain oligonucleotide (Oligo-A-B) could hybridize with Oligo-MB hairpin to open hairpin structure, leading to its fluorescence turn on. By using clickase as an alternative enzymatic label in conventional ELISAs, the established FCLISA showed high sensitivity and accuracy in detection of casein, achieving a limit of detection as low as 1.5 × 10-8 g/mL. Additionally, FCLISA has been challenged by detecting the casein in real samples, indicating a great potential in food safety assay.


Assuntos
Azidas , Química Click , Alcinos/química , Alérgenos , Azidas/química , Caseínas , Cobre/química , Imunoensaio , Oligonucleotídeos/química
5.
Ann Lab Med ; 43(1): 19-28, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-36045053

RESUMO

Background: Mass spectrometry methods exhibit higher accuracy and lower variability than immunoassays at low testosterone concentrations. We developed and validated an ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) assay for quantifying serum total testosterone. Methods: We used an ExionLC UPLC (Sciex, Framingham, MA, USA) system and a Sciex Triple Quad 6500+ (Sciex) MS/MS system in electrospray ionization and positive ion modes with multiple reaction monitoring transitions to evaluate precision, accuracy, linearity, lower limit of quantitation (LLOQ), carryover, ion suppression, stability, and reference intervals. For method comparison, we measured serum testosterone concentrations using this method in 40 subjects whose testosterone concentrations ranged from 0.14 to 55.48 nmol/L as determined using the Architect i2000 immunoassay (Abbott Diagnostics, Abbott Park, IL, USA) and in an additional 160 sera with testosterone concentrations <1.67 nmol/L. Results: The intra- and inter-run precision CVs were <2.81%, and the accuracy bias values were <3.85%, which were all acceptable. The verified linear interval was 0.03-180.84 nmol/L; the LLOQ was 0.03 nmol/L. No significant carryover and ion suppression were observed. The testosterone in serum was stable at 4°C, at -20°C, and after three freeze-thaw cycles. The reference intervals were successfully verified. The correlation was good at testosterone concentrations of 0.14-55.48 nmol/L; however, the Architect assay showed positive percent bias at concentrations <1.67 nmol/L. Conclusions: The UPLC-MS/MS assay shows acceptable performance, with a lower LLOQ than the immunoassay. This method will enable the quantitation of low testosterone concentrations.


Assuntos
Espectrometria de Massas em Tandem , Testosterona , Cromatografia Líquida/métodos , Humanos , Imunoensaio/métodos , Valores de Referência , Reprodutibilidade dos Testes
6.
Food Chem ; 399: 133970, 2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-35998499

RESUMO

Lateral flow immunoassays (LFIAs) are routine methods for rapid foodborne pollutants screening, with detection limits that are closely associated with the label probes used. The exploitation of high performance and robust probe is highly desirable, and remains a great challenge. Herein, we reported an emerging fluorescent nanobeads i.e. carbon-dots (CD) covalently incorporated mesoporous silicon nanoparticles (CD-MSNs) for LFIAs. CD-MSNs revealed brighter fluorescence, larger particle size and more modification sites in comparison with those of single CD. After bio-functionalisation, CD-MSNs probes were introduced to construct LFIA test strips, and designed for ultrasensitive detection of aflatoxin B1 (AFB1) and Staphylococcus aureus (S. aureus), two representative foodborne pollutants, based on the competitive and sandwich models, respectively. Very competitive quantitative detection limits i.e. 0.05 ng/mL and 102 cfu/mL were correspondingly obtained. Additionally, the test strips were successfully applied to rapidly and accurately screen AFB1 and S. aureus in food samples, highlighting their practicality.


Assuntos
Poluentes Ambientais , Nanopartículas , Aflatoxina B1/análise , Carbono , Corantes Fluorescentes , Imunoensaio/métodos , Limite de Detecção , Silício , Staphylococcus aureus
7.
Food Chem ; 399: 134002, 2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-36037690

RESUMO

Herein, a competitive quenching electrochemiluminescence immunosensor towards aflatoxin B1 (AFB1) detection was constructed by in-situ forming platinum nanoparticles (PtNPs) on ECL emitter COP T4VTP6 and effective ECL signal quencher Fc-CHO/Phe. In this system, cationic covalent organic polymer COP T4VTP6 emitted stronger cathode ECL signal at 765 nm, it acted as an interesting nanoreactor to immobilize PtCl62- through electrostatic adsorption, and directly in situ catalyzed the redox reaction to produce PtNPs without adding any external reducing agent, where PtNPs not only served as the substrate for antibody immobilization, but also played the role of coreaction accelerator to catalyze the production of SO4-, significantly improving more stable ECL signal. Moreover, the Fc-CHO/Phe labeled BSA-AFB1 was used for competitive reaction. Based on the efficient sensing strategy, ECL signal increased accordingly and exhibited linear signal responses with increasing AFB1, which realized a detection limit of 4.56 fg/mL, providing a promising potential on food analysis.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Técnicas Eletroquímicas , Imunoensaio , Limite de Detecção , Medições Luminescentes , Nanotecnologia , Platina
8.
Food Chem ; 399: 133955, 2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-36041336

RESUMO

Herbicides atrazine and acetochlor are used in crop production. Because of environmental and health hazards with respective maximum contamination levels of 3 and 20 ng/mL, quantifying these herbicides is important when considering presence in foods and vegetables. We utilized two Pd@Pt nanoparticle-amplified immunoassays, a colorimetric Pd@Pt nanoparticle-linked immunosorbent assay (NLISA) and differential pulse voltammetry (DPV) dependent on catalytic activity of Pd@Pt in a dual-lateral flow immunoassay (dual-LFIA-DPV). We achieved overall recoveries of 88.5-114 % in juice, fruit, and vegetable samples for both immunoassays. The NLISA yielded limits of detection (LODs) of 0.59 and 0.31 µg/kg and the dual-LFIA-DPV 0.27 and 0.51 µg/kg for the two respective species. Results for both immunoassays were validated by high-performance liquid chromatography (HPLC), for all food and drink samples though LODs are compromised when configuring the HPLC for both species with the same chromatogram. We expect Pd@Pt-based immunoassays to prove useful in various fields.


Assuntos
Herbicidas , Nanopartículas , Frutas/química , Herbicidas/análise , Imunoensaio/métodos , Imunoadsorventes/análise , Limite de Detecção , Verduras/química
9.
Anal Chim Acta ; 1207: 339774, 2022 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-35491036

RESUMO

Coordination polymers (CPs) with tunable structures and properties have been extensively explored in a variety of fields. In this work, we demonstrated the potential of stimuli-responsive CPs as a host of integrating enzymes to construct a portable immunoassay. By employing terbium ion (Tb3+) as a metal node and guanine monophosphate (GMP) as a bridge ligand, an alkaline phosphatase (ALP)-responsive Tb/GMP CPs was fabricated, which allows amyloglucosidase (GA) to be integrated to form GA@Tb/GMP composite. Owing to the size-selectivity of Tb/GMP CPs as a host, the loaded GA was physically isolated from its substrate starch. However, Tb/GMP CPs is highly sensitive to ALP, which can hydrolyze the phosphate group of GMP to destroy the structure of Tb/GMP CPs, leading to the release of GA from GA@Tb/GMP composite. The released GA can digest starch to produce glucoses and give a measured signal by personal glucose meter (PGM). This finding leads to a PGM-based portable immunoassay for the quantitative analysis of carcinoembryonic antigen (CEA), and satisfactory results with a detection limit of 0.28 ng/mL have been achieved. The successful determination of CEA in serum samples demonstrates its potential in practical application. We believe that this work can provide a remarkable insight for the rational design of stimuli-responsive CPs for a wide of applications.


Assuntos
Antígeno Carcinoembrionário , Polímeros Responsivos a Estímulos , Fosfatase Alcalina/análise , Glucose , Imunoensaio , Polímeros/química , Amido
10.
Mikrochim Acta ; 189(10): 367, 2022 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-36056240

RESUMO

A self-assembled nanozyme of iron porphyrin mediated supramolecular modified gold nanoparticles (FpA) was fabricated to determine nitrated alpha-synuclein as the Tyr 39 residue (nT39 α-Syn) of a potential biomarker for early diagnosis of Parkinson's disease (PD). Mechanically, localized surface plasmon resonance (LSPR) and the mass effect caused by catalytic deposition of the nanozyme contributed to a cascade signal amplification strategy. The sensor allowed a signal amplification and selective nT39 α-Syn bioanalysis with a 1.34-fold enhancement by cascade amplified SPR signal and double specific recognition. The detection limit was 1.78 ng/mL in the detection range of 7-240 ng/mL. Benefiting from the excellent immunosensor, this method can distinguish healthy people and PD patients using actual samples. Overall, this strategy provides a nanozyme-based biosensing platform for the early diagnosis of PD and can be applied to detect other protein biomarkers, such as PD-L1.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Doença de Parkinson , Biomarcadores , Técnicas Biossensoriais/métodos , Ouro/química , Humanos , Imunoensaio , Nanopartículas Metálicas/química , Nitratos , Ressonância de Plasmônio de Superfície , alfa-Sinucleína
11.
J Hazard Mater ; 439: 129701, 2022 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-36104918

RESUMO

Fenitrothion (FN) residue in food is a serious threat to public health. Consequently, a sensitive, cost-effective, and convenient immunoassay for FN urgently needs to be fabricated to safeguard human health. Herein, a nanobody-alkaline phosphatase fusion protein (Nb-ALP)-based fluorescent ELISA using red emissive carbon dots (r-CDs) anchored cobalt oxyhydroxide nanosheet (CoOOH NS) composite was developed for detecting FN. Briefly, a Nb-ALP was obtained by autoinduction expression and employed as a recognition, signal transduction, and amplification element. As the fluorescence signal source, r-CDs were assembled with CoOOH NS to yield the r-CDs@CoOOH NS composite, leading to the fluorescence quenching of r-CDs via Förster resonance energy transfer (FRET). After competitive immunoreaction, the Nb-ALP bounded to the immobilized antigen can mediate the production of ascorbic acid, which can reduce the CoOOH NS to Co2+, breaking the FRET between r-CDs and CoOOH NS, accompanied by the fluorescence recovery of r-CDs. This fluorescent ELISA is highly sensitive to FN with a detection limit of 0.14 ng mL-1, which is 25-fold lower than that of conventional colorimetric ELISAs. The recovery test of food samples and the validation by GC-MS/MS further demonstrated the proposed assay was an ideal tool for detecting FN.


Assuntos
Carbono , Fenitrotion , Fosfatase Alcalina/química , Carbono/química , Cobalto , Humanos , Imunoensaio , Óxidos , Espectrometria de Massas em Tandem
12.
PLoS One ; 17(9): e0274181, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36107911

RESUMO

Quantitative measurement of SARS-CoV-2 neutralizing antibodies is highly expected to evaluate immune status, vaccine response, and antiviral therapy. The Elecsys® Anti-SARS-CoV-2 S (Elecsys® anti-S) was developed to measure anti-SARS-CoV-2 S proteins. We sought to investigate whether Elecsys® anti-S can be used to predict neutralizing activities in patients' serums using an authentic virus neutralization assay. One hundred forty-six serum samples were obtained from 59 patients with COVID-19 at multiple time points. Of the 59 patients, 44 cases were included in Group M (mild 23, moderate 21) and produced 84 samples (mild 35, moderate 49), while 15 cases were included in Group S (severe 11, critical 4) and produced 62 samples (severe 43, critical 19). The neutralization assay detected 73% positive cases, and Elecsys® anti-S and Elecsys® Anti-SARS-CoV-2 (Elecsys® anti-N) showed 72% and 66% positive cases, respectively. A linear correlation between the Elecsys® anti-S assay and the neutralization assay were highly correlated (r = 0.7253, r2 = 0.5261) than a linear correlation between the Elecsys® anti-N and neutralization assay (r = 0.5824, r2 = 0.3392). The levels of Elecsys® anti-S antibody and neutralizing activities were significantly higher in Group S than in Group M after 6 weeks from onset of symptoms (p < 0.05). Conversely, the levels of Elecsys® anti-N were comparable in both groups. Three immunosuppressed patients, including cancer patients, showed low levels of anti-S and anti-N antibodies and neutralizing activities throughout the measurement period, indicating the need for careful follow-up. Our data indicate that Elecsys® anti-S can predict the neutralization antibodies in COVID-19.


Assuntos
Anticorpos Neutralizantes , COVID-19 , Anticorpos Antivirais , Antivirais , COVID-19/diagnóstico , Humanos , Imunoensaio , Testes de Neutralização , SARS-CoV-2
13.
Sci Rep ; 12(1): 15496, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-36109569

RESUMO

Since late 2019, the outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the resultant spread of COVID-19 have given rise to a worldwide health crisis that is posing great challenges to public health and clinical treatment, in addition to serving as a formidable threat to the global economy. To obtain an effective tool to prevent and diagnose viral infections, we attempted to obtain human antibody fragments that can effectively neutralize viral infection and be utilized for rapid virus detection. To this end, several human monoclonal antibodies were isolated by bio-panning a phage-displayed human antibody library, Tomlinson I. The selected clones were demonstrated to bind to the S1 domain of the spike glycoprotein of SARS-CoV-2. Moreover, clone A7 in Fab and IgG formats were found to effectively neutralize the binding of S protein to angiotensin-converting enzyme 2 in the low nM range. In addition, this clone was successfully converted to quench-based fluorescent immunosensors (Quenchbodies) that allowed antigen detection within a few minutes, with the help of a handy fluorometer.


Assuntos
Bacteriófagos , Técnicas Biossensoriais , COVID-19 , Enzima de Conversão de Angiotensina 2 , Anticorpos Monoclonais , Anticorpos Neutralizantes , Anticorpos Antivirais , Bacteriófagos/metabolismo , COVID-19/diagnóstico , Humanos , Imunoensaio , Fragmentos de Imunoglobulinas , Imunoglobulina G , Glicoproteínas de Membrana/metabolismo , Biblioteca de Peptídeos , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Proteínas do Envelope Viral/metabolismo
14.
Anal Chem ; 94(36): 12514-12522, 2022 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-36049116

RESUMO

Owing to its simplicity, high throughput, and ultrasensitivity, single-particle collision electrochemistry (SPCE) has attracted great attention in biosensing, especially labeled SPCE. However, the low signal conversion efficiency and much interference from complex samples limit its wide application. Here, a new and robust SPCE immunosensor was proposed for ultrasensitive cardiac troponin I (cTnI) detection by combining target-driven rolling circle amplification (RCA) with magnetic beads (MBs). Antibody-modified MBs have good stability, dispersity, and magnetic response capacity in complex samples, enabling efficient capture and separation of cTnI with high specificity and anti-interference ability. The presence of cTnI could specifically drive the formation of magnetic immunocomplexes followed by triggering RCA and enzyme digestion reaction. By using Pt nanoparticles (Pt NPs)-modified ssDNA as signal probes, one cTnI molecule could induce the release of 4.5 × 104 Pt NPs for collision experiments, greatly enhancing signal conversion efficiency and detection sensitivity. Based on the integration of MBs with RCA, the SPCE immunosensor realized 0.57 fg/mL cTnI detection with a wide linear range of 1 fg/mL to 50 ng/mL. Furthermore, cTnI detection in serum samples of myocardial infarction patients was successfully performed, demonstrating great application prospect of the SPCE immunosensor in clinical diagnosis.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Técnicas Biossensoriais/métodos , Humanos , Imunoensaio , Limite de Detecção , Fenômenos Magnéticos , Nanopartículas Metálicas/química , Troponina I
15.
Nat Commun ; 13(1): 5359, 2022 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-36097164

RESUMO

Enzyme-linked immunosorbent assays (ELISAs) are a cornerstone of modern molecular detection, but the technique still faces notable challenges. One of the biggest problems is discriminating true signal generated by target molecules versus non-specific background. Here, we developed a Single-Molecule Colocalization Assay (SiMCA) that overcomes this problem by employing total internal reflection fluorescence microscopy to quantify target proteins based on the colocalization of fluorescent signal from orthogonally labeled capture and detection antibodies. By specifically counting colocalized signals, we can eliminate the effects of background produced by non-specific binding of detection antibodies. Using TNF-α, we show that SiMCA achieves a three-fold lower limit of detection compared to conventional single-color assays and exhibits consistent performance for assays performed in complex specimens such as serum and blood. Our results help define the pernicious effects of non-specific background in immunoassays and demonstrate the diagnostic gains that can be achieved by eliminating those effects.


Assuntos
Anticorpos , Proteínas , Ensaio de Imunoadsorção Enzimática/métodos , Imunoensaio/métodos , Sensibilidade e Especificidade
16.
Int J Mol Sci ; 23(17)2022 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-36077387

RESUMO

Carbohydrate antigen 199 (CA199) is a serum biomarker which has certain value and significance in the diagnosis, prognosis, treatment, and postoperative monitoring of cancer. In this study, a lateral flow immunoassay based on europium (III) polystyrene time-resolved fluorescence microspheres (TRFM-based LFIA), integrated with a portable fluorescence reader, has been successfully establish for rapid and quantitative analysis of CA199 in human serum. Briefly, time-resolved fluorescence microspheres (TRFMs) were conjugated with antibody I (Ab1) against CA199 as detection probes, and antibody II (Ab2) was coated as capture element, and a "TRFMs-Ab1-CA199-Ab2" sandwich format would form when CA199 was detected by the TRFM-based LFIA. Under the optimal parameters, the detection limit of the TRFM-based LFIA for visible quantitation with the help of an ultraviolet light was 4.125 U/mL, which was four times lower than that of LFIA based on gold nanoparticles. Additionally, the fluorescence ratio is well linearly correlated with the CA199 concentration (0.00-66.0 U/mL) and logarithmic concentration (66.0-264.0 U/mL) for quantitative detection. Serum samples from 10 healthy people and 10 liver cancer patients were tested to confirm the performances of the point-of-care application of the TRFM-based LFIA, 20.0 U/mL of CA199 in human serum was defined as the threshold for distinguishing healthy people from liver cancer patients with an accuracy of about 60%. The establishment of TRFM-based LFIA will provide a sensitive, convenient, and efficient technical support for rapid screening of CA199 in cancer diagnosis and prognosis.


Assuntos
Neoplasias Hepáticas , Nanopartículas Metálicas , Biomarcadores Tumorais , Ouro , Humanos , Imunoensaio , Limite de Detecção , Microesferas
17.
Molecules ; 27(17)2022 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-36080260

RESUMO

Low-density lipoprotein (LDL) is a cardiac biomarker identified in the pathology of cardiovascular disease (CVD). Typically, the level of LDL is calculated using the Friedewald relationship based on measured values of total cholesterol, high-density lipoproteins (HDL), and triglycerides. Unfortunately, this approach leads to some errors in calculation. Therefore, direct methods that can be used for fast and accurate detection of LDL are needed. The purpose of this study was to develop an electrochemical platform for the detection of LDL based on an antibody-ferrocene conjugate. An anti-apolipoprotein B-100 antibody labeled with ferrocene was covalently immobilized on the layer of 4-aminothiophenol (4-ATP) on the surface of gold electrodes. Upon interaction between LDL and the antibody-ferrocene conjugate, a decrease in the ferrocene redox signal registered by square wave voltammetry was observed, which depends linearly on the concentration from 0.01 ng/mL to 1.0 ng/mL. The obtained limit of detection was equal to 0.53 ng/mL. Moreover, the satisfied selectivity toward human serum albumin (HSA), HDL, and malondialdehyde-modified low-density lipoprotein (MDA-LDL) was observed. In addition, the acceptable recovery rates of LDL in human serum samples indicate the possible application of immunosensors presented in clinical diagnostics.


Assuntos
Técnicas Biossensoriais , Imunoconjugados , HDL-Colesterol , Humanos , Imunoensaio , Lipoproteínas LDL , Metalocenos , Triglicerídeos
18.
Anal Chim Acta ; 1227: 340310, 2022 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-36089320

RESUMO

In this article we describe construction of a bioreceptive interface for detection of a breast cancer biomarker carbohydrate antigen CA15-3. The conductive interface was patterned by a 2D nanomaterial MXene, to which a mixed layer containing sulfobetaine and carboxybetaine was electrochemically grafted through a diazonium moiety. Such a modified interface was then applied for covalent immobilisation of anti-CA15-3 antibody as a bioreceptive probe for detection of a breast cancer biomarker. Two different strategies were applied for final construction of an immunosensor i.e. an interface finally blocked by bovine serum albumin or an immunosensor without such modification. Finally, electrochemical reading was accomplished using a soluble redox probe Ru(NH3)63+ ion for detection of CA15-3 in a clinically relevant range up to 50 U mL-1. The results indicate that immunosensor based on non-blocked interface can be applied for biosensing using two modes of action: 1. differential pulse voltammetry (a plot of a peak current vs. analyte concentration) and 2. an electrochemical impedance spectroscopy (a plot of a charge transfer resistance vs. analyte concentration). The electrode blocked by bovine serum albumin (BSA) can be used by additional 3. mode of action: through detection of changes in the potential (a plot Epvs. c). Additionally, we reveal and explain that Ru(NH3)63+ is redox probe, which can be applied as interfacial molecular nanoscale ruler to distinguish negatively charged protein molecules present in the close proximity (≤ 6 nm) of the electrode (in our case adsorbed BSA molecules) from the negatively charged protein molecules at a larger distance (>12 nm) from the electrode (i.e. CA15-3 analyte).


Assuntos
Técnicas Biossensoriais , Neoplasias da Mama , Biomarcadores Tumorais , Técnicas Biossensoriais/métodos , Neoplasias da Mama/diagnóstico , Feminino , Humanos , Imunoensaio/métodos , Mucina-1 , Oxirredução , Compostos de Rutênio , Soroalbumina Bovina
19.
Anal Chim Acta ; 1227: 340311, 2022 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-36089321

RESUMO

Since pesticide residues in food have attracted increasing concern worldwide, it is crucial to develop rapid and sensitive analytical methods to detect pesticide residues and ensure food safety. In this work, via the biotin-streptavidin (SA) recognition system, we constructed a magnetic relaxation switching (MRS) immunosensor for sensitive detection of chlorpyrifos (CPF). With a competitive immunoassay mode, H2O2 and Fe2+ were firstly optimized as the reaction substrates. Wherein, horseradish peroxidase (HRP) acted as enzyme label to catalyze H2O2 and mediated the conversion of Fe2+/Fe3+. By introducing Fe2+/Fe3+ interconversion as MRS signal output, CPF was detected by the immunoreaction induced variations of the transverse relaxation time (ΔT2). The proposed MRS immunosensor exhibited a detection linear range from 0.01 to 1000 ng mL-1 (R2 = 0.9916), with the limit of detection (LOD) of 6 pg mL-1 (3S/M, n = 3). As a proof-of-concept, CPF was easily detected among five different pesticides at a low concentration level (0.1 ng mL-1), as well as in real samples (apple, tea, and lettuce) with recoveries of 80.70-115.30%. Besides, the sensor can realize one step of "separation and detection" towards CPF with the aid of magnetic nanoparticles, which demonstrate its promising potential for pesticide residue detection in food samples and environment.


Assuntos
Técnicas Biossensoriais , Clorpirifos , Resíduos de Praguicidas , Técnicas Biossensoriais/métodos , Peróxido de Hidrogênio , Imunoensaio/métodos , Fenômenos Magnéticos
20.
ACS Appl Mater Interfaces ; 14(36): 41640-41648, 2022 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-36047566

RESUMO

The nanostructuration of biolayers has become a paradigm for exploiting nanoscopic light-matter phenomena for biosensing, among other biomedical purposes. In this work, we present a photopatterning method to create periodic structures of biomacromolecules based on a local and periodic mild denaturation of protein biolayers mediated by UV-laser irradiation. These nanostructures are constituted by a periodic modulation of the protein activity, so they are free of topographic and compositional changes along the pattern. Herein, we introduce the approach, explore the patterning parameters, characterize the resulting structures, and assess their overall homogeneity. This UV-based patterning principle has proven to be an easy, cost-effective, and fast way to fabricate large areas of homogeneous one-dimensional protein patterns (2 min, 15 × 1.2 mm, relative standard deviation ≃ 16%). This work also investigates the implementation of these protein patterns as transducers for diffractive biosensing. Using a model immunoassay, these patterns have demonstrated negligible signal contributions from non-specific bindings and comparable experimental limits of detection in buffer media and in human serum (53 and 36 ng·mL-1 of unlabeled IgG, respectively).


Assuntos
Técnicas Biossensoriais , Nanoestruturas , Fenômenos Biofísicos , Humanos , Imunoensaio/métodos , Lasers , Nanoestruturas/química , Transdutores
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