Kanamycin In Milk Research Articles

Impedimetric Determination of Kanamycin in Milk with Aptasensor Based on Carbon Black-Oligolactide Composite.

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%.

A simple and sensitive aptasensor based on SERS for trace analysis of kanamycin in milk

This study aimed to propose a novel surface-enhanced Raman spectroscopy (SERS)-based aptasensor by using 4-mercaptobenzoic acid (4-MBA) modified gold@silver core-shell nanoparticles (Au@Ag NPs) for detecting kanamycin for the first time with outstanding sensitivity and selectivity in the real milk samples. To achieve a high SERS activity substrate, the Au@Ag NPs with uniform shapes in a diameter of 32.2 ± 3.4 nm were synthesized successfully by seed-mediated growth and then modified with the Raman reporter 4-MBA. Under the optimal conditions, this aptamer sensing system exhibited a linear range between 6.67 × 10− 10 g/mL and 2 × 10− 7 g/mL with the limit of detection (142 pg/mL) of kanamycin. Moreover, the KANA-specific aptamer endowed a good specificity to this biosensor for the negative interference from other analogues (streptomycin sulfate, gentamicin sulfate, and penicillin). Finally, the aptasensor was applied in the artificially contaminated milk (1–100 ng/mL) samples and compared with the HPLC-MS technique, with the result indicating great practicality of the aptasensor. This paper has provided a potential aptasensor for simple and rapid trace-sensing of the harmful antibiotic residue in milk.

Development of Structure-Switching Aptamers for Kanamycin Detection Based on Fluorescence Resonance Energy Transfer.

The structure-switching aptamers are designed for the simple and rapid detection of kanamycin based on the signal transduction principle of fluorescence resonance energy transfer (FRET). The structure switch is composed of kanamycin-binding aptamers and the complementary strands, respectively labeled with fluorophore and quencher, denoted as FDNA and QDNA. In the absence of kanamycin, FDNA and QDNA form the double helix structure through the complementary pairing of bases. The fluorophore and the quencher are brought into close proximity, which results in the fluorescence quenching because of the FRET mechanism. In the presence of kanamycin, the FDNA specifically bind to the target due to the high affinity of aptamers, and the QDNA are dissociated. The specific recognition between aptamers and kanamycin will obstruct the formation of structure switch and reduce the efficiency of FRET between FDNA and QDNA, thus leading to the fluorescence enhancement. Therefore, based on the structure-switching aptamers, a simple fluorescent assay for rapid detection of kanamycin was developed. Under optimal conditions, there was a good linear relationship between kanamycin concentration and the fluorescence signal recovery. The linear range of this method in milk samples was 100–600 nM with the detection limit of 13.52 nM (3σ), which is well below the maximum residue limit (MRL) of kanamycin in milk. This method shows excellent selectivity for kanamycin over the other common antibiotics. The structure-switching aptamers have been successfully applied to the detection of kanamycin spiked in milk samples with the satisfying recoveries between 101.3 and 109.1%, which is well-consistent with the results from LC-MS/MS. Due to the outstanding advantages of facile operation, rapid detection, high sensitivity, excellent specificity, and low cost, the application and extension of this strategy for rapid determination of antibiotics in food samples may greatly improve the efficiency in food safety and quality supervision.

Preparation of polyclonal antibody and development of a biotin-streptavidin-based ELISA method for detecting kanamycin in milk and honey

Kanamycin is an aminoglycoside antibiotic used increasingly in human and veterinary medicine. However, kanamycin residues in food can cause serious side effects. Here we reported the preparation of polyclonal antibody and the development of an indirect competitive biotin-streptavidin-amplified-based enzyme-linked immunosorbent assay(BA-ELISA) that can sensitively and specifically detect kanamycin residues in milk and honey. The immunogen and coating antigen were synthesized by covalently linking kanamycin to carrier proteins using the carbodiimide method. The anti-kanamycin polyclonal antibodies were obtained from immunized rabbits. The key assay parameters were investigated and optimized. The results show that under optimum conditions, the limit of detection for kanamycin is 0.07 ng/mL and the IC50 is 6.48 ng/mL. Cross-reactivity values of the antibody with four kanamycin analogues are all <1%. Trace amounts of kanamycin in milk and honey samples can be detected by this novel BA-ELISA method successfully with satisfactory recoveries of 91.0%—103.3%. The developed protocol was also validated against liquid chromatography-mass spectrometry, returning a significant correlation. These results indicate that BA-ELISA is a viable option for monitoring kanamycin residues in milk and honey.

Thioglycolic acid-CdTe quantum dots sensing and molecularly imprinted polymer based solid phase extraction for the determination of kanamycin in milk, vaccine and stream water samples

Kanamycin is an antibiotic from the aminoglycoside class used to treat a myriad of infections. Aminoglycosides enhanced the photoluminescence of the thioglycolic acid modified CdTe quantum dots (TGA-CdTe QDs) following a Langmuir binding isotherm model. Binding constants were calculated indicating strong interactions between surface of nanoparticles and aminoglycosides. For kanamycin, the equilibrium binding constant was of 5.6×107Lmol−1. Based on this, a sensitive and selective method was developed for kanamycin determination based on the enhanced photoluminescence of TGA-CdTe QDs. Solid phase extraction with a new molecularly imprinted polymer, made using kanamycin as a template, provided high selectivity towards aminoglycosides. Under adjusted conditions, the two orders of magnitude analytical curve (linearized Langmuir binding isotherm) was constructed from 1.4×10−8molL−1 (limit of quantification) to 1.6×10−6molL−1 (8.1–92.8ngmL−1) of kanamycin sulphate. The proposed procedure was tested for the determination of kanamycin in milk and yellow fever vaccine with recoveries between 86.6 and 93.7%. In addition, the kanamycin effect on TGA-CdTe QDs was also demonstrated under capillary electrophoresis regime, indicating another possibility for selective sensing.

A Label-free and Turn-on Fluorescence Strategy for Kanamycin Detection Based on the NMM/G-quadruplex Structure.

In the present work, a simple design of a "label-free" fluorescence method for kanamycin was developed and explored based on a specific DNA aptamer as a recognizer and a NMM/G-quadruplex DNA system as a reporter. The titration experiment showed a linear detection range of 0.5 to 100 nM with a low detection limit of 0.5 nM. It also showed excellent selectivity in a selectivity experiment. It was then successfully employed to detect kanamycin in milk with a excellent reliability.

Fast detection of kanamycin in milk by protamine-aptamer-gold nanotechnology

Fast detection of kanamycin in milk by protamine-aptamer-gold nanotechnology

An aptasensor with dsDNA for rapid and highly sensitive detection of kanamycin in milk

Herein, we developed an aptasensor using double-stranded DNA (dsDNA) modified with cadmium sulfide (CdS) nanoparticles and gold nanoparticles (AuNPs) on a gold electrode (GE) for kanamycin detection.

A visual application of gold nanoparticles: Simple, reliable and sensitive detection of kanamycin based on hydrogen-bonding recognition

The authors here described a visual detection strategy employing 4-amino-3-hydrazino-5-mercapto-1, 2, 4-triazole (AHMT) functionalized gold nanoparticles (AuNPs) to detect kanamycin (KA) in various samples. The AHMT with mercapto group was self-assembled onto the surface of AuNPs and the AHMT functionalized AuNPs (AHMT-AuNPs) aggregated when KA existed owing to the hydrogen-bonding interaction between KA and AHMT. As a result, the color of the AHMT-AuNPs solution changed from wine red-to-deep purple with the increasing concentration of KA. Taking advantage of the hydrogen-bonding interaction, KA could be quantitatively detected by the proposed sensor in the range of 0.005–0.1μM and 0.1–20μM, with the detection limit as low as 0.004μM which is much lower than the maximum contamination level for KA in milk defined by the European Union. Furthermore, the proposed sensor was not affected by the interference chemicals including common amine acid, antibiotics, and metal ions. The sensor was also used to detect KA from various real samples, and the results were excellent in accord with the values measured by the high performance liquid chromatography (HPLC). This sensor exhibited a promising potential for simple and real-time detection of KA in various real samples.

Fluorometric determination of the antibiotic kanamycin by aptamer-induced FRET quenching and recovery between MoS2 nanosheets and carbon dots

A rapid and sensitive aptamer-based assay is described for kanamycin, a veterinary antibiotic with neurotoxic side effects. It is based on a novel FRET pair consisting of fluorescent carbon dots and layered MoS2. This donor-acceptor pair (operated at excitation/emission wavelengths of 380/440 nm) shows fluorescence recovery efficiencies reaching 93 %. By taking advantages of aptamer-induced fluorescence quenching and recovery, kanamycin can be quantified in the of 4–25 μM concentration range, with a detection limit of 1.1 μM. The method displays good specificity and was applied to the determination of kanamycin in spiked milk where it gave recoveries ranging from 85 % to 102 %, demonstrating that the method serves as a promising tool for the rapid detection of kanamycin in milk and other animal-derived foodstuff.

Highly sensitive aptasensor based on synergetic catalysis activity of MoS2-Au-HE composite using cDNA-Au-GOD for signal amplification

Single or few-layer nanosheets of MoS2 (MoS2 nanosheets) and a composite composed of MoS2 nanosheets, Au nanoparticles (AuNPs) and hemin (HE) (denoted as MoS2-Au-HE) were prepared. The composites possessed high synergetic catalysis activity towards the electroreduction of hydrogen peroxide. Furthermore, glucose oxidase (GOD) and AuNPs were used as marker of the complementary DNA (cDNA) strand of kanamycin aptamer to prepare a conjugate (reffered as cDNA-Au-GOD) that was designed as the signal probe. Both cDNA-Au-GOD and MoS2-Au-HE were applied to fabricate aptasensor for kanamycin. MoS2-Au-HE acted as solid platform for kanamycin aptamer and signal transmitters. AuNPs were employed as the supporter of cDNA and GOD which catalyze dissolved oxygen to produce hydrogen peroxide in the presence of glucose. Then cathodic peak current of H2O2 was recorded by differential pulse voltammetry (DPV). The electrochemical reduction of H2O2 was catalyzed by MoS2-Au-HE that was modified onto the surface of a glassy carbon electrode (GCE). The cathodic peak current of H2O2 was highly linearly decreased with an increase of kanamycin concentrations from 1.0ng/L to 1.0×105ng/L, with a detection limit of 0.8ng/L. This aptasensor can be used to detect kanamycin in milk with high specificity, sensitivity and selectivity.

A selective and sensitive fluorescent aptasensor for detection of kanamycin based on catalytic recycling activity of exonuclease III and gold nanoparticles

Kanamycin is an aminoglycoside antibiotic used in human and veterinary medicine. Sensitive and selective methods to detect kanamycin residues for food safety and clinical diagnosis are of great interest. In this study a fluorescent aptasensor was designed to detect kanamycin based on exonuclease III activity (Exo III), gold nanoparticles (AuNPs) and FAM-Labeled complimentary strand of aptamer (CS).In the absence of kanamycin, aptamer binds to its CS to form a double-stranded DNA (dsDNA) with 3′-overhang end at aptamer and 3′-blunt end at CS. The formed dsDNA leaves the surface of AuNPs. Upon addition of Exo III, aptamer is recycled from dsDNA and the cycle goes on, leading to a very strong fluorescence emission. In the presence of kanamycin, aptamer binds to its target and CS remains on the surface of AuNPs, resulting in a weak fluorescence emission.The designed aptasensor showed high selectivity toward aminoglycoside antibiotics, including kanamycin and gentamicin, with a limit of detection (LOD) as low as 321pM for kanamycin.This aptasensor was successfully used to detect kanamycin in milk and serum.

A label-free electrochemical aptasensor for the detection of kanamycin in milk

Kanamycin is detected based on the conformational change of the aptamer attached to the electrode surface and the corresponding SWV current change in [Fe(CN)6]3−/4−solution.

Aptamer-based spectrophotometric detection of kanamycin in milk

Aminoglycoside antibiotics are widely used drugs. The residual antibiotics in animal-derived food are proven to be harmful to human health and therefore should be strictly controlled in the food industry. A spectrophotometric method based on a kanamycin-specific aptamer and gold nanoparticles (AuNPs) was developed for the quantitative detection of kanamycin. Two types of functionalized AuNPs were synthesized by self-assembly of thiol-modified single-stranded DNAs (ssDNAs), which were complementary to the 5′ terminal and 3′ terminal sequences of the kanamycin aptamer. As the kanamycin aptamer is mixed with these AuNPs, the AuNPs aggregate due to the hybridization of the aptamer with the complementary ssDNAs on the surface of AuNPs. However, when different concentrations of kanamycin are present, it can bind with the aptamer specifically and competitively, which leads to the disaggregation of the AuNPs aggregate, and a change in the UV-visible absorption spectrum of AuNPs. The absorbance at 527 nm is utilized to detect the concentration of kanamycin. The detection range of this method is 1–500 nM, with a detection limit of 1 nM. The developed method was then successfully employed to detect kanamycin in milk samples, and a similar response range and detection limit were obtained.

Analysis of the Kanamycin in Raw Milk Using the Suspension Array

With the monoclonal antibody against kanamycin being prepared successfully, a bead‐based indirect competitive fluorescent immunoassay was developed to detect kanamycin in milk. The fact that there was no significant cross‐reaction with other aminoglycoside antibiotics implied that the monoclonal antibody was highly specific for kanamycin. The limit of detection (LOD) and the 50% inhibition concentration (IC50) in raw milk were 3.2 ng/mL and 52.5 ng/mL, respectively. Using the method developed in this study, the kanamycin concentrations were monitored in raw milk after the intramuscular administration of kanamycin in sick cows. Compared to the conventional enzyme‐linked immunosorbent assay (ELISA), the method using the suspension array system was more sensitive. The results obtained in the present study showed a good correlation with that of the ELISA.

P-3-357 - A cranioorbital-subfrontal aproach to anteromedial skull base lesions

Single or few-layer nanosheets of MoS2 (MoS2 nanosheets) and a composite composed of MoS2 nanosheets, Au nanoparticles (AuNPs) and hemin (HE) (denoted as MoS2-Au-HE) were prepared. The composites possessed high synergetic catalysis activity towards the electroreduction of hydrogen peroxide. Furthermore, glucose oxidase (GOD) and AuNPs were used as marker of the complementary DNA (cDNA) strand of kanamycin aptamer to prepare a conjugate (reffered as cDNA-Au-GOD) that was designed as the signal probe. Both cDNA-Au-GOD and MoS2-Au-HE were applied to fabricate aptasensor for kanamycin. MoS2-Au-HE acted as solid platform for kanamycin aptamer and signal transmitters. AuNPs were employed as the supporter of cDNA and GOD which catalyze dissolved oxygen to produce hydrogen peroxide in the presence of glucose. Then cathodic peak current of H2O2 was recorded by differential pulse voltammetry (DPV). The electrochemical reduction of H2O2 was catalyzed by MoS2-Au-HE that was modified onto the surface of a glassy carbon electrode (GCE). The cathodic peak current of H2O2 was highly linearly decreased with an increase of kanamycin concentrations from 1.0 ng/L to 1.0×105 ng/L, with a detection limit of 0.8 ng/L. This aptasensor can be used to detect kanamycin in milk with high specificity, sensitivity and selectivity.