Electrospun composite of polypyrrole-polyamide as a micro-solid phase extraction sorbent

A micro-solid phase extraction technique was developed using a novel polypyrrole-polyamide nanofiber sheet, fabricated by electrospinning method. The applicability of the new nanofiber sheet was examined as an extracting medium to isolate malathion as a model pesticide from aqueous samples. Solvent desorption was subsequently performed in a microvial, and an aliquot of extractant was injected into gas chromatography–mass spectrometry. Various parameters affecting the electrospinning process including monomer concentration, polyamide content, applied voltage, and electrospinning time were examined. After fabricating the most suitable preparation conditions, influential parameters on the extraction and desorption processes were optimized. The developed method proved to be rather convenient and offers sufficient sensitivity and good reproducibility. The limit of detection (S/N = 3) and limit of quantification (S/N = 10) of the method under optimized conditions were 50 and 100 ng L⁻¹, respectively. The relative standard deviation at concentration level of 1 ng mL⁻¹ was 2% (n = 3). The calibration curve of analyte showed linearity in the range of 0.1–1 ng mL⁻¹ (R ² = 0.9975). The developed method was successfully applied to tap and Zayanderood river water samples, while the relative recovery percentages of 98% and 96% were obtained, respectively. The whole procedure showed to be conveniently applicable and quite easy to be manipulated.     

A sensitive liquid chromatographic-mass spectrometric method for simultaneous determination of dehydroevodiamine and limonin from Evodia rutaecarpa in rat plasma

A liquid chromatographic–mass spectrometric (LC–MS) method has been developed and validated for simultaneous determination of dehydroevodiamine and limonin from Evodia rutaecarpa in rat plasma. After addition of the internal standard, domperidone, plasma samples were extracted by liquid–liquid extraction with ethyl acetate and separated on an Apollo C₁₈ column (250 mm × 4.6 mm, 5 μm), with methanol–0.01% formic acid water (60:40, v/v) as mobile phase, within a runtime of 12.0 min. The analytes were detected without interference in the selected ion monitoring (SIM) mode with positive electrospray ionization. The linear range was 1.0–500 ng mL⁻¹ for dehydroevodiamine and 2.0–1,000 ng mL⁻¹ for limonin, with lower limits of quantitation of 1.0 and 2.0 ng mL⁻¹, respectively. Intra-day and inter-day precision were within 6.0% and 10.9%, respectively, for both analytes, and the accuracy (relative error, RE, %) was less than 4.8% and 6.5%, respectively. The validated method was successfully applied to a comparative pharmacokinetic study of dehydroevodiamine and limonin in rat plasma after oral administration of dehydroevodiamine, limonin, and an aqueous extract of Evodiae fructus. The results indicated there were obvious differences between the pharmacokinetic behavior after oral administration of an aqueous extract of Evodiae fructus compared with single substances.     

Determination of vanadium(V) with CdTe quantum dots as fluorescent probes

CdTe quantum dots (QDs) were modified with thioglycolic acid (TGA) and synthesized in aqueous medium. The optimum fluorescence intensity was found to be at pH 6.24 with a CdTe QDs concentration of 4.96 × 10⁻⁷ mol L⁻¹. The quenched fluorescence intensity of CdTe QDs is linearly proportional to V(V) concentration from 10 to 200 ng mL⁻¹ with correlation coefficient R = 0.9985. The limit of detection for V(V) was 2.07 ng mL⁻¹. The proposed method was successfully applied to the analysis of trace amounts of V(V) in water samples with recovery of 96.5–101.8%, and the results were in good agreement with those of electrothermal atomic absorption spectrometry.     

Speciation analysis of chromium in natural water samples by electrothermal atomic absorbance spectrometry after separation/preconcentration with nanometer-sized zirconium oxide immobilized on silica gel

Silica gel was prepared by the sol–gel method, modified with nanometer-sized zirconium oxide, and this material was characterized by X-ray diffraction. A micro-column packed with silica gel modified with nanometer zirconium oxide as sorbent has been developed for the quantitative separation and preconcentration of trace amounts of chromium(III) prior to their determination by electrothermal atomic absorption spectrometry. Total chromium was determined after the reduction of chromium(VI) to chromium(III) by 10% (m/v) of aqueous ascorbic acid as reducing reagent. The adsorption capacity for chromium(III) was found to be 2.36 mg g⁻¹. The detection limit for chromium(III) was 15 ng L⁻¹ with an enrichment factor of 100. The relative standard deviation was 3.2% (n = 7, c = 2.0 ng mL⁻¹).     

New catalytic ultrasound method for derivatization of 11-nor-Δ9-tetrahydrocannabinol-9-carboxylic acid in urine, with analysis by GC-MS/MS

A new procedure is described for the derivatization by silylation of 11-nor-Δ⁹-tetrahydrocannabinol-9-carboxylic acid (THCCOOH) present in urine, followed by analysis using gas chromatography–tandem mass spectrometry. A conventional procedure for derivatization of the analyte was evaluated using two types of experimental design. A 2³ factorial design considered the parameters temperature, reaction time, and the solvent/derivatization agent ratio. A central composite design (CCD) was applied to optimize the values of the significant variables. The optimum conditions were a reaction temperature of 50 °C, a reaction time of 30 min, and a BSTFA/acetone ratio of 40:20. The use of imidazole as a catalyst, together with ultrasonication, reduced the reaction time to 5 min and increased the efficiency of derivatization of THCCOOH, compared with the conventional method. The operating conditions of the tandem mass spectrometer were also optimized. The method was linear in the concentration range 1–50 ng mL⁻¹ (R ² = 0.9951). Intra- and inter-day precisions were 7.7–12.3% and 11.1–13.9%, respectively, recoveries ranged between 91 ± 8% and 101 ± 12%, accuracy (as % bias) was between –11.7% and +0.7%, and limits of detection and quantification were 0.5 and 1.0 ng mL⁻¹, respectively.     

Application of chitosan functionalized with 3,4-dihydroxy benzoic acid moiety for on-line preconcentration and determination of trace elements in water samples

Chitosan resin functionalized with 3,4-dihydroxy benzoic acid (CCTS-DHBA resin) was used as a packing material for flow injection (FI) on-line mini-column preconcentration in combination with inductively coupled plasma-atomic emission spectrometry (ICP-AES) for the determination of trace elements such as silver, bismuth, copper, gallium, indium, molybdenum, nickel, uranium, and vanadium in environmental waters. A 5-mL aliquot of sample (pH 5.5) was introduced to the minicolumn for the adsorption/preconcentration of the metal ions, and the collected analytes on the mini-column were eluted with 2 M HNO₃, and the eluates was subsequently transported via direct injection to the nebulizer of ICP-AES for quantification. The parameters affecting on the sensitivity, such as sample pH, sample flow rate, eluent concentration, and eluent flow rate, were carefully examined. Alkali and alkaline earth metal ions commonly existing in river water and seawater did not affect the analysis of metals. Under the optimum conditions, the method allowed the determination of metal ions with detection limits of 0.08 ng mL⁻¹ (Ag), 0.9 ng mL⁻¹ (Bi), 0.07 ng mL⁻¹ (Cu), 0.9 ng mL⁻¹ (Ga), 0.9 ng mL⁻¹ (In), 0.08 ng mL⁻¹ (Mo), 0.09 ng mL⁻¹ (Ni), 0.9 ng mL⁻¹ (U), and 0.08 ng mL⁻¹ (V). By using 5 mL of sample solution, the enrichment factor and collection efficiency were 8–12 fold and 96–102%, respectively, whereas the sample throughput was 7 samples/hour. The method was validated by determining metal ions in certified reference material of river water (SLRS-4) and nearshore seawater (CASS-4), and its applicability was further demonstrated to river water and seawater samples.     

Determination of marker pteridines in urine by HPLC with fluorimetric detection and second-order multivariate calibration using MCR-ALS

A liquid chromatographic method has been developed, in combination with the multivariate curve resolution-alternating least squares algorithm (MCR-ALS), for the simultaneous determination of marker pteridines in urine samples. A central composite design has been applied to optimize the factors influencing the separation (buffer concentration, buffer pH, flow rate, oven temperature, mobile-phase composition). A set of 15 calibration samples were randomly prepared, in a concentration range of 0.5–10.5 ng mL⁻¹ for neopterin, biopterin, and pterin; 4.0–8.0 ng mL⁻¹ for xanthopterin; and 0.5–4.5 ng mL⁻¹ for isoxanthopterin. The validation was carried out with fortified urine samples from healthy adults. The optimized conditions were a mobile-phase composition of 10 mM citric buffer at pH 5.44 and acetonitrile (94.5/5.5, v/v), a flow rate of 1.0 mL min⁻¹, and an oven temperature of 25 °C. The detection system consisted of a fast-scanning spectrofluorimeter, which allows obtaining of second-order data matrices containing the fluorescence intensity as a function of retention time and emission wavelength. In this work, MCR-ALS was used to cope with coeluting interferences, on account of the second-order advantage inherent to this algorithm which, in addition, is able to handle data sets deviating from trilinearity, like the high-performance liquid chromatography data analyzed in the present report. The developed approach enabled us to determine five pteridines, some of them with overlapped profiles, reducing the experimental time and reagent consumption. Ratio values for pteridines/creatinine in urine, for infected children with different pathologies, are reported in this work.     

Rapid Fluorometric Screening of Antibiotics in Seafood

Simple and rapid fluorometric screening methods have been developed based on the competitive binding between the target and an intercalating fluorophore dye to double-stranded-DNA (dsDNA). In this study, the long-wavelength fluorescente dye TOTO-3 was employed as the indicator. Compounds that interact with dsDNA will affect the binding of TOTO-3 to the nucleic acid thereby changing the fluorescence intensity. The analyte concentration is indirectly determined by the decrease in fluorescence intensity. A fiber optic fluorescence screening system was developed for rapid and convenient sample processing. Lambda DNA (48.5 kb) was chosen as a suitable sensing nucleic acid material. Detection of sulfathiazole and chloramphenicol in shrimps using this method was studied in the range of 0.5–25 ng mL⁻¹ of sulfathiazole and of 1–50 ng mL⁻¹ of chloramphenicol. Detection limits of 0.5 ng mL⁻¹ of sulfathiazole and 1 ng mL⁻¹ of chloramphenicol were achieved. This approach is useful as a routine test in the monitoring of antibiotics in the environment or aquaculture products. The easy operation and the rapid and sensitive detection make this a potential high-throughput screening method.     

Fast and selective extraction of nicotine from human plasma based on magnetic strong cation exchange resin followed by liquid chromatography-tandem mass spectrometry

In the study, a fast and selective method based on magnetic separation has been developed for the extraction of nicotine from human plasma using magnetic strong cation exchange (MSCX) resins as adsorbent. MSCX resins were prepared using hydrophobic Fe₃O₄ magnetite as magnetically susceptible component, styrene and acrylic acid as polymeric matrix components, and acetyl sulfonate as the sulfonation agent. The extraction procedure was carried out in a single step by stirring the mixture of diluted plasma sample and MSCX resins in the vortex for 5 min. Then, the resins with adsorbed nicotine were separated from the sample matrix by applying an appropriate magnetic field. Main factors affecting the extraction of nicotine such as the amount of MSCX resins, pH value of the extraction solvent, extraction time, and washing and eluting conditions were optimized. The nicotine eluted from the resins was determined by liquid chromatography–tandem mass spectrometry. The calibration curve obtained by analyzing matrix-matched standards shows excellent linear relationship (r ² = 0.9998) in the concentration range of 10–2,500 ng mL⁻¹. The limit of detection and quantification obtained are 2.9 and 9.7 ng mL⁻¹, respectively. The relative standard deviations of intra- and inter-day obtained are in the range of 1.9–6.9% and 2.5–7.8% with the recoveries ranging from 78.7% to 99.1%. The proposed method was successfully applied to determine nicotine in human plasma phlebotomized from ten male smokers. Nicotine was detectable with the contents ranging from 44.4 to 221.9 ng mL⁻¹ in five samples.     

Fast screening immunoassay of sulfonamides in commercial fish samples

An indirect competitive enzyme-linked immunosorbent assay (ELISA) was developed in plate to detect three sulfonamide residues (sulfamerazine (SMR), sulfadimetoxine (SDM), and sulfadiazine (SDZ)) in gilthead sea bream (Sparus aurata) samples. Different extraction methodologies—using methanol/water 1:1 (v/v) + ethylene diamine tetraacetic acid (EDTA) 0.5% (m/v), acetonitrile, phosphate-buffered saline (PBS) 10 mmol L⁻¹ pH 7 and acetate buffer 100 mmol L⁻¹ pH 5—and cleanup steps, based on solid-phase extraction (C₁₈, SCX, Si) or liquid extraction with hexane, were assayed. As optimum, a fast and simple method using acetonitrile was selected to extract the sulfonamide residues from the edible muscle of fish. Due to matrix effects, a standard addition calibration curve in fish extract is necessary for quantification purposes. Sulfonamide-free samples were spiked at different concentration levels (between 30 and 90 ng g⁻¹, 5–15 ng mL⁻¹ in plate) and average recoveries (n = 8), ranging from 71% to 95%, 65% to 79%, and 72% to 95%, were obtained for SMR, SDM, and SDZ, respectively. The assay detection limits for these antibiotics were lower than 100 μg kg⁻¹ (maximum residue level established by the European Union). The accuracy was evaluated by spiking blank fish extracts at different concentrations (10–40 ng mL⁻¹, 5–20 ng mL⁻¹ in plate), and the relative errors ranged between ±20%. Finally, in order to confirm the utility of the developed ELISA as a screening methodology, fish samples from different supermarkets were analyzed, and results were compared with those obtained by a validated high-performance liquid chromatography (HPLC) method. The correlation between the results obtained by both ELISA and HPLC methods is satisfactory.      

Development of a liquid chromatography-mass spectrometry method for the determination of ursolic acid in rat plasma and tissue: application to the pharmacokinetic and tissue distribution study

A fast and sensitive liquid chromatography–mass spectrometry method was developed for the determination of ursolic acid (UA) in rat plasma and tissues. Glycyrrhetinic acid was used as the internal standard (IS). Chromatographic separation was performed on a 3.5 μm Zorbax SB-C18 column (30 mm × 2.1 mm) with a mobile phase consisting of methanol and aqueous 10 mM ammonium acetate using gradient elution. Quantification was performed by selected ion monitoring with (m/z)⁻ 455 for UA and (m/z)⁻ 469 for the IS. The method was validated in the concentration range of 2.5 − 1470 ng mL⁻¹ for plasma samples and 20 − 11760 ng g⁻¹ for tissue homogenates. The intra- and inter-day assay of precision in plasma and tissues ranged from 1.6% to 7.1% and 3.7% to 9.0%, respectively, and the intra- and inter-day assay accuracy was 84.2 − 106.9% and 82.1 − 108.1%, respectively. Recoveries in plasma and tissues ranged from 83.2% to 106.2%. The limits of detections were 0.5 ng mL⁻¹ or 4.0 ng g⁻¹. The recoveries for all samples were >90%, except for liver, which indicated that ursolic acid may metabolize in liver. The main pharmacokinetic parameters obtained were T _max = 0.42 ± 0.11 h, C _max = 1.10 ± 0.31 μg mL⁻¹, AUC = 1.45 ± 0.21 μg h mL⁻¹ and K _a = 5.64 ± 1.89 h⁻¹. The concentrations of UA in rat lung, spleen, liver, heart, and cerebellum were studied for the first time. This method is validated and could be applicable to the investigation of the pharmacokinetics and tissue distribution of UA in rats.     

Dispersive liquid–liquid microextraction followed by gas chromatography–mass spectrometry for the rapid and sensitive determination of UV filters in environmental water samples

The performance of the dispersive liquid–liquid microextraction (DLLME) technique for the determination of eight UV filters and a structurally related personal care species, benzyl salicylate (BzS), in environmental water samples is evaluated. After extraction, analytes were determined by gas chromatography combined with mass spectrometry detection (GC-MS). Parameters potentially affecting the performance of the sample preparation method (sample pH, ionic strength, type and volume of dispersant and extractant solvents) were systematically investigated using both multi- and univariant optimization strategies. Under final working conditions, analytes were extracted from 10 mL water samples by addition of 1 mL of acetone (dispersant) containing 60 μL of chlorobenzene (extractant), without modifying either the pH or the ionic strength of the sample. Limits of quantification (LOQs) between 2 and 14 ng L⁻¹, inter-day variability (evaluated with relative standard deviations, RSDs) from 9% to 14% and good linearity up to concentrations of 10,000 ng L⁻¹ were obtained. Moreover, the efficiency of the extraction was scarcely affected by the type of water sample. With the only exception of 2-ethylhexyl-p-dimethylaminobenzoate (EHPABA), compounds were found in environmental water samples at concentrations between 6 ± 1 ng L⁻¹ and 26 ± 2 ng mL⁻¹.     

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometry for determination of drugs of abuse in human urine

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometric detection (SPE–CE–MS) has been used for determination of 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), codeine (COD), hydrocodeine (HCOD), and 6-acetylmorphine (6AM) in urine. The preconcentration system consists of a small capillary filled with Oasis HLB sorbent and inserted into the inlet section of the electrophoresis capillary. The SPE–CE–MS experimental conditions were optimized as follows: the sample (adjusted to pH 6.0) was loaded at 930 mbar for 60 min, elution was performed with methanol at 50 mbar for 35 s, 60 mmol L⁻¹ ammonium acetate at pH 3.8 was used as running buffer, the separation voltage was 30 kV, and the sheath liquid at a flow rate of 5.0 μL min⁻¹ was isopropanol–water 50:50 (v/v) containing 0.5% acetic acid. Analysis of urine samples spiked with the four drugs and diluted 1:1 (v/v) was studied in the linear range 0.08–10 ng mL⁻¹. Detection limits (LODs) (S/N = 3) were between 0.013 and 0.210 ng mL⁻¹. Repeatability (expressed as relative standard deviation) was below 7.2%. The method developed enables simple and effective determination of these drugs of abuse in urine samples at the levels encountered in toxicology and doping.     

Development of a colloidal gold-based lateral-flow immunoassay for the rapid simultaneous detection of clenbuterol and ractopamine in swine urine

A multianalyte lateral-flow immunochromatographic technique using colloidal gold-labeled polyclonal antibodies was developed for the rapid simultaneous detection of clenbuterol and ractopamine. The assay procedure could be accomplished within 5 min, and the results of this qualitative one-step assay were evaluated visually according to whether test lines appeared or not. When applied to the swine urines, the detection limit and the half maximal inhibitory concentration (IC₅₀) of the test strip under an optical density scanner were calculated to be 0.1 ± 0.01 ng mL⁻¹ and 0.1 ± 0.01 ng mL⁻¹, 0.56 ± 0.08 ng mL⁻¹, and 0.71 ± 0.06 ng mL⁻¹, respectively, the cut-off levels with the naked eye of 1 ng mL⁻¹ and 1 ng mL⁻¹ for clenbuterol and ractopamine were observed. Parallel analysis of swine urine samples with clenbuterol and ractopamine showed comparable results obtained from the multianalyte lateral-flow test strip and GC-MS. Therefore, the described multianalyte lateral-flow test strip can be used as a reliable, rapid, and cost-effective on-site screening technique for the simultaneous determination of clenbuterol and ractopamine residues in swine urine.      

Analysis of isoflavones and flavonoids in human urine by UHPLC

A rapid, ultra high-performance liquid chromatographic (UHPLC) method has been developed and validated for simultaneous identification and analysis of the isoflavones genistein, daidzein, glycitin, puerarin, and biochanin A, and the flavonoids (±)-catechin, (−)-epicatechin, rutin, hesperidin, neohesperidin, quercitrin, and hesperetin in human urine. Urine samples were incubated with β-glucuronidase/sulfatase. UHPLC was performed with a Hypersil Gold (50 × 2.1 mm, 1.9 μm) analytical column. Elution was with a gradient prepared from aqueous trifluoroacetic acid (0.05%) and acetonitrile. UV detection was performed at 254 and 280 nm. The calibration curves were indicative of good linearity (r ² ≥ 0.9992) in the range of interest for each analyte. LODs ranged between 15.4 and 107.0 ng mL⁻¹ and 3.9 and 20.4 ng mL⁻¹ for flavonoids and isoflavones, respectively. Intra-day and inter-day precision (C.V., %) was less than 3.9% and 3.8%, respectively, and accuracy was between 0.03% and 5.0%. Recovery was 70.35–96.58%. The method is very rapid, simple, and reliable, and suitable for pharmacokinetic analysis. It can be routinely used for simultaneous determination of these five isoflavones and seven flavonoids in human urine. The method can also be applied to studies after administration of pharmaceutical preparations containing isoflavones and flavonoids to humans.     

DNA Quantification in Nanoliter Volumes

A novel method for the determination of proteins at nanogram levels was proposed based on the decrease of resonance light scattering (RLS) signal resulting from the interaction of dibromo-o-nitrophenylfluorone (DBONPF)-sodium lauroyl glutamate (SLG) with proteins. At pH 2.97, the decrease RLS intensity was proportional to the concentration of proteins in the range of nanogram levels with 3σ detection limits being 3.4 ng mL⁻¹ for bovine serum albumin (BSA), 1.7 ng mL⁻¹ for human serum albumin (HSA), 4.1 ng mL⁻¹ for γ-globulin (γ-IgG), 4.4 ng mL⁻¹ for egg albumin, 6.2 ng mL⁻¹ for pepsin (Pep) and 3.7 ng mL⁻¹ for α-chymotrypsin (Chy). The method is no protein-to-protein variability, simple, rapid, practical and relatively free from interference from coexisting substance, as well as much more sensitive than most of the reported methods. The proposed method was successfully applied to determine total protein in human serum samples.     

Electrochemiluminescent aptamer biosensor for the determination of ochratoxin A at a gold-nanoparticles-modified gold electrode using N-(aminobutyl)-N-ethylisoluminol as a luminescent label

A highly selective electrochemiluminescent biosensor for the detection of target nephrotoxic toxin, ochratoxin A (OTA), was developed using a DNA aptamer as the recognition element and N-(4-aminobutyl)-N-ethylisoluminol (ABEI) as the signal-producing compound. The electrochemiluminescent aptamer biosensor was fabricated by immobilizing aptamer complementary DNA 1 sequence onto the surface of a gold-nanoparticle (AuNP)-modified gold electrode. ABEI-labeled aptamer DNA 2 sequence hybridized to DNA 1 and was utilized as an electrochemiluminescent probe. A decreased electrochemiluminescence (ECL) signal was generated upon aptamer recognition of the target OTA, which induced the dissociation of DNA 2 (ABEI-labeled aptamer electrochemiluminescent probe) from DNA 1 and moved it far away from the electrode surface. Under the optimal conditions, the decreased ECL intensity was proportional to an OTA concentration ranging from 0.02 to 3.0 ng mL^-1, with a detection limit of 0.007 ng mL^-1. The relative standard deviation was 3.8% at 0.2 ng mL^-1 (n = 7). The proposed method has been applied to measure OTA in naturally contaminated wheat samples and validated by an official method. This work demonstrates the combination of a highly binding aptamer with a highly sensitive ECL technique to design an electrochemiluminescent biosensor, which is a very promising approach for the determination of small-molecule toxins.     

Development of a multichannel flow-through chemiluminescence microarray chip for parallel calibration and detection of pathogenic bacteria

Pathogen detection is important for health and safety reasons. Several outbreaks all over the world have shown the need for rapid, qualitative, quantitative, and, particularly, multianalyte detection systems. Hence, a multichannel flow-through chemiluminescence microarray chip for parallel detection of pathogenic bacteria was developed. The disposable chip made of acrylonitrile–butadiene–styrene (ABS) copolymer was devised as a support for a multiplexed sandwich immunoassay. Calibration and measurement was possible in one experiment, because the developed chip contains six parallel flow-through microchannels. Polyclonal antibodies against the pathogenic bacteria Escherichia coli O157:H7, Salmonella typhimurium, and Legionella pneumophila were immobilized on the chip by microcontact printing in order to use them as specific receptors. Detection of the captured bacteria was carried out by use of specific detection antibodies labelled with biotin and horseradish peroxidase (HRP)–streptavidine conjugates. The enzyme HRP generates chemiluminescence after adding luminol and hydrogen peroxide. This signal was observed by use of a sensitive CCD camera. The limits of detection are 1.8 × 10⁴ cells mL⁻¹ for E. coli O157:H7, 7.9 × 10⁴ cells mL⁻¹ for L. pneumophila, and 2.0 × 10⁷ cells mL⁻¹ for S. typhimurium. The overall assay time for measurement and calibration is 18 min, enabling very fast analysis.      

Label-free amperometric immunobiosensor based on a gold colloid and Prussian blue nanocomposite film modified carbon ionic liquid electrode

A novel experimental methodology based on a Prussian blue (PB) and gold nanoparticles (AuNPs) modified carbon ionic liquid electrode (CILE) was developed for use in a label-free amperometric immunosensor for the sensitive detection of human immunoglobulin G (HIgG) as a model protein. The CILE was fabricated by using the ionic liquid 1-octyl-3-methylimidazolium hexafluorophosphate as binder. Controllable electrodeposition of PB on the surface of the CILE and coating with 3-aminopropyl triethylene silane (APS) formed a film with high electronic catalytic activity and large surface area for the assembly of AuNPs and further immobilization of HIgG antibody. The electrochemistry of the formed nanocomposite biofilm was investigated by electrochemical techniques including cyclic voltammetry, differential pulse voltammetry, and electrochemical impedance spectroscopy. The HIgG concentration was measured through the decrease of amperometric responses in the corresponding specific binding of antigen and antibody. The decreased differential pulse voltammetric values were proportional to the HIgG concentration in two ranges, 0.05–1.25 ng mL⁻¹ and 1.25–40 ng mL⁻¹, with a detection limit of 0.001 ng mL⁻¹ (S/N = 3). This electrochemical immunoassay combined the specificity of the immunological reaction with the sensitivity of the AuNPs, ionic liquid, and PB amplified electrochemical detection and would therefore be valuable for clinical immunoassays.     

Development of an immunoassay and a sol-gel-based immunoaffinity cleanup method for coplanar polychlorinated biphenyls from soil and sediment samples

Two polychlorinated biphenyls (PCB) enzyme linked immunosorbent assays (ELISAs) were developed using goat PCB purified immunoglobulin (IgG) antibodies (Abs). The IgGs exhibited the highest affinity toward PCB-77 (24 ng mL⁻¹) with sensitivities in the range of 6-11 ng mL⁻¹. The Abs cross-reacted with PCB-126 and the heptachlorodibenzofuran 1,2,3,4,6,7,8-HpCDF but not with PCB-169, PCB-118, Aroclor 1232, 1248, 1260 or the hexachlorodibenzofuran 2,3,4,6,7,8-HxCDF. The IgGs were also used to develop a sol-gel-based immunoaffinity purification (IAP) method for cleanup of PCB-126. Recovery efficiencies depended on the sol-gel formats; a 1:12 format resulted in the highest binding capacity. Net binding capacity ranged from 112 to 257 ng, and 90% of the analyte could be eluted with only 2 mL of ethanol. The method was also very efficient in purifying PCB-126 from spiked soil and sediment samples from contaminated sites; and eliminating matrix interferences to a degree that enabled analysis of the purified samples by ELISA. The approaches developed in the course of the study form a basis for the development of additional IAP methods for other PCBs, and their implementation in high-throughput screening programs for PCB in food, soil, and other environmental and biological samples.