Europium Chelates Research Articles

A custom-made time-resolved fluoroimmunoassay for the quantitation of the host cell protein of Vero in rabies vaccine

Host cell proteins (HCPs) are the process-specific and inevitable impurities during the manufacture via a host cell, which affect the safety or efficacy of the bio-product. However, the commercial HCP enzyme-linked immunosorbent assay (ELISA) kits may not apply to specific products such as rabies vaccine from Vero cells. More advanced and process-specific assay methods are needed in the quality control of rabies vaccine throughout the whole manufacturing process. Therefore, a novel time-resolved fluoroimmunoassay (TRFIA) for the detection of process-specific HCP of Vero cells in rabies vaccine was established in this study. Liquid chromatography coupled tandem mass spectrometry (LC-MS/MS) was used during the preparation of HCP antigen. Based on a sandwich-type immunoassay format, analytes in samples were captured by one antibody coating in the wells and "sandwiched" by another antibody labeled with europium chelates. Due to the complex composition of HCP, both the capture and detected antibodies are polyclonal antibodies from the same anti-HCP antibodies pool. Multiple experiments have identified the optimal conditions to allow the valid and reliable detection of HCP in rabies vaccine. The TRFIA had a satisfactory limit of detection value (0.011 μg/ml) under optimal conditions, with the linear range from 0.0375 to 2.4 μg/ml of HCP. The coefficient variations (CVs) were all < 10%, and the recoveries were in the range of 97.00–102.42%. All the test results of Vero cell protein reference substance were included in the expected concentration, which demonstrated that the present method was available for the test of HCP in rabies vaccine. Based on these results, the novel TRFIA to detect HCP appears to be important for application in modern vaccine quality control during the whole manufacturing process.

Exploration of photophysical and biological attributes of red light emitting europium (III) chelates with β-keto acid and nitrogen donor ancillary ligands

Exploration of photophysical and biological attributes of red light emitting europium (III) chelates with β-keto acid and nitrogen donor ancillary ligands

Enhanced Red Emissions of Europium (III) Chelates in RNA–OTMA Complexes

In recent years, biopolymers have been widely used in optoelectronic devices because of their unique structure, novel optical and physical properties. In this paper, novel RNA-based photofunctional materials were fabricated by associating an RNA–OTMA (octadecyltrimethylammonium chloride) lipid complex with a luminescent Eu (III) complex. The fluorescence emission intensity, fluorescence quantum yield and fluorescence lifetime were characterized in both solution and thin film states, and the interaction between the RNA–OTMA lipid complex and Eu (III) complexes was analyzed. This study suggests that the luminescence performance of the Eu (III) complexes doped with RNA–OTMARNA–OTMA was significantly enhanced compared with those of the complex in a conventional poly (methyl methacrylate) matrix, thereby providing an important basis for the application of biomaterials in optoelectronic devices.

Optoelectronic and biological quantification of semi-conducting, crimson europium chelates with fluorinated β-keto acid and N-donor ancillary ligands

Optoelectronic and biological quantification of semi-conducting, crimson europium chelates with fluorinated β-keto acid and N-donor ancillary ligands

Three-dimensional temperature and velocity measurements in fluids using thermographic phosphor tracer particles

Many flows of technical and scientific interest are intrinsically three-dimensional. Extracting slices using planar measurement techniques allows only a limited view into the flow physics and can introduce ambiguities while investigating the extent of 3D regions. Nowadays, thanks to tremendous progress in the field of volumetric velocimetry, full 3D-3C velocity information can be gathered using tomographic PIV or PTV hence eliminating many of these ambiguities (Discetti and Coletti, 2018; Westerweel et al., 2013). However, for scalar quantities like temperature, 3D measurements remain challenging. Previous approaches for coupled 3D thermometry and velocimetry combined astigmatism PTV with encapsulated europium chelates particles (Massing et al., 2018) or tomographic PIV with thermochromic liquid crystals particles (Schiepel et al., 2021). Here we present a new technique based on solid thermographic phosphor tracer particles, which have been extensively used for planar fluid temperature and velocity measurements (Abram et al., 2018) and are applicable in a wide range of temperatures. The particles are seeded into a gas flow where their 3D positions are retrieved by triangulation from multiple views and their temperatures are derived from two-colour luminescence ratio imaging. In the following, the experimental setup and key processing steps are described before a demonstration of the concept in a turbulent heated jet is shown.

POF-Based Solar Concentrators Incorporating Dyes and Europium Chelates.

In this paper, useful models that enable time-efficient computational analyses of the performance of luminescent solar concentrators (LSCs) are developed and thoroughly described. These LSCs are based on polymer optical fibers codoped with organic dyes and/or europium chelates. The interest in such dopants lies in the availability of new dyes with higher quantum yields and in the photostability and suitable absorption and emission bands of europium chelates. Time-efficiency without compromising accuracy is especially important for the simulation of europium chelates, in which non-radiative energy transfers from the absorbing ligands to the europium ion and vice versa are so fast that the discretization in time, in the absence of some simplifying assumptions, would have to be very fine. Some available experimental results are also included for the sake of comparison.

Dual-label time-resolved fluoroimmunoassay for simultaneous measurement of human epidermal growth factor receptor 2 and human epididymis protein 4 in serum

In this study, a novel dual-label time-resolved fluoroimmunoassay (TRFIA) is described for simultaneous quantification of human epidermal growth factor receptor 2 (HER-2) and human epididymis protein 4 (HE4) in serum to screen gynecologic cancers. A double-antibody sandwich TRFIA was introduced with europium and samarium chelates to simultaneously detect the concentrations of HER-2 and HE4. Under optimal conditions, the proposed method exhibited wide linear ranges for HER-2 of 0.07–500 ng ml−1 and for HE4 of 0.32–1000 pmol l−1 with the average coefficient of variation below 10%. The specificity was satisfied through determining the other common tumor markers. The recovery rates were 94.5% and 96.6% on average for HER-2 and HE4, respectively. Good correlations were observed in clinical samples between developed method and commercial chemiluminescence immunoassay kits. The results demonstrated that dual-label TRFIA for HER-2 and HE4 was rapid and precise, and therefore could have a promising use in large sample detection for gynecological cancer screening.

Five‐, Four‐ and Three‐Dentate Europium Chelates for Anion Sensing and Their Applicability to Enzymatic Dephosphorylation Reactions

AbstractThe design of anion‐sensitive probes with sufficient sensitivity and selectivity is a demanding task in analytical sciences and chemical sensor technology. The reversible binding of anions to lanthanide centers is a possible approach for the development of molecular anion sensors, as reversibility is a prerequisite for continuous sensing and monitoring of enzymatic reactions. Some anion species lead to a strong increase of luminescence intensities and lifetimes by the replacement of luminescence quenching water molecules, though the selectivity of the luminescence response is still a major problem. We synthesized a series of positively charged pyridyl‐based multidentate europium complexes (five‐, four‐ and three dentate) including sensitizing chromophores and studied their luminescence intensity and lifetime responses to different polyphosphates, pyrophosphate, phosphate anions, and carboxyanions. The results revealed that the number and symmetry of the binding sites have a significant impact on the response. The five‐dentate complex was used for the real‐time monitoring of the activity of the ATP hydrolyzing enzyme apyrase.

One-step synthesis of europium complexes containing polyamino acids through ring-opening polymerization and their potential for biological imaging applications

Lanthanide-doped nanoprobes have received significant attentions for utilization in biological sensing and imaging due to their unique optical properties. However, only few works were focused on fabrication of europium complexes based fluorescence polymeric nanoparticles (FPNs) with excellent biocompatibility and biodegradability. In this study, we fabricated the FPNs (named as Eu(TTA)3Phen-GluEG FPNs) with encapsulated europium complexes which show low cytotoxicity, high sensitivity and deep penetration. Free amine group present on europium complexes initiated the ring-opening polymerization (ROP) of side-chain modified glutamic acid NCAs, offering a simple and effective method to prepare europium core FPNs with a uniform size distribution. Europium (III) chelates were furnished with a functional polyamino acid shell to fabricate biocompatible and biodegradable FPNs. Biological evaluation results demonstrate that such fabricated FPNs process excellent biocompatibility and dyeing performance. Therefore, we can expect that the fabrication approach will attract much research interest and effort on the preparation of biodegradable probes for various biological applications.

Facile fabrication of biodegradable lanthanide ions containing fluorescent polymeric nanoparticles: Characterization, optical properties and biological imaging

The fluorescent polymeric nanoparticles (FPNs) with encapsulated europium complexes have recently emerged as one of the most promising luminescent probes for biomedical applications owing to their unique optical properties, biocompatibility and structure designability. Significant progresses have been made for the preparation of biocompatible trivalent lanthanide ions (Ln3+) fluorescent nanoparticles, whereas, only limited attention was focused on the fabrication of europium ion containing FPNs with biodegradability, high sensitivity and deep penetration. In this work, a facile and simple method was developed to construct biodegradable and biocompatible luminescent polyurethanes (PUs) through a two-step polymerization. In this construction system, diisocyanate-terminating polyethylene glycol (NCO-PEG-NCO) was first synthesized and subsequently conjugated with dihydroxyl modified europium (III) chelates (Eu(TTA)3Phen-OH) to obtain these luminescent PUs. These PUs show amphipathic property and can assemble into core-shell FPNs in aqueous solution and exhibit high water dispersibility. Biological evaluation results suggest that Eu(TTA)3Phen-PEG2000 FPNs possess desirable biocompatibility and great potential for biological imaging applications. Therefore, we expect that the fabrication approach will inspire much research enthusiasm and effort for the preparation of biodegradable luminescent probes.

A magnetic nanoparticle-labeled immunoassay with europium and samarium for simultaneous quantification of serum pepsinogen I and II

Objective: To develop a novel immunoassay for the simultaneous determination of serum pepsinogen I (PG I) and pepsinogen II (PG II) by combining established methods of time-resolved fluoroimmunoassay (TRFIA) and magnetic nanoparticles separation.Materials and methods: This new immunoassay method was characterised by immobilising primary antibodies on the surface of magnetic particles and labelled with stable fluorescent chelates of europium and samarium.Results: Using magnetic nanoparticles, the TRFIA immunoassay exhibited broad dynamic assay ranges for PG I with detection limit of 0.33 ng/mL, while for PG II with detection limit of 0.38 ng/mL. Cross-reactivity between PGs I and II were <15. The intra- and inter-assay coefficient variations of the method were <3%, and the recoveries were in the range of 97–103% for serum samples. Bland–Altman analysis of 124 serum samples showed good consistency with a commercial TRFIA kit. For PG I, the mean (95% confidence interval) difference was 0.97 (–14.3–12.3) ng/mL, whilst for PG II the difference was 0.6 (–4.4–3.2) ng/mL.Conclusions: Our data suggest that the method is feasible and could be developed into a platform for the routine clinical determination of PG I and PG II levels in human serum.

Simultaneous determination of free and total prostate‐specific antigen by a magnetic particle‐based time‐resolved fluoroimmunoassay

In this paper, a novel, sensitive and rapid method to simultaneously determine the free and total prostate-specific antigen (fPSA and tPSA) in serum by combining a time-resolved fluoroimmunoassay (TRFIA) and immunomagnetic separation was described. The new approach uses magnetic particles as an immobilization matrix and means of separation, whereas the luminescent europium (Eu) and samarium (Sm) chelates are used as probes. The proposed method was evaluated via a single-step, sandwich-type TRFIA immunoassay of tPSA and fPSA as model analytes in serum. A new one-step method to simultaneously detect fPSA and tPSA in less than 15minutes was built up with the detection limits 0.006ng/mL for fPSA and 0.05ng/mL for tPSA. The assay ranges for fPSA and tPSA were both 0.5-100ng/mL, whereas the average recovery of those two obtained by our original mode were 95.9% and 95.3%, respectively. The present new TRFIA method carrying larger reproducibility, higher recovery, and sensitive specificity was demonstrated to be widely acceptable. This simultaneous determination method containing a fast and sensitive approach can be put into an important position to screening large quantities of specimen, and be commonly applied to the clinical determination of fPSA and tPSA in human serum.

Dual-Labeled Time-Resolved Immunofluorometric Assay for the Simultaneous Quantitative Detection of Hepatitis B Virus Antigens in Human Serum.

In this paper, a novel time-resolved fluorescence immunoassay (TRFIA) is described that allows the simultaneous quantitative detection of hepatitis B virus surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in human serum to aid the diagnosis and monitoring of hepatitis B virus infection. The proposed method was developed based on a two-step sandwich immunoassay protocol in which monoclonal antibodies against HBsAg and HBeAg were co-coated in 96 microtitration wells, then tracer polyclonal antibodies against HBsAg labeled with samarium and tracer monoclonal antibodies against HBeAg labeled with europium chelates were used for detection. The detection range was 0.1-150IU/mL for HBsAg and 0.5-160 PEIU/mL for HBeAg, and the detection limits were 0.03IU/L and 0.09 PEIU/ml, respectively. The intra- and inter-assay coefficients of variation were below 8% for both virus antigens. The dilution linearity and accuracy of the assay were satisfactory. No statistically significant differences were observed in sensitivity or specificity for the serum samples between the dual-label TRFIA and a commercial single-label TRFIA. These results demonstrate that an effective, reliable and convenient HBsAg/HBeAg dual-label TRFIA was successfully developed that may be clinically applicable for blood screening to monitor the course of hepatitis B virus infection and predict treatment responses.

A Novel Europium Chelate Coated Nanosphere for Time-Resolved Fluorescence Immunoassay.

A novel europium ligand 2, 2’, 2’’, 2’’’-(4, 7-diphenyl-1, 10-phenanthroline-2, 9-diyl) bis (methylene) bis (azanetriyl) tetra acetic acid (BC-EDTA) was synthesized and characterized. It shows an emission spectrum peak at 610 nm when it is excited at 360 nm, with a large Stock shift (250 nm). It is covalently coated on the surface of a bare silica nanosphere containi free amino groups, using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-Hydroxysuccinimide. We also observed an interesting phenomenon that when BC-EDTA is labeled with a silica nanosphere, the chelate shows different excitation spectrum peaks of about 295 nm. We speculate that the carboxyl has a significant influence on its excitation spectrum. The BC-EDTA/Eu3+coated nanosphere could be used as a fluorescent probe for time-resolved fluorescence immunoassay. We labeled the antibody with the fluorescent nanosphere to develop a nanosphere based hepatitis B surface antigen as a time-resolved fluorescence immunoassay reagent, which is very easy to operate and eliminates potential contamination of Eu3+ contained in the environment. The analytical and functional sensitivities are 0.0037 μg/L and 0.08 μg/L (S/N≥2.0) respectively. The detection range is 0.08-166.67 μg/L, which is much wider than that of ELISA (0.2-5μg/L). It is comparable to the commercial dissociation-enhanced lanthanide fluoro-immunoassay system (DELFIA) reagents (0.2-145μg/L). We propose that it can fulfill clinical applications.

A magnetic nanoparticle-based time-resolved fluoroimmunoassay for determination of the cytokeratin 19 fragment in human serum.

A sensitive, rapid and novel measurement method for cytokeratin 19 fragment (CYFRA 21-1) in human serum by magnetic particle-based time-resolved fluoroimmunoassay (TRFIA) is described. Built on a sandwich-type immunoassay format, analytes in samples were captured by one monoclonal antibody coating onto the surface of magnetic beads and "sandwiched" by another monoclonal antibody labeled with europium chelates. The coefficient variations of the method were lower than 7%, and the recoveries were in the range of 90-110% for serum samples. The lower limit of quantitation of the present method for CYFRA 21-1 was 0.78 ng/ml. The correlation coefficient of CYFRA 21-1 values obtained by our novel TRFIA and CLIA was 0.980. The present novel TRFIA demonstrated high sensitivity, wider effective detection range and excellent reproducibility for determination of CYFRA 21-1 can be useful for early screening and prognosis evaluation of patients with non-small cell lung cancer.

Spectrophotometric determination and removal of unchelated europium ions from solutions containing Eu-diethylenetriaminepentaacetic acid chelate–peptide conjugates

Europium chelates conjugated with peptide ligands are routinely used as probes for conducting in vitro binding experiments. The presence of unchelated Eu ions in these formulations gives high background luminescence and can lead to poor results in binding assays. In our experience, the reported methods for purification of these probes do not achieve adequate removal of unchelated metal ions in a reliable manner. In this work, a xylenol orange-based assay for the quantification of unchelated metal ions was streamlined and used to determine levels of metal ion contamination as well as the success of metal ion removal on attempted purification. We compared the use of Empore chelating disks and Chelex 100 resin for the selective removal of unchelated Eu ions from several Eu-diethylenetriaminepentaacetic acid chelate–peptide conjugates. Both purification methods gave complete and selective removal of the contaminant metal ions. However, Empore chelating disks were found to give much higher recoveries of the probes under the conditions used. Related to the issue of probe recovery, we also describe a significantly more efficient method for the synthesis of one such probe using Rink amide AM resin in place of Tentagel S resin.

A time-resolved fluoroimmunoassay for the quantitation of rabies virus nucleoprotein in the rabies vaccine

Sensitive, precise and rapid detection tests are needed in the quality control of rabies vaccine for rabies virus nucleoprotein. Previous studies for quantitation of rabies virus nucleoprotein focused on enzyme-linked immunosorbent assay (ELISA). A novel immunoassay for rapid determination of rabies virus nucleoprotein in rabies vaccine was first established by time-resolved fluoroimmunoassay (TRFIA). Based on a sandwich-type immunoassay format, analytes in samples were captured by one monoclonal antibody coating in the wells and “sandwiched” by another monoclonal antibody labeled with europium chelates. The immunocomplex was retained after washing, and then adopted treatment with enhancement solution; fluorescence was then measured according to the number of europiumions dissociated. Levels of the rabies virus nucleoprotein were measured in a linear range (5–2500mEU/mL) with a lower limit of quantitation (0.95mEU/mL) under optimal conditions. The repeatability, recovery, and linearity of the immunoassay were demonstrated to be acceptable. The correlation coefficient of nucleoprotein values obtained by novel TRFIA method and ELISA method was 0.981. These results showed good correlation and confirmed that this sensitive, precise and rapid TRFIA was feasible and could be more suitable for the quality control in the process of rabies vaccine production than ELISA.

Multidentate Europium Chelates as Luminoionophores for Anion Recognition: Impact of Ligand Design on Sensitivity and Selectivity, and Applicability to Enzymatic Assays

The design of photoluminescent molecular probes for the selective recognition of anions is a major challenge for the development of optical chemical sensors. The reversible binding of anions to lanthanide centers is one promising option for the realization of anion sensors, because it leads in some cases to a strong luminescence increase by the replacement of quenching water molecules. Yet, it is an open problem to gain control of the sensitivity and selectivity of the luminescence response. Primarily, the selective detection of (poly)phosphate species such as nucleotides has emerged as a demanding task, because they are involved in many biological processes and enzymatic reactions. We designed a series of pyridyl-based multidentate europium complexes (seven-, six-, and five-dentate) including sensitizing chromophores and studied their luminescence intensity and lifetime responses to different (poly)phosphates (adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), cyclic adenosine monophosphate (cAMP), pyrophosphate, and phosphate anions), and carboxyanions (citrate, malate, oxalacetate, succinate, α-ketoglutarate, pyruvate, oxalate, carbonate). The results reveal that the number of free coordination sites has a significant impact on the sensitivity and selectivity of the response. Because of its reversibility, the lanthanide probes can be applied to monitor the activity of ATP-consuming enzymes such ATPases and apyrases, which is demonstrated by means of the five-dentate complex.

First observation of enhanced luminescence from single lanthanide chelates on silver nanorods.

We used near-field interactions with a silver nanorod (AgNR) to greatly enhance luminescence of a lanthanide (Ln) chelate. The enhancement factor was 280-fold, making single lanthanide luminescence detectable. This is also the first observation on single molecule detection (SMD) of a lanthanide dye.

Substitutional Photoluminescence Modulation in Adducts of a Europium Chelate with a Range of Alkali Metal Cations: A Gas-Phase Study

We present gas-phase dispersed photoluminescence spectra of europium(III) 9-hydroxyphenalen-1-one (HPLN) complexes forming adducts with alkali metal ions ([Eu(PLN)3M](+) with M = Li, Na, K, Rb, and Cs) confined in a quadrupole ion trap for study. The mass selected alkali metal cation adducts display a split hypersensitive (5)D0 → (7)F2 Eu(3+) emission band. One of the two emission components shows a linear dependence on the radius of the alkali metal cation whereas the other component displays a quadratic dependence thereon. In addition, the relative intensities of both components invert in the same order. The experimental results are interpreted with the support of density functional calculations and Judd-Ofelt theory, yielding also structural information on the isolated [Eu(PLN)3M](+) chromophores.