Chemiluminescence Detection Method Research Articles

Development of a chemiluminescent detection method for absolute activity measurement of Taq DNA polymerase through dATP consumption

Being one of the most extensively researched and widely used thermostable DNA polymerases, Taq DNA polymerase derived from Thermus aquaticus has played pivotal roles in numerous molecular biology applications. The accurate and convenient determination of enzymatic activity is a fundamental prerequisite for both academic research and the industrial application of Taq DNA polymerase. To address the limitations of traditional methods for polymerase activity determination, such as low accuracy, complicated operation, and radioactive hazards, we have devised an absolute-quantitation method for Taq DNA polymerase based on chemiluminescence detection of the consumption of dATP (CDC-dATP). The CDC-dATP method offers several advantages, including simplicity suitable for routine laboratory use, high sensitivity (detecting as low as 0.0025 U/μL), excellent reproducibility, a broad linear range for Taq DNA polymerase (0.0025–0.1 U/μL) with a strong linear relationship (R2 > 0.99), and, significantly, absolute activity quantitation without the associated radioactive hazards. We delved into the molecular behavior of Taq DNA polymerase in the CDC-dATP method. Furthermore, we explored the extensive application potential of the CDC-dATP methodology for measuring the activity of various other enzymes involved in dATP/ATP generation or consumption, encompassing other DNA polymerases, reverse transcriptases, RNA polymerases, hydrolases, and protein kinases, covering all seven major categories of enzymes.

A Novel Flow Injection Method with Chemiluminescence Detection for the Determination of Carmoisine in Gelatin Desserts.

Carmoisine dye, a red azo food colorant commonly utilized to impart a red color to synthetic food products, is the subject of this study. Here, we present a novel reversed flow injection analysis with a chemiluminescence detection (FIA-CL) method employing a newly developed homemade flow cell to determine carmoisine dye. This developed method is based on the inhibition effect of the dye on the chemiluminescence light (CL) emission generated from a luminal-hypochlorite system, whereby the reduction in CL intensity correlates directly with the concentration of carmoisine dye. Investigations into various analytical parameters were conducted to enhance method efficiency and applicability. A linear calibration graph of 4.0 to 100.0µg mL-1 was established (R² = 0.9993), with a detection limit of LOD = 2.93µg mL-1. Subsequent application of the proposed method to analyze gelatine dessert samples yielded results in reasonable agreement with those obtained using the reported HPLC method, as evidenced by student t-test and F-test analyses.

Application of a genetically engineered macrophage cell line for evaluating cellular effects of UV/US-treated poly(ethylene terephthalate) microplastics

Microplastic (MP) pollution is a global environmental problem. To understand the biological effects of MPs on humans, it is essential to evaluate the response of human cells to model plastic particles that mimic environmental MPs in a sensitive and non-invasive manner. In this study, we investigated the preparation of poly(ethylene terephthalate) (PET) fragments with properties similar to those of environmental MPs by combining photo-oxidative degradation via ultraviolet (UV) irradiation with mechanical pulverization and hydrolysis via ultrasound (US) exposure. Combination of UV and US treatments decreased the particle size of PET fragments to 10.2 µm and increased their crystallinity and Young's modulus to 35.7 % and 0.73 GPa, respectively, while untreated PET fragments showed the particle size of 18.9 µm, the crystallinity of 33.7 %, and Young’s modulus of 0.48 GPa. In addition, an increase in negative surface potential and O/C ratio were observed for UV/US-treated PET fragments, suggesting surface oxidation via UV/US treatment. Cytokine secretion from human macrophages was evaluated by a highly sensitive inflammation evaluation system using the HiBiT-based chemiluminescence detection method developed by genome editing technology. UV/US-treated PET fragments induced a 1.4 times higher level of inflammatory cytokine secretion on inflammatory macrophages than untreated ones, suggesting that the biological responses of PET fragments could be influenced by changes in material properties via oxidation. In conclusion, UV/US treatment enables efficient preparation of model plastic particles and is expected to provide new insights into the evaluation of biological effects using human cells. (240 words)

Methyl nitrate as a byproduct in advanced water treatment systems: Liquid chromatographic determination method and cause of formation

Neutral low-molecular-weight organics such as methyl nitrate that can readily pass through reverse osmosis (RO) membranes employed in potable water reuse facilities attract interest owing to public health considerations. In this study, a novel determination method based on high-performance liquid chromatography, online photochemical conversion to peroxynitrite, and luminol chemiluminescence detection was developed for methyl nitrate measurement in treated water. The maximum photochemical conversion efficiency of methyl nitrate to peroxynitrite was found to be 6.5% using a 222-nm excimer lamp. The calibration curve for the developed method was linear between 1.0 × 10−9 and 1.0 × 10−7 M, and the limit of detection was 0.3 nM (0.03 μg/L) given an injection volume of 200 μL. The methyl nitrate concentrations in RO permeate from reclaimed wastewater and product water after subsequent treatment by a UV/H2O2 advanced oxidation process (AOP) were 2.2 and 22.5 nM (0.17 and 1.7 μg/L), respectively. UV irradiation of RO permeate in the laboratory using a low-pressure Hg lamp confirmed the formation of methyl nitrate in the permeate in the absence of H2O2 and residual chloramines. This chemiluminescent detection method for methyl nitrate will promote a greater understanding of the origin and formation of this treatment byproduct in reclaimed wastewater.

β-galactosidase activatable chemiluminescent probe for in vivo imaging and point-of care test

As a highly effective tool with a superior signal-to-noise ratio, chemiluminescent probes find extensive utility in the in vivo detection of diverse biochemical aspects, including inflammation and the tumor microenvironment. However, the widespread adoption of chemiluminescence detection is hindered by the requirement of large precision instruments, thereby limiting its application scope. Consequently, further development and improvement of the chemiluminescence detection method are imperative. This study presents a novel chemiluminescent probe that enables precise detection of β-galactosidase (β-gal) both in vitro and in vivo by modifying the specific cleavage recognition groups of β-gal to the hydroxyl group of the dioxane-based chemiluminescent scaffold. In comparative tests against traditional chemical probes, this probe exhibited an exceptional signal-to-background ratio that significantly surpasses its counterparts. To enhance detection capabilities, a customized Point-of-Care Testing (POCT) instrument specifically designed for chemiluminescence detection was developed. This compact, user-friendly, and cost-effective instrument demonstrated comparable performance to large precision instruments. Notably, the structural characteristics of the probe, utilizing the 'hydroxyl deprotection' strategy, allow for convenient, real-time, and accurate detection of diverse targets by modifying the recognition groups tailored to specific targets, in conjunction with the POCT instrument. This research introduces a pioneering approach for generating optical signal tests in the laboratory using chemiluminescent probes, thereby unlocking the potential for rapid clinical detection and self-diagnosis of certain diseases in a home setting.

Highly sensitive determination of capsaicin with tris(2,2′-bipyridyl)ruthenium(II) electrogenerated chemiluminescence

A highly sensitive tris(2,2′-bipyridyl)ruthenium(II) [Ru(bpy)32+] electrogenerated chemiluminescence (ECL) detection method for the determination of capsaicin has been developed. In this study, capsaicin was worked as a coreactant in the oxidative-reduction pathway of the Ru(bpy)32+ ECL. The oxidation of Ru(bpy)32+ together with coreactant capsaicin on a bare glassy carbon electrode at an applied potential of + 1.10 V vs. Ag/AgCl (3 M NaCl) led to a strong ECL emission. The Ru(bpy)32+ ECL emission of capsaicin was strongly dependent upon the buffer solution pH, in which a strong ECL was obtained in alkaline buffer solution. Under the optimized conditions, the present Ru(bpy)32+ ECL detection method showed a linear response to capsaicin in the linear range from 1.0 × 10-7 M to 1.0 × 10-4 M with a limit of detection of 9.4 × 10-8 M (S/N = 3). In addition, the present method can detect dihydrocapsaicin and nordihydrocapsaicin with a detection limit of 1.9 × 10-7 M and 2.2 × 10-7 M, respectively. The present ECL method shows good analytical performance for capsaicinoids in terms of linear dynamic range and detection limit compared to those obtained at other electrochemical methods even without the employment of additional modification of electrode surfaces to enhance the detection responses. In addition, no analyte preconcentration was required. The recovery tests performed for spiked capsaicin in real pepper samples were satisfactory with the recovery from 95% to 103%. To the best of our knowledge, this is the first report on the application of ECL for the quantitative determination of capsaicinoids. Since the present method is very simple, fast and sensitive, it could be applied for the determination of capsaicin in food and pharmaceuticals.

Portable and sensitive detection of cancer cells via a handheld luminometer.

Portable detection of cancer cells is important for early-stage diagnostic applications and prognosis of cancer. Herein, a simple and sensitive chemiluminescence method was proposed for portable detection of cancer cells via a handheld luminometer. It is based on the cancer cell triggered cyclic strand displacement reaction (SDR) for signal amplification. Cancer cells, CCRF-CEM, bind to their aptamer and release the trigger DNA (TDNA) of SDR. The TDNA initiates the cyclic SDR between magnetic bead modified hairpin DNA 1 (MB-H1) and HRP-tagged hairpin DNA 2 (HRP-H2), resulting in an enrichment of HRP onto the surface of magnetic beads by forming H1/H2 duplexes. HRP catalyzed luminol/H2O2/PIP solution to produce a strong chemiluminescence (CL) signal. CCRF-CEM cells were sensitively detected by combining magnetic enrichment with the signal amplification of SDR. The CL intensity showed an excellent linear relationship with the number of CCRF-CEM cells in the range of 100 to 5 × 104 cells per mL. The detection limit was as low as 85 cells per mL. Therefore, it offers a sensitive, cheap and portable method for the chemiluminescence detection of cancer cells and provides a new option for the early diagnosis of cancers.

Determination of organophosphate flame retardant tris(2-chloroethyl)phosphine based on the luminol-H2 O2 chemiluminescence system.

Organophosphorus flame retardants (OPFRs) are new types of environmental pollutants, therefore the rapid and sensitive detection of OPFRs is a very important objective. A new experimental phenomenon was found in which tris(2-chloroethyl)phosphine (TCEP), a type of OPFR, could effectively enhance the signal of the luminol-H2 O2 chemiluminescence (CL) system. Combined with the controllability of flow injection analysis, a rapid, stable, and sensitive CL method was established. The CL intensity responded linearly to the concentration of TCEP in the range 0.5-100 μg/L (R2 = 0.999) with a low detection limit of 33 ng/L. Relative standard deviation (RSD) was 2.2% (n = 7, c = 100 μg/L). Water samples were labelled and recycled with RSDs of 1.1-5.7% and recoveries of 88.7-116.1%. Based on these results, this study established a new CL detection method for the environmental pollutant TCEP.

A sensitive chemiluminescence detection approach for determination of 2,4-dinitrophenylhydrazine derivatized aldehydes using online UV irradiation – luminol CL reaction. Application to the HPLC analysis of aldehydes in oil samples

Aldehydes are toxic carbonyl compounds that are identified in various matrices surrounding us. For instance, aldehydes could be formed during the cooking and frying of foods which affects the food quality and safety. Derivatization is a must for the determination of aldehydes as they lack intrinsic chromophoric groups. 2,4-Dinitrophenyl hydrazine (DNPH) is the most used derivatizing reagent for aldehydes and the formed hydrazones could be determined by either HPLC-UV or LC-MS. However, UV detection is non-sensitive, and the MS equipment is expensive and not widely available. Thus, herein we report a smart chemiluminescence (CL) detection method for the DNPH aldehydes derivatives. These derivatives are supposed to possess photosensitization ability due to the presence of strong chromophoric structures; nitrobenzene and phenyl hydrazone. Upon their UV irradiation, singlet oxygen is found to be produced which then converts the DNPH-aldehyde derivative into hydroperoxide. Next, the hydroperoxide reacts with luminol in an alkaline medium producing a strong CL. An HPLC system with online UV irradiation and online reaction with luminol followed by CL detection was constructed and used for the determination of aldehydes after their derivatization with DNPH. The developed method showed excellent sensitivity with detection limits down to 1.5–18.5 nM. The achieved sensitivity is superior to that obtained by HPLC-UV and LC-MS detection methods for DNPH-aldehydes derivatives. Additionally, our approach is an chemiluminogenic where the DNPH reagent itself does not produce CL which is an excellent advantage. The method was applied successfully for the determination of aldehydes in canola oil samples using simple liquid-liquid extraction showing good recovery (87.0–106.0%), accuracy (87.2–106.6), and precision (RSD≤10.2%). After analysis of fresh and heated oil samples, it was demonstrated that heating of oil, even for short time, strongly elevated the level of their aldehydes' content. At last, it was found that the results of the analysis of aldehydes in oil samples using the proposed method perfectly matched those obtained by a reference LC-MS method.

Cobalt-imidazole metal-organic framework loaded with luminol for paper-based chemiluminescence detection of catechol with use of a smartphone.

Chemiluminescence (CL) reagent luminol was loaded into the porous structure of cobalt-imidazole metal-organic framework (MOF) ZIF-67 to obtain luminol-functionalized ZIF-67 (luminol@ZIF-67) with CL property. The morphology, composition, CL property, and CL mechanism of luminol@ZIF-67 were carefully investigated. The obtained luminol@ZIF-67 exhibited strong, stable, and visible CL emission that reacted with H2O2, attributed to the strong catalytic effect of ZIF-67 combined with the shortened diffusion distance between luminol and the catalytic center. The CL intensity of luminol@ZIF-67 was more than 550 times higher than that of luminol. Catechol can effectively quench the CL emission of luminol@ZIF-67 that reacted with H2O2. Then, a simple paper-based CL imaging detection method was developed for the detection of catechol by using a smartphone as a portable detector. The linear calibration curve of the developed CL assay for catechol ranged from 5 to 100mg/L with detection limit of 1.1mg/L (S/N = 3δ). The strong CL emission of luminol@ZIF-67 combined with the effective quench ability of catechol guaranteed high sensitivity of the detection method. The practical application ability of the developed CL assay was tested by the determination of catechol in tea and tap water samples, resulting in acceptable results. This work provides an effective paper-based CL detection method for catechol and enriches the species of the chemiluminescent MOF material.

Chemiluminescence Detection in the Study of Free-Radical Reactions. Part 1.

The present review, consisting of two parts, considers the application of the chemiluminescence detection method in evaluating free radical reactions in biological model systems. The first part presents a classification of experimental biological model systems. Evidence favoring the use of chemiluminescence detection in the study of free radical reactions, along with similar methods of registering electromagnetic radiation as electron paramagnetic resonance, spectrophotometry, detection of infrared radiation (IR spectrometry), and chemical methods for assessing the end products of free radical reactions, is shown. Chemiluminescence accompanying free radical reactions involving lipids has been the extensively studied reaction. These reactions are one of the key causes of cell death by either apoptosis (activation of the cytochrome c complex with cardiolipin) or ferroptosis (induced by free ferrous ions). The concept of chemiluminescence quantum yield is also discussed in this article. The second part, which is to be published in the next issue, analyzes the application of chemiluminescence detection using luminescent additives that are called activators, a.k.a. chemiluminescence enhancers, and enhance the emission through the triplet–singlet transfer of electron excitation energy from radical reaction products, followed by light emission with a high quantum yield.

Comparison of electrochemical nitric oxide detection methods with chemiluminescence for measuring nitrite concentration in food samples

Nitrite is a naturally occurring species present in various food samples and also present in our bodies as a product of nitric oxide (NO) oxidation. Considering the ubiquity of nitrite, its determination is of great importance in both biological and food samples. Herein, a very facile indirect method of nitrite determination in meat samples via selective reduction to nitric oxide (NO) is presented. The resulting gaseous product is quantified via portable and cost-effective electrochemical sensors. Both a novel laboratory prepared Pt-Nafion based NO sensor and a commercially available amperometric NO sensor are compared. Excellent correlations between the nitrite amount found in tested samples using both of the electrochemical sensors and a reference chemiluminescence method are demonstrated (r = 0.997 and r = 0.999 for Pt-Nafion based and commercially available NO-B4 electrochemical sensors, respectively, n = 12). Moreover, the slope of the linear regression curves are very close to unity for the comparison of the three systems tested. The amperometric sensors compared within this work exhibit good precision and accuracy and are shown to be an attractive alternative to the costly chemiluminescence detection method for accurately determining nitrite levels in food samples.

Determination of morphine in culinary poppy seed tea extractions using high performance liquid chromatography with chemiluminescence detection

ABSTRACTThis study assesses the morphine content of different preparations of poppy seed tea using an acidic potassium permanganate chemiluminescence detection method. Four ‘model’ teas were prepared utilizing extractants commonly described in online drug forums and compared with a ‘control’ solution containing acetic acid. High performance liquid chromatography, coupled with acidic potassium permanganate chemiluminescence detection, was able to determine that significant concentrations of morphine can be extracted from the culinary poppy seeds with all four tea extractants (between 155 µg/g and 223 µg/g). The results of this study provide a process of determining the presence of morphine in a range of different poppy seed tea extracts in a fast, efficient and selective manner. This protocol will afford better understanding of the extent of poppy seed tea extracts in the community and allow for extension of forensic practice.

Molecularly Imprinted Polymer Based Chemiluminescence Method for Detection of Nitrofurans

In this study, a molecularly imprinted polymer capable of simultaneously recognising seven nitrofurans is synthesised. The polymer particles coated the wells of a conventional 96-well microplate as the recognition element. After sample loading, the analytes were absorbed and a highly sensitive imidazole-enhanced bis(2,4,6-trichlorophenyl) oxalate–H2O2 system was added to excite light emission. After optimisation of several parameters, the chemiluminescence method was used to determine the seven nitrofurans in animal feeds. Results showed that the method achieved ultrahigh sensitivity for the seven drugs with limits of detection of 5–12pgmL−1, and one assay was finished within 10min. In addition, the polymer-coated plate could be reused five times. The recoveries from the standard fortified blank feed samples were in the range of 74.8–97.4%. From a comparison with a high performance liquid chromatography method, the molecularly imprinted polymer based chemiluminescence method could be used as a simple, rapid, sensitive, and recyclable tool to monitor the abuse of nitrofurans in animal feeds.

A chemiluminescence method for the determination of mercury(ii) ions by tuning the catalytic activity of gold nanoparticles with ethylenediamine

A turn-on and an ultrasensitive chemiluminescence detection method for mercuric ions was developed.

Chemiluminescence of Lucigenin/Riboflavin and Its Application for Selective and Sensitive Dopamine Detection.

Lucigenin-riboflavin chemiluminescence is reported for the first time. Moreover, most dopamine chemiluminescence (CL) detection methods are based on the quenching of CL by dopamine. In contrast, we find that dopamine can significantly enhance lucigenin/riboflavin CL and establish a highly sensitive turn-on method for dopamine detection based on the enhancement of lucigenin/riboflavin CL. Under the optimal conditions, the CL intensity of the lucigenin/riboflavin system increased linearly with the concentration of dopamine in the range of 0.0056-55.56 μM, and the limit of detection is 1.87 nM.

The IAGOS NO<sub><i>x</i></sub> instrument – design, operation and first results from deployment aboard passenger aircraft

Abstract. We describe the nitrogen oxide instrument designed for the autonomous operation on board passenger aircraft in the framework of the European Research Infrastructure IAGOS (In-service Aircraft for a Global Observing System). We demonstrate the performance of the instrument using data from two deployment periods aboard an A340-300 aircraft of Deutsche Lufthansa. The well-established chemiluminescence detection method is used to measure nitrogen monoxide (NO) and nitrogen oxides (NOx). NOx is measured using a photolytic converter, and nitrogen dioxide (NO2) is determined from the difference between NOx and NO. This technique allows measuring at high time resolution (4 s) and high precision in the low ppt range (NO: 2σ = 24 pptv; NOx: 2σ = 35 pptv) over different ambient temperature and ambient pressure altitude ranges (from surface pressure down to 190 hPa). The IAGOS NOx instrument is characterized for (1) calibration stability and total uncertainty, (2) humidity and chemical interferences (e.g., ozone; nitrous acid, HONO; peroxyacetyl nitrate, PAN) and (3) inter-instrumental precision. We demonstrate that the IAGOS NOx instrument is a robust, fully automated, and long-term stable instrument suitable for unattended operation on airborne platforms, which provides useful measurements for future air quality studies and emission estimates.

Chemiluminescent Detection of HIV DNA Based on Allosteric Activation of Peroxidase-Mimicking DNAzyme

Homogeneous chemiluminescent method for HIV DNA detection based on allosteric activation of peroxidase-mimicking DNAzyme (PMDNAzyme) was developed. The probes used in the assay contain PMDNAzyme fragment and the additional oligonucleotide sequence complementary to HIV DNA. The interaction of PMDNAzyme fragment and the additional oligonucleotide sequence results in changes in G-quadruplex structure of the PMDNAzyme and decreases peroxidase-like activity of the probe. In the presence of HIV DNA such interaction was destroyed due to the formation of stable duplex between the additional fragment of the probe and DNA-analyte. Consequently, some reorganizations in G-quadruplex structure of the probe are observed, which are accompanied by enhancement of catalytic activity of the PMDNAzyme. The mechanism of the DNA-dependent activation of PMDNAzyme containing probes was confirmed by CD spectroscopy as well as modeling of the probes and their complexes with DNA target. The calibration curves for HIV DNA determination allowed estimating the analytical parameters of the assay. The detection limit value and the linear range were shown to be 0.3 nM and 0.3–15 nM, respectively. The assay sensitivity was high (190000 nM–1). The values of coefficient of variation (CV) measured within the working range varied less than 4%, which indicates the high accuracy of the proposed assay.

Ultra-sensitive chemiluminescence imaging DNA hybridization method in the detection of mosquito-borne viruses and parasites

BackgroundMosquito-borne viruses (MBVs) and parasites (MBPs) are transmitted through hematophagous arthropods-mosquitoes to homoiothermous vertebrates. This study aims at developing a detection method to monitor the spread of mosquito-borne diseases to new areas and diagnose the infections caused by MBVs and MBPs.MethodsIn this assay, an ultra-sensitive chemiluminescence (CL) detection method was developed and used to simultaneously detect 19 common MBVs and MBPs. In vitro transcript RNA, virus-like particles (VLPs), and plasmids were established as positive or limit of detection (LOD) reference materials.ResultsMBVs and MBPs could be genotyped with high sensitivity and specificity. The cut-off values of probes were calculated. The absolute LODs of this strategy to detect serially diluted in vitro transcribed RNAs of MBVs and serially diluted plasmids of MBPs were 102–103copies/μl and 101–102copies/μl, respectively. Further, the LOD of detecting a strain of pre-quantified JEV was 101.8–100.8PFU/ml, fitted well in a linear regression model (coefficient of determination = 0.9678).ConclusionsUltra-sensitive CL imaging DNA hybridization was developed and could simultaneously detect various MBVs and MBPs. The method described here has the potential to provide considerable labor savings due to its ability to screen for 19 mosquito-borne pathogens simultaneously.

Detection of Anti-Domain I β-2 Glycoprotein I Antibodies as New Potential Target in Antiphospholipid Syndrome Diagnosis

Anti-cardiolipin antibodies (ACL) and anti-β-2-glycoprotein I antibodies (anti-2GPI) represent two out of three laboratory criteria for detection of antiphospholipid syndrome (APS). The domain I (DI) in anti-β-2-glycoprotein I is a new target for better identification of antibodies and may be associated with thrombotic risk in antiphospholipid syndrome. Anti-β2GPI antibodies specifically reacting with DI have a particular clinical importance being more commonly detected among patients with APS and other autoimmune diseases. This observation implies that compared with antibodies targeting the whole molecule, anti-DI antibodies have higher specificity for APS. Routine testing for anti- DI antibodies in clinical practice can be used for an easy differentiation of subjects carrying clinically meaningful anti- β2GPI antibodies from those individuals with a benign autoantibody profile. The aim of our study was to determine the significance of the domain I in the anti-β-2-glycoprotein I as a new biomarker for determining thrombotic risk in antiphospholipid syndrome. We investigate the DI anti-β2GPI on a group of 74 patients with antiphospholipid syndrome diagnosis. All patients have positive antibodies in at least one class ACL and anti-β2GPI antibodies. We detect DI anti-β2GPI positivity in 21 samples in our group. The thrombotic complications had been observed in 21 from 74 patients. The incidence of thrombotic complications in the total group was established as 28.4%, in comparison to the group DI anti-β2GPI positive with the incidence of thrombotic complications 57%. Performing of assay improved a positive predictive value from 25% pre-test to 68% for patients with positive test. The new chemiluminescent method for detection of DI anti-β2GPI shows a better compliance with clinical outcome than the actual diagnostic scheme.