Immunochromatographic Assay Research Articles

Development and evaluation of a lateral flow immunochromatographic assay for the rapid detection of donkey meat in beef as a tool for meat adulteration identification

Development and evaluation of a lateral flow immunochromatographic assay for the rapid detection of donkey meat in beef as a tool for meat adulteration identification

Establishment of Broad-Specificity Monoclonal Antibody-Based Immunoassay for Rapid Detection of Indole-Type and Indazole-Type Synthetic Cannabinoids and Metabolites.

Synthetic cannabinoids (SCs) have emerged as one of the most severely abused categories of new psychoactive substances (NPS), exacerbating the global drug problem and posing significant threats to public health. Presently, a class of new amide-type SCs featuring an indazole or indole core has been identified in numerous cases of illegal drug use, but there is still a lack of comprehensive analysis methods of SC detection. Herein, monoclonal antibodies (mAbs) 2E4 and AE6 targeting 36 indole-type and indazole-type SCs and their metabolites with IC50 ranging from 0.14 to 85.28 ng/mL were prepared and the molecular mechanism of antibody recognition was elaborated. We established the indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and gold immunochromatography assay (GICA) based on mAbs 2E4 and AE6 to detect indazole-type and indole-type SCs in urine and hair samples. Under optimal conditions, the proposed method detected ADB-BUTINACA (an indazole-type SC) with limits of detection (LODs) of 0.11 ng/mL for urine and 0.024 ng/mg for hair by ic-ELISA, and 1.02 ng/mL for urine and 0.046 ng/mg for hair by GICA; the LODs of 4F-MDMB-BUTICA (an indole-type SC) detection was 0.036 ng/mL for urine and 0.012 ng/mg for hair by ic-ELISA, and 0.54 ng/mL for urine and 0.03 ng/mg for hair by GICA. Collectively, our study provides a comprehensive foundation for the rapid screening and quantitation of SC derivatives in biological samples.

Dendritic mesoporous silica loaded with gold nanoclusters and their application in immunochromatographic assay.

Novel green-emitting fluorescent microspheres (GreFMPs) were assembled by loading highly luminescent gold nanoclusters (Arg/ATT/AuNCs) ondendritic mesoporous silica nanoparticles (DMSNs) via PEI-mediated electrostatic adsorption. The fluorescence microspheres exhibit excellent monodispersion with average diameters about (254.5 ± 34.6nm). Compared with free Arg/ATT/AuNCs, the GreFMPs have similar fluorescent properties and biocompatibility but superior environmental tolerance. Subsequently, an immunochromatographic assay based on GreFMPs (GreFMP-ICA) has been successfully developed, which is highly selective toward alphafetoprotein (AFP) in human serum. Under the optimal parameters, there is a good linear range between 0.5 and 160.0ng/mL, the limit of detection was found to be about 0.5ng/mL, and the recovery and relative standard deviation are 81.0 ~ 100.6% and 3.5 ~ 16.6%, respectively. These results manifested that the GreFMPs are beneficial for the rational design of the ICA platform, and the proposed GreFMP-ICA has the potential to screen AFP in clinical applications.

Quantitative immunochromatographic assay for rapid detection of rosemary herb extract in ham sausage and dairy products

Quantitative immunochromatographic assay for rapid detection of rosemary herb extract in ham sausage and dairy products

Epidemiological Dynamics of Canine Morbillivirus in Resident Dogs of Makurdi Metropolis, Benue State, Nigeria

Canine distemper is an endemic viral disease of dogs in Nigeria. Knowledge of the carrier status of dogs is key to successful control of the disease. In this study, the carrier status of apparently healthy dogs’ resident in parts of Makurdi metropolis was determined using the immunochromatographic rapid antigen assay kit for qualitative detection of canine morbillivirus antigens in ante-mortem samples. Ocular-, nasal-, and rectal- swabs, as well as serum were taken from each of 204 dogs bringing a total of 816 samples. The results showed that 26.96 % of the dogs sampled were positive for Canine morbillivirus antigens. Viral antigens were detected in 9.8%, 11.27%, and 4.41% of nasal, ocular, and rectal swaps respectively, and in 6.37% of the serum samples collected. An age-related susceptibility was observed as viral antigens detection rate was higher in younger dogs compared to older ones. Similarly, 40.43% of vaccinated and 22.93% of unvaccinated dogs were positive for canine morbillivirus antigens. Of significant importance is the prevalence rate in unvaccinated population of dogs. In terms of breed-related detection rate, 23.93% of Nigerian local dogs and 40.63% of exotic breeds tested positive for viral antigens, and it was observed that 25.54% of dogs which had history of contact with other dogs and 40% of dogs which had no such history carried canine morbillivirus antigens. The significance of this study is that it details the current epidemiological dynamics of canine morbillivirus in resident dogs of Makurdi metropolitan area, and the findings are discussed. Subject Areas: Animal infectious disease epidemiology.

Rapid and sensitive detection of staphylococcal enterotoxin E using a time-resolved fluorescence immunochromatography assay

Staphylococcal food poisoning is a significant foodborne illness caused by staphylococcal enterotoxins (SEs). Immunoassays have become the primary method for rapidly detecting harmful bacteria and toxins because of their excellent sensitivity and specificity. However, these assays have limitations in that they cannot differentiate between types of SEs and do not provide rapid, on-site, quantitative testing. In this study, a time-resolved fluorescence immunochromatography assay (TRFICA) was developed specifically for detecting staphylococcal enterotoxin E (SEE), which is commonly found in dairy products. Compared with a double antibody sandwich enzyme-linked immunosorbent assay, which had a detection limit of 0.028 ng/mL, TRFICA demonstrated comparable sensitivity, enabling SEE quantification with a detection limit as low as 0.081 ng/mL in infant formula. Validation by spiking infant formula samples confirmed no cross-reactivity with analogs (recoveries ranged from 93.17% to 128.77%). Furthermore, with an 8-min reaction time and interpretation delivered by a portable TRFICA strip reader, our method demonstrates potential for use in mobile and on-site detection. This study describes a rapid, easy, and reliable method for detecting trace levels of SEE in infant formula, which could serve as an early screening tool toward preventing food poisoning in infants and children.

Development of a colloidal gold immunochromatographic test strip for detecting the smooth Brucella

SummaryBrucellosis, caused by Gram-negative Brucella, spreads in human and animal populations through contact with infected animals and products. Developing a rapid and sensitive detection technology for pathogen is crucial to reduce the risk of this disease transmitting between animal populations and to humans. We produced a monoclonal antibody LPS-6B5, which shows high affinity to LPS and limited cross-reactivity with other bacteria. Based on LPS-6B5, a colloidal gold immunochromatographic assay (GICA) was developed which demonstrates high sensitivity and specificity in detecting cultured B. melitensis, B. abortus and B. suis. The Gold Immunochromatographic Assay (GICA) strips exhibited the most sensitive detection limits, with a value of 7.8125 × 105 CFU/mL for Brucella melitensis, surpassing the sensitivity levels observed for Brucella abortus and Brucella suis. It is also suitable for clinical and field samples, providing a cost-effective and user-friendly alternative to traditional methods.

Low cost and real-time surveillance of enteric infection and diarrhoeal disease using rapid diagnostic tests in Cox’s Bazar, Bangladesh

BackgroundReal-time disease surveillance is an important component of infection control in at-risk populations. However, data on cases or from lab testing is often not available in many low-resource settings. Rapid diagnostic tests (RDT), including immunochromatographic assays, may provide a low cost, expedited source of infection data.MethodsWe conducted a pilot survey-based prevalence mapping study of enteric infection in Camp 24 of the camps for the forcibly displaced Rohingya population from Myanmar in Cox’s Bazar, Bangladesh. We randomly sampled the population and collected and tested stool from under-fives for eight pathogens using RDTs in January–March 2021 and September–October 2021. A Bayesian geospatial statistical model allowing for imperfect sensitivity and specificity of the tests was adapted.ResultsWe collected and tested 396 and 181 stools in the two data collection rounds. Corrected prevalence estimates ranged from 0.5% (Norovirus) to 27.4% (Giardia). Prevalence of Escherichia coli O157, Campylobacter, and Cryptosporidium were predicted to be higher in the high density area of the camp with relatively high probability (70–95%), while Adenovirus, Norovirus, and Rotavirus were lower in the areas with high water chlorination. Clustering of cases of Giardia and Shigella was also observed, although associated with relatively high uncertainty.ConclusionsWith an appropriate correction for diagnostic performance RDTs can be used to generate reliable prevalence estimates, maps, and well-calibrated uncertainty estimates at a significantly lower cost than lab-based studies, providing a useful approach for disease surveillance in these settings.

Evaluation of a point-of-care multiplex immunochromatographic assay for the diagnosis of typhoid: results from a retrospective diagnostic accuracy study.

Currently available diagnostic tests for typhoid have several limitations, including low sensitivity and specificity. Dual Path Platform Typhoid assay is a multiplex rapid test that detects IgA antibodies to lipopolysaccharide and hemolysin E antigen. It is considered to have high sensitivity and specificity, and its results were found to be highly correlated with ELISA results. However, very few studies have been conducted to evaluate this test and limited information about the accuracy of this test is present. Hence, this study evaluated the new typhoid test.

Human Papillomaviruses 16 and 18 E6 Oncoprotein Detection Test in Primary Oropharyngeal Carcinomas and Metastatic Lymph Nodes: A Cross-Sectional Study.

Accuracy in the diagnosis of HPV-associated oropharyngeal carcinoma (OPSCC) of a rapid, low-cost lateral flow immunochromatographic assay for detecting E6 oncoprotein of HPV-16 and HPV-18 was previously evaluated in a small pilot study. This cross-sectional study aimed to assess on a large case series the sensitivity and specificity of E6 oncoprotein as a diagnostic marker for HPV-associated carcinogenesis in OPSCC. 137 consecutive patients with histologically confirmed OPSCC were enrolled in two hospitals in Northeast Italy. HPV status was determined by PCR for HPV DNA and p16INK4a immunohistochemistry on primary tumor biopsies. An OPSCC was defined as HPV-associated when double positive for high-risk HPV-DNA and p16INK4a overexpression in primary lesion. Cytological samples from primary tumors and metastatic lymph nodes were obtained and tested for HPV16/18 E6 oncoproteins using the lateral flow immunochromatographic assay, which requires between 90 and 120 min to provide a result. Diagnostic performances were calculated as percentage with confidence intervals (CI). Of the 137 OPSCC cases, 68 (49.6%) were HPV-associated, testing positive for both high-risk HPV-DNA and p16INK4a, with HPV16 predominating (82.4%). An average waiting time of 22 days was observed to obtain the results of p16INK4a and HPV-DNA after primary lesions biopsy. In patients with HPV16/18-associated OPSCC, the HPV16/18 E6 oncoprotein was detected in 59 out of 60 cytological samples from the primary lesion (sensitivity: 98.3%; 95% CI: 91.1-100%) and in 45 out of 51 cytological samples from lymph node metastases (sensitivity: 88.2%; 95% CI: 76.1-95.6%). The E6 oncoprotein assay showed a specificity of 100% in both primary tumors and lymph node metastases. The low-cost lateral flow immunochromatographic assay for detecting HPV16/18 E6 oncoproteins confirmed high accuracy for identifying HPV-associated OPSCC, particularly in primary tumors, suggesting its potential as a valuable diagnostic tool in clinical practice. Its rapid diagnostic capability could significantly accelerate the process of treatment decision-making, enhancing the timely management of patients.

A curious case of a negative control line on a dengue duo assay due to interference - A case report.

An elderly patient who presented with 1week of fever and respiratory symptoms was tested for dengue infection with an Abbott Bioline™ Dengue Duo immunochromatographic assay. Unexpectedly, the control line of the dengue non-structural protein 1 (NS1) component was absent, necessitating result invalidation. It remained absent when the test was repeated on the SD Biosensor Standard™ Q Dengue Duo, but present on the Wells Bio careUSTM Dengue Combo and Asan Easy Test® Dengue DUO assays, suggesting potential interference. Dilution, polyethylene glycol (PEG) precipitation and centrifugation with a 100kDa filter were performed to reduce/remove the potential interferent. Sera from other patients that showed a control line, and a test line that was either positive or negative for NS1, were used as controls. Upon dilution with negative control serum, a faint control line emerged. PEG precipitation resulted in disappearance of control and test lines in the positive control. Filtration led to emergence of the control line for the patient's serum but caused the test line for the positive control serum to disappear. Overall, investigations suggested the presence of a high molecular weight (>100kDa) substance which interferes with chicken IgY-anti-chicken IgY binding at the control line of affected assays. Our results highlight two important points: firstly, some commonly used laboratory procedures (e.g. PEG or filtration) may inadvertently remove the target biomarker (e.g. multimeric NS1) and should be interpreted with appropriate controls. Secondly, alternative kits that use a different antigen-antibody combination for the control line can be considered when similar patients are encountered in future.

Ultra-rapid detection of nuclear protein of severe fever with thrombocytopenia syndrome virus by colloidal gold immunochromatography assay.

In 2009, severe fever with thrombocytopenia syndrome virus (SFTSV), also known as the Dabie bandavirus (DBV), was first discovered in Henan, China. It is a tick-borne zoonotic virus with a fatality rate ranging from 6% to 30%. Currently, we lack safe and effective vaccines or antiviral drugs to treat SFTSV infection. Therefore, the development of a specific, sensitive, and cost-effective detection method is crucial. Using inactivated SFTSV and recombinant SFTSV nucleocapsid protein (SFTSV-NP), we repeatedly immunized mice with different adjuvants and obtained two monoclonal antibodies against SFTSV-NP, which were used to develop a colloidal gold immunochromatographic assay (ICA) rapid test kit for SFTSV. Compared with nucleic acid testing (gold standard), the ICA test strips are 97.67% accurate in testing clinical serum samples (36 cases of clinical serum samples and seven cases of whole blood samples). The test kit was 100% accurate in detecting different SFTSV strains. No false-positive results were generated when detecting other arboviruses. Therefore, our developed SFTSV test kit conveniently, rapidly, and effectively detects SFTSV.

Highly sensitive time-resolved fluorescent microspheres lateral flow immunoassay for the quantitative detection of triadimefon and its metabolite residues in fruits and vegetables.

Ageneral one-step lateral flow immunochromatographic assay (LFIA) for the quantitative detection of triadimefon (TDF) and triadimenol(TDN) in fruit and vegetable samples was developedusing time-resolved fluorescence microspheres (TRFM) as labels. A specific anti-triadimefon monoclonal antibody (mAb) was conjugated with TRFM to fabricate LFIA test strips. A time-resolved fluorometer as an LFIA reader was applied to obtain quantitative results and assess risk ranges for the LFIA test strips. Under the optimized experimental conditions, the limits of detection (LODs) in buffer/cucumbers/tomatoes/oranges were 0.046ng/mL, 0.135µg/kg, 1.047µg/kg, and 5.811µg/kg, respectively, which are ca. 1000 times lower than that of colloidal gold-labeled strips. The recovery in cucumber/tomato/orange samples was 109.4-116.7%, 87.7-110.9%, and 88.0-111.9%, respectively, indicating that the test strips had good reliability. Coupled with the easily customizable pretreatment procedures for various samples, the LFIA results were obtained within 18min without the need for professional personnel or complicated equipment. TRFM-LFIA for TDF and TDN also shows remarkable specificity and precision. The test strips were also low-cost, portable, and convenientto use. These results indicate the test strips could be utilized as a novel strategy for on-site detection of TDF and TDN, which has the potential to expand and detect other pesticide or insecticide residues in food.

The prevalence of Tuberculosis in HIV patients and their correlation with CD4+ count in a tertiary care hospital in Rishikesh, gateway to the Himalayas

IntroductionTuberculosis is the most common serious opportunistic infection in HIV-positive patients. TB can occur at any time during the course of HIV infection. The estimated number of people living with HIV in India in 2019 was 23.48 lakh. With an estimated 2.5 million cases, or 195 cases per 1 lakh people, India has the highest incidence of tuberculosis cases worldwide, accounting for about 23% of all cases, according to MOHFW India. Information about HIV/TB co-infection is still scarce in India, especially in the hilly areas of Uttarakhand. Material and methodsIn this retrospective study, subjects are the patients who are diagnosed with MTB in HIV-positive patients of all age groups and sex and were followed up from January 2021 till August 2022 in a tertiary referral hospital AIIMS Rishikesh.Diagnosis of MTB was based on clinical, laboratory, and neuroimaging signs, and HIV was based on dot blot, immunochromatographic, and chemiluminescence assay. ResultA total of 520 patients screened for HIV were positive. Out of which 145 were MTB in which 133 had pulmonary tuberculosis and 12 had extra-pulmonary tuberculosis. The patients' average age was 35.5 years. Among males, females and transgender, 97 (66.89%), 46 (31.72%) and 2 (1.38%) have both TB and HIV co-infections respectively.Mean CD4 count prior to ATT was 350.21, After completion was 486.48. The prevalence of Tuberculosis among HIV positive cases was 23.8%.Baseline and latest CD4 count was conducted before and after ART and ATT treatment. It was observed that there was significant increase in the CD4 count after treatment. ConclusionHIV co-infection with TB is higher in this region. There is a role of CD4+ count in the prognosis and progression of the disease. More strategic measures are required for early diagnosis and treatment of HIV to combat MTB.

Comparative analysis of the results of live anthrax vaccine identification by immunofluorescence and immunochromatography

INTRODUCTION. The quality evaluation of live anthrax vaccines will benefit from the implementation of alternative testing methods that are capable of specific identification of the Bacillus anthracis spore antigen using the appropriate diagnostic products that are authorised in the Russian Federation for the detection of B. anthracis spores.AIM. This study aimed to investigate the applicability of immunofluorescence analysis to the identification of live anthrax vaccines and compare this method with immunochromatography.MATERIALS AND METHODS. The study used commercial batches of a live anthrax vaccine and Russian diagnostic products, including diagnostic dry adsorbed fluorescent anti-anthrax spore immunoglobulins and an immunochromatographic assay (ICA) reagent kit for rapid detection and identification of B. anthracis spores (ICA system for B. anthracis). For smears and ICA system reactions, the authors prepared working solutions of bacterial suspensions at spore concentrations typical of the B. anthracis STI-1 vaccine strain. The spore concentrations were achieved using the pharmacopoeial reference standard (RS) for the opacity of bacterial suspensions of 10 international units (IU). Identification reactions involved the registration of immune complex formation.RESULTS. Immunofluorescence tests of the live anthrax vaccine demonstrated bright greenish-yellow envelope fluorescence with an intensity score of 3+ to 4+ for smears stained with diagnostic immunoglobulins at 1:32 and 1:64 dilutions. Immunochromatographic tests of the live anthrax vaccine detected the spore antigen at vaccine concentrations of 109 and 108 spores/mL, with the test strips showing two distinct dark-pink lines indicative of immunoprecipitation. According to the results obtained using the selected methods, the tested microbial culture was confirmed as B. anthracis.CONCLUSIONS. Immunochromatography and immunofluorescence tests with appropriate diagnostic preparations are convenient and reliable tools for the species-specific detection of B. anthracis STI-1 spores in the live anthrax vaccine. The results obtained in the anthrax vaccine identification tests provide a basis for recommending the above methods as supplementary alternatives to Ziehl–Neelsen bacteriological staining, which is currently prescribed by the State Pharmacopoeia of the Russian Federation.

Rapid screening of prednisolone acetate adulterants in health foods using colloidal gold immunochromatographic assay

In this study, a rapid detection method utilizing colloidal gold immunochromatography (CG-ICA) was developed for the detection of illegally added prednisone acetate in health foods. Initially, the preparation conditions of colloidal gold solution were optimized. The optimal potassium carbonate dosage, antibody diluent type, antibody dosage, probe labeling time, blocking time and BSA dosage were determined. Technical analysis was performed to ensure that the established CG-ICA exhibited satisfactory color development and inhibition rates. Under optimized conditions, the cut-off value of CG-ICA was 250 μg/kg. The assay demonstrated a sensitivity of 100 %, a false positive rate of 8 %, and a false negative rate of 0, indicating high specificity for prednisone acetate. The results obtained from testing actual samples were consistent with those obtained using LC-MS/MS, thereby verifying the reliability of the developed method. This method offers robust support for the rapid detection of illegally added prednisone acetate in health foods.

Dual-mode immunochromatography based on Ti3C2Tx functionalized CdTe quantum dots for chloramphenicol detection in animal-derived foods

In this study, Ti3C2Tx was coupled with CdTe quantum dots via plyethylenimine (PEI) mediation to form composite nanomaterials with dual colorimetric and fluorescent properties, which had strong pH, high ion strength, and organic solvent resistance. Then, this novel Ti3C2Tx@CdTe was firstly applied to develop a dual-mode immunochromatographic assay (ICA) for chloramphenicol (CAP) detection in animal-derived foods. The limits of detection of the colorimetric and fluorescent modes for CAP in milk/chicken/fish were 0.01 and 0.005 μg/kg, the limits of quantification were 0.025 and 0.016 μg/kg, the recovery rates were between 85.7% and 129.0% and 84.0%-119.2%, with the corresponding variations ranged from 6.5% to 14.8% and 6.4%-14.9%, respectively. The sensitivity of fluorescence mode was up to 100-fold higher than that of the reported ICA for CAP detection. The results of the confirmatory experiment on 30 samples showed good consistency with that of our established method (R2>0.93). The above results indicated that Ti3C2Tx@CdTe had superior detection performance and can provide a dual-mode signal calibration, high stability and sensitivity selection for immunoassay detection platforms.

A-240 Validation of the OSOM Flu SARS-CoV-2 Combo Home Test

Abstract Background COVID-19 and Influenza respiratory infections often cause similar symptoms, however, treatment is different for each infection. Delay in diagnosis can be fatal in some cases, therefore rapid detection and differentiation are critical for appropriate medical treatment. The OSOM Flu SARS-CoV-2 Combo Home Test is a lateral flow immunoassay intended for the qualitative and differential detection of nucleocapsid protein from SARS-CoV-2 and influenza A/B proteins using self-collected anterior nasal swab samples (self-collection is only recommended for those 14 years of age and older). An interpretation card, included with the test, is used to reliably distinguish among possible testing results. Methods A prospective study was conducted at multiple geographically distinct sites in the U.S during the 2023-2024 season for the validation of the OSOM Flu SARS-CoV-2 Combo Home Test to detect SARS-CoV-2/Flu A/Flu B in self-collected anterior nasal swab samples. The study evaluated the performance in symptomatic individuals only. A total of 703 prospective samples were evaluable for SARS. For Flu A/B, four additional subject samples were unevaluable due to comparator results not being available and a total of 699 samples were included in the data analysis. The comparator tests were FDA 510(k)-cleared molecular assays. Results Influenza A was detected with a sensitivity of 93.1% (95% CI: 84.8%-97.0%) and a specificity of 99.5% (95% CI: 98.6% - 99.8%). Influenza B was detected with a sensitivity of 89.1% (95% CI: 77.0% - 95.3%) and a specificity of 99.7% (95% CI: 98.9% - 99.9%). SARS CoV-2 was detected with a sensitivity of 87.0% (95% CI 78.6% - 92.4%) and a specificity of 99.1% (95% CI: 97.9% - 99.6%) when Ct values are standardized and 10% of samples are considered low positives, having a Ct value of ≥ 30. Conclusions With respiratory infections often showing indistinguishable symptoms and the possibility of co infection, lowering the testing hurdles for individuals to be able to perform a rapid differential test at home without the aid from a healthcare worker is the most effective way to mitigate spread and patient management from a public health perspective. Comparing the OSOM Flu SARS-CoV-2 Combo Home test to the FDA cleared comparators in a clinical setting, the OSOM Flu SARS-CoV-2 Combo Home test retains acceptable sensitivity and specificity. Using self-collected anterior nasal swabs which are minimally invasive alongside the low cost of a lateral flow immunochromatographic assay remains the most effective way to perform the serial screening testing recommended by the CDC in situations where transmission and severe outcome risk is high.

B-205 Performance of a Novel Fluorogenic Assay for Detection of Carbapenemase-producing Enterbacteriaceae

Abstract Background In recent years, under the selective challenge of antibiotics, the variety and number of drugresistant pathogenic microorganisms have increased significantly, which brings great challenges to clinical diagnosis and treatment, especially the infection caused by carbapenem resistant Enterobacteriaceae (CRE). Carbapenemases production is the main mechanism of drug resistance of Enterobacteriaceae to carbapenems. Drug resistance of Carbapenems can be caused by three mechanisms, resulting in the production of five major Carbapenemases. These are Klebsiella pneumoniae enzyme (KPC), New Delhi metal β-lactamase (NDM), carbapenem hydrolyzed oxalase (OXA-48 like), integrin-encoded metal β- lactamase (VIM) and IMP (Imipenemase). The real-time fluorescence quantitative PCR (qPCR) method based on molecular beacons was combined with the melting curve analysis to identify five drug resistance genes simultaneously by a single PCR reaction, with rapid detection, high sensitivity and strong specificity. Based on this principle, we developed the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae (Dynamiker Biotechnology (Tianjin) Co., Ltd.). Methods We evaluated the performance of the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae, including the limit of detection (LoD) and cross-reactivity, and compared it with the lateral flow immunochromatography assay (LFA). Results The LoD ranged from 75-450 CFU/mL for the five carbapenemase genes. The analytical specificity for target genes was 100%, as assessed with a panel of 15 pathogens, which indicated no cross-reactions. Comparison of qPCR and LFA results from twenty-three CRE clinical isolates with characterized carbapenemase content demonstrated a complete agreement (Table 1). Conclusions The novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae is an accurate and rapid method to identify KPC, NDM, VIM, IMP and OXA-48-like carbapenemases in the clinical microbiology laboratory, which can guide infection control programs to limit the spread of these organisms.

Lychee-shaped magneto-gold nanohybrid with highly retained magnetic-plasmonic activity for ultrasensitive and multiplex immunochromatographic assay

Utilizing nanomaterials with magnetic and plasmonic properties can significantly enhance the sensitivity of immunochromatographic assay (ICA) through magnetic enrichment, and signal amplification. Herein, we successfully synthesized a lychee-shaped magneto-gold nanohybrid (designated as MSiAuTA). This nanohybrid comprises a magnetic nanoparticle core, a flesh-fiber-like mesoporous silica template hosting numerous gold nanoparticles (AuNPs), and a lychee-exocarp-like tannic acid layer. The resultant MSiAuTA exhibits rapid magnetic response, strong plasmonic activity, and excellent biocompatibility. We further developed a multiplex ICA (mICA) platform utilizing MSiAuTA as labels for target enrichment and colorimetric signal amplification. This platform enables simultaneous and quantitative detection of Escherichia coli O157:H7 and Salmonella typhimurium. The MSiAuTA-mICA supports portable strip reader and smartphone readout and demonstrates exceptional sensitivity, achieving detection limits of 9.8 × 102 CFU/mL for E. coli O157:H7 and 4.9 × 102 CFU/mL for S. typhimurium. Additionally, we validated the accuracy, robustness, and practicality of MSiAuTA-mICA in detecting both spiked and naturally contaminated samples, including water, lettuce, chicken, apple juice, and milk. In conclusion, the lychee-shaped MSiAuTA nanohybrid shows significant promise as a versatile label for enhancing ICA’s detection capabilities in complex sample matrices.