Investigation of Chikungunya virus genotype at tertiary care center in Western Maharashtra, India

Background & objectives: Chikungunya is a reemerging arbovirus infection. Laboratory diagnosis can be done by Classical test involving Rapid Immunochromatography, Enzyme-Linked Immunosorbent assay and Molecular methods. The present study was undertaken to know the genotype of the Chikungunya virus (CHICKV) among patients suspected of CHICKV and investigated by virus culture, partial sequencing, Rapid Immunochromatography, and Enzyme-linked Immunosorbent assay (ELISA). To understand different techniques used in Chikungunya diagnosis viz., virus culture, partial sequencing along with Immunochromatography and ELISA. Methods: This is a prospective, laboratory-based study at a tertiary care center. Lateral flow chromatography and ELISA was carried out on serum samples. All 50 samples were cultured and indirect Immunofluorescence was performed on positive samples at Interactive Research School for Health Affairs (IRSHA), Bharati Vidyapeeth Medical College Pune, Maharashtra, India. Virus isolates were subjected to partial sequencing for identification of genotype after confirmation by PCR. Statistical Package of Social Science (SPSS) version 22.0 software was used to calculate the Receiver operating curve (ROC) for different tests. Results: Out of 50 samples, 20 were positive by Immunochromatography, 23 by ELISA, and 3 by culture, PCR confirmed CHIKV isolates and sequencing identified genotypes as East Central South African type. Interpretation & conclusion: CHIKV culture isolates of East Central South African type lineage were predominantly found in the present study. These are also common genotypes present in Asia including India.

Leptospira interrogans insoluble fraction as a potential antigen source for lateral flow immunochromatography

BACKGROUND Leptospirosis is an emerging zoonosis that affects humans and animals. Immunochromatography rapid test is widely used for early diagnosis of leptospirosis, but with low sensitivity and specificity. OBJECTIVES To evaluate Leptospira interrogans insoluble fraction as a potential antigen source for lateral flow immunochromatography. METHODS Insoluble fraction derived from the crude bacterial extract was obtained by serial centrifugation. The polypeptide profile was determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Immune reactivity of this fraction was assessed by Western Blotting and lateral flow immunochromatography (LFI). It was tested 160 microagglutination test (MAT)-positive sera from patients in the acute phase, 100 MAT-negative sera from patients with acute febrile illness, and 45 patients with other infectious diseases. FINDINGS There was a predominance of low molecular mass-polypeptide bands, ranging from 2 to 37 kDa. The antibody reactivity of theses polypeptides was found to range from 13-50%, especially between 10 and 38 kDa. Among MAT-positive sera of patients with leptospirosis in the acute phase, 97% were also positive in LFI, indicating high sensitivity. Among MAT-negative sera, all were negative in LFI, indicating high specificity. Only 2% of cross-reactivity was detected. CONCLUSION The insoluble fraction can be a valuable antigen source for development of point-of-care diagnosis test for leptospirosis.

Subclinical signs of podocyte injury associated with Circulating Anodic Antigen (CAA) in Schistosoma mansoni-infected patients in Brazil

ABSTRACT Background: The long-term effects of schistosomiasis on the glomerulus may contribute to the development of chronic kidney disease. This study aimed to investigate baseline Schistosoma mansoni-Circulating Anodic Antigen (CAA) levels and their association with kidney biomarkers related to podocyte injury and inflammation in long-term follow-up after praziquantel (PZQ) treatment. Methods: Schistosoma infection was diagnosed by detecting CAA in urine using a quantitative assay based on lateral flow using luminescent up-converting phosphor reporter particles. A cutoff threshold of 0.1 pg/mL CAA was used to diagnose Schistosoma infection (baseline) in a low-prevalence area in Ceará, Northeast, Brazil. Two groups were included: CAA-positive and CAA-negative individuals, both of which received a single dose of PZQ at baseline. Urinary samples from 55 individuals were evaluated before (baseline) and at 1, 2, and 3 years after PZQ treatment. At all time points, kidney biomarkers were quantified in urine and adjusted for urinary creatinine levels. Results: CAA-positive patients had increased baseline albuminuria and proteinuria and showed greater associations between kidney biomarkers. CAA levels correlated only with Vascular Endothelial Growth Factor (VEGF) (podocyte injury) levels. Increasing trends were observed for malondialdehyde (oxidative stress), monocyte chemoattractant protein-1 (inflammation marker), and VEGF. In the follow-up analysis, no relevant differences were observed in kidney biomarkers between the groups and different periods. Conclusions: S. mansoni-infected individuals presented subclinical signs of glomerular damage that may reflect podocyte injury. However, no causal effect on long-term renal function was observed after PZQ treatment.

Development of lateral flow immunoassay for SARS-CoV-2 antigen detection / 中国胸心血管外科临床杂志

@#This article summarizes the development of lateral flow immunoassay for SARS-CoV-2 antigen detection. Lateral flow immunoassay is a rapid, low cost, and ease of use detection tool that has been widely applied in clinical and public health sectors. Since the outbreak of COVID-19, the technique has been adopted for rapid antigen diagnostic test of SARS-CoV-2, including commonly used colloidal gold nanoparticle-based lateral flow immunoassays as well as various fluorescence-based lateral flow immunoassays. With innovations in labelling methods, this detection technique has been in continuous development and is shifting from qualitative toward quantitative as well as gaining sensitivity.

Kaedah Diagnostik Semasa dan Penggunaan Ujian Titik Penjagaan Pantas (POC) Bagi Mendiagnos Hiperkolesterolemia Famili (FH) (Current Diagnostic Techniques and the Use of Rapid Point-of-Care (POC) Testing to Diagnose Familial Hypercholesterolemia (FH)) / Jurnal Sains Kesihatan Malaysia

@#Familial hypercholesterolemia (FH) is an autosomal dominant inherited genetic disease characterized by increased concentrations of low-density lipoprotein (LDL-C) cholesterol in the blood. The risk of premature coronary heart disease in FH patients may increase without early treatment. Advancement in molecular biology techniques has enable early detection and diagnosis of FH. These techniques are cost-effective and have a shorter turnaround time. The current diagnostic tools available for FH diagnosis involving algorithm-based scoring criteria and various molecular diagnosis methods including next-generation sequencing (NGS), Sanger sequencing, Multiplex ligation-dependent probe amplification (MLPA) and DNA hybridisation assay are discussed in this review. However, molecular genetic testing is not widely available due to time-consuming procedures, high cost and requires trained personnel. Thus, this 36 review highlights the use of point of care (POC) testing as an approach to diagnose FH, particularly in countries lacking infrastructure and expertise in this field. Lateral flow testing (LFA) has gained attention as a POC diagnostic tool due to its simplicity, low cost and involved simple procedure and settings. The advantages of LFA made this technique a potential tool in addressing challenges in diagnosing FH, particularly for early diagnosis of family members.

Low performance of a SARS-CoV-2 point-of-care lateral flow immunoassay in symptomatic children during the pandemic

Abstract Objective: to evaluate the accuracy of an antibody point-of-care lateral flow immunoassay (LFI -Wondfo Biotech Co., Guangzhou, China) in a pediatric population. Methods: children and adolescents (2 months to 18 years) with signs and symptoms suggestive of acute SARS-CoV-2 infection were prospectively investigated with nasopharyngeal RT-PCR and LFI at the emergency room. RT-PCR was performed at baseline, and LFI at the same time or scheduled for those with less than 7 days of the clinical picture. Overall accuracy, sensitivity and specificity were assessed, as well as according to the onset of symptoms (7-13 or ≥14 days) at the time of the LFI test. Results: In 175 children included, RT-PCR and LFI were positive in 51 (29.14%) and 36 (20.57%), respectively. The overall sensitivity, specificity, positive and negative predictive value was 70.6% (95%CI 56.2-82.5), 96.8% (95%CI 91.9-99.1), 90.0% (95%CI 77.2-96.0), and 88.9% (95%CI 83.9-92.5), respectively. At 7-13 and ≥14 days after the onset of symptoms, sensitivity was 60.0% (95%CI 26.2-87.8) and 73.2% (95%CI 57.1-85.8) and specificity was 97.9% (95%CI 88.7-99.9) and 96.1% (95%CI 89.0-99.2), respectively. Conclusion: Despite its high specificity, in the present study the sensitivity of LFI in children was lower (around 70%) than most reports in adults. Although a positive result is informative, a negative LFI test cannot rule out COVID-19 in children.

Technical performance of a lateral flow immunoassay for detection of anti-SARS-CoV-2 IgG in the outpatient follow-up of non-severe cases and at different times after vaccination: comparison with enzyme and chemiluminescent immunoassays

ABSTRACT This study assessed the technical performance of a rapid lateral flow immunochromatographic assay (LFIA) for the detection of anti-SARS-CoV-2 IgG and compared LFIA results with chemiluminescent immunoassay (CLIA) results and an in-house enzyme immunoassay (EIA). To this end, a total of 216 whole blood or serum samples from three groups were analyzed: the first group was composed of 68 true negative cases corresponding to blood bank donors, healthy young volunteers, and eight pediatric patients diagnosed with other coronavirus infections. The serum samples from these participants were obtained and stored in a pre-COVID-19 period, thus they were not expected to have COVID-19. In the second group of true positive cases, we chose to replace natural cases of COVID-19 by 96 participants who were expected to have produced anti-SARS-CoV-2 IgG antibodies 30-60 days after the vaccine booster dose. The serum samples were collected on the same day that LFIA were tested either by EIA or CLIA. The third study group was composed of 52 participants (12 adults and 40 children) who did or did not have anti-SARS-CoV-2 IgG antibodies due to specific clinical scenarios. The 12 adults had been vaccinated more than seven months before LFIA testing, and the 40 children had non-severe COVID-19 diagnosed using RT-PCR during the acute phase of infection. They were referred for outpatient follow-up and during this period the serum samples were collected and tested by CLIA and LFIA. All tests were performed by the same healthcare operator and there was no variation of LFIA results when tests were performed on finger prick whole blood or serum samples, so that results were grouped for analysis. LFIA's sensitivity in detecting anti-SARS-CoV-2 IgG antibodies was 90%, specificity 97.6%, efficiency 93%, PPV 98.3%, NPV 86.6%, and likelihood ratio for a positive or a negative result were 37.5 and 0.01 respectively. There was a good agreement (Kappa index of 0.677) between LFIA results and serological (EIA or CLIA) results. In conclusion, LFIA analyzed in this study showed a good technical performance and agreement with reference serological assays (EIA or CLIA), therefore it can be recommended for use in the outpatient follow-up of non-severe cases of COVID-19 and to assess anti-SARS-CoV-2 IgG antibody production induced by vaccination and the antibodies decrease over time. However, LFIAs should be confirmed by using reference serological assays whenever possible.

Development of a Recombinase-aided Amplification Combined With Lateral Flow Dipstick Assay for the Rapid Detection of the African Swine Fever Virus / 生物医学与环境科学（英文）

OBJECTIVE@#To establish a sensitive, simple and rapid detection method for African swine fever virus (ASFV) B646L gene.@*METHODS@#A recombinase-aided amplification-lateral flow dipstick (RAA-LFD) assay was developed in this study. Recombinase-aided amplification (RAA) is used to amplify template DNA, and lateral flow dipstick (LFD) is used to interpret the results after the amplification is completed. The lower limits of detection and specificity of the RAA assay were verified using recombinant plasmid and pathogenic nucleic acid. In addition, 30 clinical samples were tested to evaluate the performance of the RAA assay.@*RESULTS@#The RAA-LFD assay was completed within 15 min at 37 °C, including 10 min for nucleic acid amplification and 5 minutes for LFD reading results. The detection limit of this assay was found to be 200 copies per reaction. And there was no cross-reactivity with other swine viruses.@*CONCLUSION@#A highly sensitive, specific, and simple RAA-LFD method was developed for the rapid detection of the ASFV.

Evaluation of seven SARS-CoV-2 rapid IgM/IgG tests

RESUMEN Introducción: A finales de 2019, se detectó un nuevo coronavirus en China que provocó una enfermedad respiratoria aguda conocida como COVID-2019. Objetivo: Evaluar siete sistemas comerciales para la detección rápida de anticuerpos para determinar su sensibilidad, especificidad y robustez en nuestras condiciones para ser utilizados por el Sistema Nacional de Salud. Métodos: Se evaluaron siete sistemas para la detección de anticuerpos IgM/IgG. Se conformó un panel de evaluación con muestras de individuos negativos, sueros de otras afecciones previas a la pandemia y de pacientes positivos con la enfermedad. Resultados: Las cifras de sensibilidad general oscilan entre el 25 % y el 88 %, siendo los sistemas Realy Tech y Deep Blue los que mostraron los mejores resultados. La especificidad para ambos fue del 100 %. La tasa de IgM positiva según Realy Tech o Deep Blue aumentó a 94,1 % o 81,8 % en la etapa tardía de la enfermedad. Conclusiones: Los sistemas Realy Tech y Deep Blue detectaron IgM/IgG en suero y en sangre total con adecuada sensibilidad y especificidad. La reactividad cruzada no parece ser un problema. La serología en el caso de COVID-19 no puede utilizarse como diagnóstico pero permite a la vigilancia epidemiológica conocer el estado inmunológico de las poblaciones. Es fundamental analizar la respuesta inmune frente a la infección para realizar la caracterización epidemiológica y potencialmente informar el riesgo individual de futuras enfermedades y el estudio de posibles vacunas.

ABSTRACT Introduction: In late 2019, a new coronavirus was detected in China causing an acute respiratory illness known as COVID-2019. Objective: Evaluate seven commercial systems for the rapid detection of antibodies to determine their sensitivity, specificity and robustness in our conditions to be used by the National Health System. Methods: Seven systems were evaluated for the detection of IgM/IgG antibodies. Evaluation panel with samples from negative individuals, sera from other pathologies prior to the pandemic and from positive patients with the disease were conformed. Results: General sensitivity figures range between 25 and 88%, with the Realy Tech and Deep Blue systems showed the best results. The specificity for both was 100%. The IgM positive rate according to Realy Tech or Deep Blue increased to 94.1 or 81.8% in the late stage of the disease. Conclusions: Realy Tech and Deep Blue systems detected IgM/IgG in serum and in whole blood with adequate sensitivity and specificity. Cross-reactivity does not seem to be a problem. Serology in the case of COVID-19 cannot be used as a diagnostic but it allows epidemiological surveillance to know the immune status of populations. It's essential to analyze the immune response against the infection to carry out epidemiological characterization and potentially inform individual risk of future disease and the study of potential vaccines.

Antigenemia temprana de Cryptococcus en pacientes asintomáticos VIH positivos en Barranquilla, Colombia / Cryptococcus early antigenemia in HIV positive asymptomatic patients in Barranquilla, Colombia

Resumen La criptococosis meníngea presenta alta mortalidad mundial, especialmente en población VIH/sida. La OMS recomienda detectar el antígeno capsular de Crypto coccus como estrategia para un diagnóstico temprano y poder minimizar complicaciones. Objetivo: realizar antigenemia temprana de Cryptococcus mediante in munocromatografía/ensayo de flujo lateral en pacientes asintomáticos VIH+. Material y método: estudio descriptivo observacional; entre julio-2016 y mayo-2019 se procesaron mediante ensayo de flujo lateral, muestras de suero de 169 pacientes asintomáticos VIH+, con CD4 ≤120 cel/μL en Barranquilla, Colombia. Ante resultado positivo, se indicó profilaxis con fluconazol; se hizo seguimiento a todos los casos. Resultados: la antigenemia fue positiva en cinco pacientes (2,96%); uno falleció, cuatro recibieron profilaxis y la prueba se negativizó en dos. Los pacientes con resultado negativo inicial no desarrollaron durante el estudio sinto matología compatible con esta micosis. Discusión: el ensayo de flujo lateral de Cryptococcus está recomendado para el diagnóstico temprano de la criptococosis en población VIH/sida. Conclusión: detectar tempranamente el antígeno circulante de Cryptococcus mediante ensayo de flujo lateral en pacientes asintomáticos VIH+, permitió instaurar profilaxis oportuna, hacer seguimiento y control para reducir la mortalidad asociada con la criptococosis meníngea.

Abstract Meningeal cryptococcosis presents high levels of global mortality, especially in the HIV/AIDS population. The WHO recommends detecting the capsular antigen as an important strategy for early diagnosis and be able to minimize complications. Objective: Perform early cryptococcal antigenemia by immunochromatographic/ lateral flow assay in asymptomatic HIV+ patients. Material and method: descriptive observational study; between July-2016 and May-2019, serum samples from 169 asymptomatic HIV+ patients with CD4 ≤120 cells/μL were processed by lateral flow assay in Barranquilla, Colombia. Given a positive result, prophylaxis with fluconazole was indicated; all cases were followed up. Results: antigenemia was positive in five (2.96%) patients; one died; four received prophylaxis, and the test turned negative in two. The patients with an initial negative result, did not developed symptoms compatible with this mycosis during the study period. Discussion: lateral flow assay for Cryptococcus is recommended for the early diagnosis of cryptococcosis in the HIV/AIDS population. Conclusion: early detection of circulating Cryptococcus antigen by lateral flow assay in HIV+ patients allowed the establishment of timely prophylaxis, follow-up, and control to reduce mortality associated with meningeal cryptococcosis.

Comparison of two commercial tests (Immy vs. Dynamiker) for cryptococcal capsular antigen

Abstract INTRODUCTION Lateral flow assay is an advanced method useful in the early diagnosis of cryptococcal meningitis. We aimed to compare two commercial tests for cryptococcal capsular antigen in the sera of asymptomatic patients with human immunodeficiency virus in Barranquilla, Colombia. METHODS Thawed (n=162) previously collected serums (2016-2019) were processed using IMMY and Dynamiker cryptococcal antigen lateral flow assay. RESULTS Compared to IMMY's results, Dynamiker's sensitivity, specificity, positive predictive value, negative predictive value, and kappa index were 100%, 89.9%, 48.3%, 100.0%, and 0.61, respectively. CONCLUSIONS The Dynamiker test had excellent sensitivity, acceptable specificity, and a low detection threshold for cryptococcal antigen in the tested samples.

Novel ractopamine–protein carrier conjugation and its application to the lateral flow strip test for ractopamine detection in animal feed / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

In this work, a novel conjugate of ractopamine and bovine serum albumin (RAC–BSA) has been developed via the Mannich reaction, with a mole coupling ratio for RAC–BSA of 9:1. The proposed conjugation method provides a simple and one-step method with the use of fewer reagents compared with other conjugation methods for competitive immunoassays. RAC–BSA conjugation was used to fabricate a competitive lateral flow strip test for RAC detection in animal feed. For sample preparation, RAC was spiked in swine feed purchased from the local markets in Thailand, and methanol and running buffer at a volume ratio of 10:90 was used as extraction buffer. The procedures for sample preparation were completed within 25 min. Under optimal conditions, the limit of detection (LOD), assessed by the naked eye within 5 min, was found to be 1 ng/g. A semi-quantitative analysis was also conducted using a smart phone and computer software, with a linearity of 0.075–0.750 ng/g, calculated LOD of 0.10 ng/g, calculated limit of quantitation of 0.33 ng/g, and good correlation of 0.992. The recoveries were found in the range of 96.4%–103.7% with a relative standard deviation of 2.5%–3.6% for intra- and inter-assays. Comparison of the results obtained by the strip test with those obtained by enzyme-linked immunosorbent assay had a good agreement in terms of accuracy. Furthermore, this strip test exhibited highly specific RAC detection without cross reactivity with related compounds. Therefore, the RAC–BSA conjugation via the Mannich reaction can be accepted as a one-step and easy conjugation method and applied to the competitive lateral flow strip test.

Novel ractopamine-protein carrier conjugation and its application to the lateral flow strip test for ractopamine detection in animal feed / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

In this work, a novel conjugate of ractopamine and bovine serum albumin (RAC-BSA) has been developed via the Mannich reaction, with a mole coupling ratio for RAC-BSA of 9:1. The proposed conjugation method provides a simple and one-step method with the use of fewer reagents compared with other conjugation methods for competitive immunoassays. RAC-BSA conjugation was used to fabricate a competitive lateral flow strip test for RAC detection in animal feed. For sample preparation, RAC was spiked in swine feed purchased from the local markets in Thailand, and methanol and running buffer at a volume ratio of 10:90 was used as extraction buffer. The procedures for sample preparation were completed within 25 min. Under optimal conditions, the limit of detection (LOD), assessed by the naked eye within 5 min, was found to be 1 ng/g. A semi-quantitative analysis was also conducted using a smart phone and computer software, with a linearity of 0.075-0.750 ng/g, calculated LOD of 0.10 ng/g, calculated limit of quantitation of 0.33 ng/g, and good correlation of 0.992. The recoveries were found in the range of 96.4%-103.7% with a relative standard deviation of 2.5%-3.6% for intra- and inter-assays. Comparison of the results obtained by the strip test with those obtained by enzyme-linked immunosorbent assay had a good agreement in terms of accuracy. Furthermore, this strip test exhibited highly specific RAC detection without cross reactivity with related compounds. Therefore, the RAC-BSA conjugation via the Mannich reaction can be accepted as a one-step and easy conjugation method and applied to the competitive lateral flow strip test.

Development of an Internally Controlled Reverse Transcription Recombinase-aided Amplification Assay for the Rapid and Visual Detection of West Nile Virus / 生物医学与环境科学（英文）

West Nile virus (WNV) causes West Nile fever and West Nile encephalitis. Because infection by WNV creates serious public health problems, its simple, rapid, and visual detection is very important in clinical practice, especially in resource-limited laboratories. We have developed a rapid, specific, and highly sensitive internally controlled reverse transcription recombinase-aided amplification (RTRAA) assay to detect WNV, using both real-time fluorescence and the lateral flow dipstick (LFD) at 39.0 °C for 30 min. The analytical sensitivity of the RT-RAA assay was 10 plasmid copies and 1.6 pfu per reaction with real-time fluorescence, and 1,000 plasmid copies per reaction with the LFD. No crossreaction with other control viruses was observed. Compared with the RT-qPCR assay, the RT-RAA assay demonstrated 100% sensitivity and 100% specificity for WNV.

Optimization of ultra-fast convection polymerase chain reaction conditions for pathogen detection with nucleic acid lateral flow immunoassay

Recently, the importance of on-site detection of pathogens has drawn attention in the field of molecular diagnostics. Unlike in a laboratory environment, on-site detection of pathogens is performed under limited resources. In this study, we tried to optimize the experimental conditions for on-site detection of pathogens using a combination of ultra-fast convection polymerase chain reaction (cPCR), which does not require regular electricity, and nucleic acid lateral flow (NALF) immunoassay. Salmonella species was used as the model pathogen. DNA was amplified within 21 minutes (equivalent to 30 cycles of polymerase chain reaction) using ultra-fast cPCR, and the amplified DNA was detected within approximately 5 minutes using NALF immunoassay with nucleic acid detection (NAD) cassettes. In order to avoid false-positive results with NAD cassettes, we reduced the primer concentration or ultra-fast cPCR run time. For singleplex ultra-fast cPCR, the primer concentration needed to be lowered to 3 µM or the run time needed to be reduced to 14 minutes. For duplex ultra-fast cPCR, 2 µM of each primer set needed to be used or the run time needed to be reduced to 14 minutes. Under the conditions optimized in this study, the combination of ultra-fast cPCR and NALF immunoassay can be applied to on-site detection of pathogens. The combination can be easily applied to the detection of oral pathogens.

Evaluación del Aspergillus lateral flow device para el diagnóstico de aspergilosis invasora, experiencia en un hospital universitario / Evaluation of the lateral flow Aspergillus assay for the diagnosis of invasive aspergillosis, experience in a university hospital

Resumen Introducción: El diagnóstico de aspergilosis invasora (AI) se realiza mediante criterios clínicos y microbiológicos los que incluyen marcadores séricos. Recientemente, el test inmunocromatográfico Aspergillus lateral flow device (LFD), ha sido evaluado como método para diagnóstico de AI. Objetivo: Evaluar el desempeño de este test para el diagnóstico de AI. Material y Método: Estudio transversal en que se evaluaron muestras de suero y lavado bronco-alveolar (LBA) procesadas para galactomanano provenientes de pacientes adultos con sospecha de AI, atendidos en el Hospital Clínico de Red de Salud UCCHRISTUS. Resultados: Se procesó un total de 142 muestras de 98 pacientes, correspondientes a AI probada 5,6%, AI probable 41,5%, AI posible 12,7% y ausencia de AI 40,1%. Al confrontar los resultados con las categorías diagnósticas según criterios EORTC/MSG se obtuvo una sensibilidad y especificidad de LFD para diagnóstico de AI de 70,9 y 53,5% para muestras de suero y 83,3 y 38,5% para muestras de LBA. La concordancia entre galactomanano y LFD fue de 62,4% (54,1-69,9) con un índice Kappa de 0,202 (0,03682-0,3669). Conclusiones: Aspergillus LFD presentó una adecuada sensibilidad; sin embargo, la especificidad fue baja por lo que un resultado positivo requiere ser confirmado.

Background: The incidence of invasive aspergillosis is increasing. Its diagnosis is based on clinical and microbiological criteria which include the determination of serological markers such as galactomannan. Recently, the Aspergillus lateral flow device, an inmunocromatograph assay has been described for its diagnosis. Aim: To evaluate the performance of the lateral flow device for the diagnosis of invasive aspergillosis (IA) in adult patients. Material and Method: In this cross-sectional study, frozen samples that had been previously evaluated for galactomannan from patients classified with proven/probable/possible or no AI according to the EORTC/MSG criteria were selected. Results: A total of 142 samples from 98 patients were processed, corresponding to proven AI 5.6%, probable IA 41.5%, possible IA 12.7% and no-IA 40.1%. The sensitivity and specificity of the Aspergillus lateral flow was 70.9% and 53.5% for serum samples and 83.3% and 38.5% for BAL samples. The concordance between the galactomannan and Aspergillus lateral flow was 62.4% (54.1 - 69.9) with a Kappa index of 0.202 (0.03682 - 0.3669). Conclusions: We observed a good sensitivity but low specificity, a positive result need a confirmatory test.

Development of a Rapid Automated Fluorescent Lateral Flow Immunoassay to Detect Hepatitis B Surface Antigen (HBsAg), Antibody to HBsAg, and Antibody to Hepatitis C

BACKGROUND: Accurate, rapid, and cost-effective screening tests for hepatitis B virus (HBV) and hepatitis C virus (HCV) infection may be useful in laboratories that cannot afford automated chemiluminescent immunoassays (CLIAs). We evaluated the diagnostic performance of a novel rapid automated fluorescent lateral flow immunoassay (LFIA). METHODS: A fluorescent LFIA using a small bench-top fluorescence reader, Automated Fluorescent Immunoassay System (AFIAS; Boditech Med Inc., Chuncheon, Korea), was developed for qualitative detection of hepatitis B surface antigen (HBsAg), antibody to HBsAg (anti-HBs), and antibody to HCV (anti-HCV) within 20 minutes. We compared the diagnostic performance of AFIAS with that of automated CLIAs—Elecsys (Roche Diagnostics GmbH, Penzberg, Germany) and ARCHITECT (Abbott Laboratories, Abbott Park, IL, USA)—using 20 seroconversion panels and 3,500 clinical serum samples. RESULTS: Evaluation with the seroconversion panels demonstrated that AFIAS had adequate sensitivity for HBsAg and anti-HCV detection. From the clinical samples, AFIAS sensitivity and specificity were 99.8% and 99.3% for the HBsAg test, 100.0% and 100.0% for the anti-HBs test, and 98.8% and 99.1% for the anti-HCV test, respectively. Its agreement rates with the Elecsys HBsAg, anti-HBs, and anti-HCV detection assays were 99.4%, 100.0%, and 99.0%, respectively. AFIAS detected all samples with HBsAg genotypes A-F and H and anti-HCV genotypes 1, 1a, 1b, 2a, 2b, 4, and 6. Cross-reactivity with other infections was not observed. CONCLUSIONS: The AFIAS HBsAg, anti-HBs, and anti-HCV tests demonstrated diagnostic performance equivalent to current automated CLIAs. AFIAS could be used for a large-scale HBV or HCV screening in low-resource laboratories or low-to middle-income areas.

A novel carbon nanoparticle probe-based ultrasensitive lateral flow assay for rapid detection of Ebola virus / 生物工程学报

Ebola virus (EBOV) is an extremely contagious pathogen first discovered in Africa associated with severe hemorrhagic disease in humans and nonhuman primates, which has resulted in at least 28 500 suspected cases and 11 300 confirmed deaths in 2014-2016 Ebola epidemic in West Africa. Rapid and sensitive detection of EBOV is the key to increasing the probability of survival and reducing infection rates in pandemic regions. Here, we report an ultrasensitive and instrument-free EBOV detection assay based on colloidal carbon immunochromatography. Carbon nanoparticle-labeled rabbit anti-EBOV-VP40 IgG were concentrated in the conjugate pad, monoclonal antibody (McAb, 4B7F9) against EBOV-VP40 and goat anti-rabbit IgG were immobilized on the nitrocellulose membrane with 2 μL/cm at a concentration of 1 mg/mL as test and control lines, respectively. Then the sample application pad, conjugate release pad, nitrocellulose membrane and absorbent pad were assembled into a lateral flow test strip. The test strip shows strong specificity against related viruses that share similar clinical symptoms and geographic range with EBOV, including marburg virus, influenza virus, yellow fever virus and dengue virus. In addition, 1 500 negative serums were tested with false-positive rate of 1.3‰ which significantly lower than that of ReEBOV™ colloidal gold test kit recommended by World Health Organization (WHO). The sensitivity of this strip was analyzed using inactivated EBOV with detection limit of 100 ng/mL (10⁶ copies/mL) which clearly higher than that of ReEBOV™ dipstick (10⁸ copies/mL). Furthermore, the strip showed excellent thermal stability characteristics in room temperature and could be as a point-of-care (POC), ultra-sensitive and specific promising candidate for EBOV serological screening in rural Africa or entry/exit ports.

Research advance of lateral flow assay labels / 生物医学工程学杂志

Lateral flow assay is widely used in the point-of-care testing on-site and in-home testing with the advantage of being simple, rapid, sensitive and cost-effective. Proper labels are the key factors in lateral flow assay. Traditional labels include colloidal gold, selenium nanoparticle, and carbon nanoparticle, among which the colloidal gold is most commonly used. Lateral flow assay has been improved as a result of the discovery of new labels, such as quantum dots and nanozyme recently. Meanwhile, transformation of qualitative detection to quantitative detection is gradually realized. This article aims at introducing the most often used and the latest lateral flow assay labels, providing a basis theoretical investigation on screening proper labels for lateral flow assay researchers.

Rapid visual detection of Mycobacterium avium subsp. paratuberculosis by recombinase polymerase amplification combined with a lateral flow dipstick

Paratuberculosis (Johne's disease) is a chronic debilitating disease of domestic and wild ruminants. However, widespread point-of-care testing is infrequent due to the lack of a robust method. The isothermal recombinase polymerase amplification (RPA) technique has applied for rapid diagnosis. Herein, RPA combined with a lateral flow dipstick (LFD) assay was developed to estimate DNA from Mycobacterium avium subsp. paratuberculosis. First, analytical specificity and sensitivity of the RPA-nfo primer and probe sets were assessed. The assay successfully detected M. paratuberculosis DNA in 30 min at 39℃ with a detection limit of up to eight copies per reaction, which was equivalent to that of the real-time quantitative polymerase chain reaction (qPCR) assay. The assay was specific, as it did not amplify genomes from five other Mycobacterium spp. or five pathogenic enteric bacteria. Six hundred-twelve clinical samples (320 fecal and 292 serum) were assessed by RPA-LFD, qPCR, and enzyme-linked immunosorbent assay, respectively. The RPA-LFD assay yielded 100% sensitivity, 97.63% specificity, and 98.44% concordance rate with the qPCR results. This is the first report utilizing an RPA-LFD assay to visualize and rapidly detect M. paratuberculosis. Our results show this assay should be a useful method for the diagnosis of paratuberculosis in resource-constrained settings.