Padronização de um ELISA indireto a partir da utilização de uma proteína de superfície do vírus da anemia infecciosa equina como antígeno para o diagnóstico dessa enfermidade / Standardization of an indirect ELISA using a surface protein of the equine infectious anemia virus as an antigen for the diagnosis of this disease

A anemia infecciosa equina é uma importante enfermidade que acomete os equídeos em todo o mundo, se apresentando de forma aguda, crônica e assintomática causando grandes prejuízos para a economia tanto para criadores que vivem do trabalho desses animais quantos aos criadores que investem no melhoramento das raças, impedindo o acesso ao mercado tanto nacional quanto internacional. O Ministério da Agricultura, Pecuária e Abastecimento considera o IDGA como teste oficial para diagnóstico dessa enfermidade, porém essa técnica é demorada e muita vez acaba sendo subjetiva, dependendo da experiencia particular de cada Laboratorista. Além de não conseguir detectar animais no início da infecção. Logo, a necessidade de se buscar novas técnicas como o ELISA indireto que aperfeiçoem o tempo de análise dos resultados, facilita a automação e obtém resultados confiáveis. O estudo realizado teve como objetivo padronizar uma técnica de ELISA indireto utilizando uma proteína de envelope viral GP90 como antígeno para diagnóstico da anemia infecciosa equina. Avaliando o desempenho do teste a partir da sensibilidade, especificidade e valores preditivos positivo e negativo. Os valores obtidos foram: 91,11%, 93,33%, 91,11% e 93,33% respectivamente. Concluiu-se que o teste apresenta bom desempenho, além da possibilidade de detectar amimais positivos no início da infecção.

Equine infectious anemia is an important disease that affects horses all over the world, presenting in an acute, chronic and asymptomatic way, causing great damage to the economy, both for breeders who live off the work of these animals and for breeders who invest in the improvement of breeds, preventing access to both national and international markets. The Ministry of Agriculture, Livestock and Food Supply considers AGID to be the official test for diagnosing this disease, but this technique takes time and often ends up being subjective, depending on the particular experience of each laboratory worker. In addition to not being able to detect animals at the beginning of the infection. Therefore, the need to seek new techniques such as indirect ELISA that improve the time of analysis of results, facilitate automation and obtain reliable results. The aim of this study was to standardize an indirect ELISA technique using a GP90 viral envelope protein as an antigen for the diagnosis of equine infectious anemia. Evaluating test performance based on sensitivity, specificity and positive and negative predictive values. The values obtained were 91.11%, 93.33%, 91.11 and 93.33 respectively. It was concluded that the test performs well, in addition to the possibility of detecting positive animals at the beginning of the infection.

Application of LAg-avidity EIA in the estimate of HIV-1 incidence among key surveillance populations in Hubei Province / 公共卫生与预防医学

Objective Limiting antigen avidity enzyme immunoassay (LAg-Avidity EIA) was applied to calculate the HIV-1 incidence of new HIV infections among men who have sex with men(MSM), drug users(DUS), female sex workers(FSW) and male patients in sexually transmitted diseases clinics(STD) in 2017-2020 of Hubei. Methods HIV-1 antibody positive samples were collected from four key groups in the national HIV sentinel surveillance from 2017 to 2020, and tested by the LAg-Avidity EIA to estimate the annual new infection rate of each group. Results The prevalence of MSM was 2.90%, 4.48%, 3.63 and 2.78% from 2017-2020, respectively, which was the highest among the four groups. The rate of late diagnosis increased from 16.67% in 2017 to 28.81% in 2020.The incidence of MSM was 1.64%(0.85%-2.44%),1.41%(0.59%-2.23%),1.46% (0.71%-2.21%) and 0.96%(0.34%-1.57%), respectively, and the difference was not statistically significant (χ2=1.491, P=0.684). The prevalence of STD from 2017-2020 was 0.03%, 0.01%, 0.06% and 0.08%, respectively, with the incidence was 0.09% (-0.06%-0.25%), 0.10(-0.06%-0.26%), 0.26% (0.02%-0.50%) and 0.04%(-0.07%-0.15%). The prevalence and incidence among FSW remains very low, while the prevalence of DUS fluctuated slightly, but no new infections have been detected. Conclusion The incidence of new HIV infection among MSM in Hubei province from 2017-2020 is relatively high compared with other key groups, but the overall trend is declining, thus the intervention of MSM should be further strengthened and expand the coverage of testing in key populations.

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.

Comparison of samples found positive by anti-HCV screening test with line immunoassay and determination of threshold value

SUMMARY OBJECTIVE: This study aimed to compare the serum samples found reactive (≥1-≤20 signal-to-cutoff ratio) with Elecsys antibodies to hepatitis C virus screening test with innogenetics-line immunassay hepatitis C Virus Score test and to determine the most appropriate threshold value for our country, since positive results close to the cutoff value cause serious problems in routine diagnostic laboratories. METHODS: Antibodies to hepatitis C virus-positive samples from 687 different patients were included in the study. Antibodies to hepatitis C virus antibody detection was performed using Elecsys antibodies to hepatitis C virus II kits (Roche Diagnostics, Germany), an electrochemiluminescence method based on the double-antigen sandwich principle, on the Cobas e601 analyzer (Roche Diagnostics) in accordance with the recommendations of the manufacturer. Samples that were initially identified as reactive were studied again. Samples with ≥1-≤20 signal-to-cutoff ratio reagents as a result of retest were included in the study to be validated with the third-Generation Line immunassay kit (innogenetics-line immunassay hepatitis C Virus, Belgium). RESULTS: A total of 687 samples with antibodies to hepatitis C virus positive and levels between 1-20 S/Co were found to be 56.1% negative, 14.8% indeterminate, and 29.1% positive by innogenetics-line immunassay hepatitis C Virus confirmation test. When the cases with indeterminate innogenetics-line immunassay hepatitis C Virus test results were accepted as positive, the signal-to-cutoff ratio value for antibodies to hepatitis C virus was determined as 5.8 (95% confidence interval) in distinguishing the innogenetics-line immunassay hepatitis C Virus negative and positive groups. CONCLUSION: It was concluded that with further studies on this subject, each country should determine the most appropriate S/Co value for its population, and thus it would be beneficial to reduce the problems such as test repetition and cost increase.

Detección de trazas de soja y de leche en productos libres de gluten: desarrollo de dos enzimoinmunoensayos competitivos / Detection of traces of soy and milk in gluten-free products: development of two competitive enzyme immunoassays / Detecção de vestígios de soja e leite em produtos livres de glúten: desenvolvimento de dois enzimoimunoensaios competitivos

Resumen El objetivo del presente trabajo fue el desarrollo de dos enzimoinmunoensayos competitivos (EIC) para la detección de trazas de soja y de leche en productos libres de gluten. Como anticuerpos primarios se utilizaron antisueros policlonales de conejo específicos contra proteínas de soja o de leche. Se determinaron las concentraciones óptimas de antígenos a inmovilizar en la placa y las concentraciones de anticuerpos primarios a utilizar en la competencia. Las curvas de calibración se ajustaron utilizando concentraciones crecientes de un extracto de producto de soja y de un extracto de leche descremada en polvo. El producto de soja y la leche descremada se extrajeron con buffer Tris-HCl 0,0625 M con dodecilsulfato de sodio al 3% y sulfito de sodio 0,1 M al 2%. Se evaluaron los parámetros de validación: linealidad, límites de detección y de cuantificación, recuperación y precisión en el día y entre días, los cuales resultaron adecuados. Se analizaron 9 productos libres de gluten con los EIC desarrollados y con kits de ELISA comerciales. Ambos EIC se comportaron de manera similar con respecto a los kits comerciales. Los EIC permitieron confirmar la presencia de leche en las muestras que la declaraban. En algunas muestras que no declaraban ni leche ni soja, ambos EIC detectaron su presencia (resultados confirmados con los kits comerciales). Los EIC desarrollados poseen menor costo que los kits y, por lo tanto, éstos podrían utilizarse como métodos de screening. Cuando esta metodología resulte negativa, debe confirmarse con un método más sensible (comercial) para garantizar la ausencia de proteínas de soja o de leche.

Abstract The aim of this study was to develop two competitive enzyme immunoassays (CEI) to detect the presence of traces of soy and milk in gluten-free products. Specific rabbit polyclonal antiserums against soy protein and other against elemilk protein were used as primary antibodies. Optimal antigen concentrations to be immobilized on the plate and primary antibody concentrations to be used in competition were determined. The calibration curves were fitted using increasing concentrations of an extract of soy product and of defatted milk powder. The soy product and the defatted milk were extracted with Tris-HCl buffer 0,0625 M with 3% sodium dodecyl sulfate and 2% sodium sulfite 0.1 M. The validation parameters were evaluated: linearity, limit of detection and quantification, recovery and precision on the day and in between days. They were appropriate. Nine commercial samples of gluten-free products were analyzed with these developed CEI and commercial ELISA kits. It was observed that both CEI behaved similarly with respect to the commercial kits. The enzyme immunoassays confirmed the presence of milk in samples that declared it. In some samples that did not declare the presence of milk or soy, both enzyme immunoassays detected their presence -these results were confirmed using commercial kits. The developed CEI have a lower cost than the commercial kits, so these could be used as screening methods. When this methodology is negative, it should be confirmed with a more sensitive (commercial) method to ensure the absence of soy or milk protein.

Resumo O objetivo do presente trabalho foi o desenvolvimento de dois enzimoimunoensaios competitivos (EIC), para a detecção de vestígios de soja e leite em produtos livres de glúten. Antissoros policlonais de coelho específicos contra proteínas de soja ou de leite foram utilizados como anticorpos primários. Foram determinadas as concentrações ótimas de antígenos a serem imobilizados na placa e as concentrações de anticorpos primários a serem utilizadas na competição. As curvas de calibração foram ajustadas usando concentrações crescentes de um extrato de produto de soja e de um extrato de leite em pó desnatado. O produto de soja e o leite desnatado foram extraídos com tampão Tris-HCl 0,0625 M com dodecil sulfato de sódio a 3% e sulfito de sódio 0,1 M a 2%. Os parâmetros de validação foram avaliados: linearidade, limite de detecção e quantificação, recuperação e precisão no dia e entre os dias, os quais resultaram adequados. Nove produtos livres de glúten foram analisados com os EIC desenvolvidos e com kits de ELISA comerciais. Os dois EICs se comportaram de maneira semelhante em relação aos kits comerciais. Os EIC permitiram confirmar a presença de leite nas amostras que o declararam. Em algumas amostras que declaravam nem leite nem soja, ambos os EIC detectaram sua presença (resultados confirmados usando kits comerciais). Os EIC desenvolvidos têm um custo menor que os kits, portanto, eles poderiam ser utilizados como métodos de triagem. Quando esta metodologia é negativa, deve ser confirmada com um método mais sensível (comercial) para garantir a ausência de proteínasda soja ou do leite.

Epidemiological and clinical aspects of snakebites in the upper Juruá River region, western Brazilian Amazonia

This study addresses the clinical and epidemiological aspects of envenoming cases resulting from snakebites treated at a hospital in Cruzeiro do Sul, in the upper Juruá River region, western Brazilian Amazonia. The specific identity of snakes that caused the envenomings was inferred (a) from the diagnosis of patient symptoms and signs upon hospital admission, (b) by enzyme immunoassay for detection of Bothrops atrox and Lachesis muta venom from serum samples taken from patients before antivenom therapy, or (c) by direct identification of the snake, when it was brought along to the hospital or photographed. There were 133 snakebites (76.2 cases per 100,000 inhabitants) registered during one year (July 2017 to June 2018). Most snakebites (88.7%) were caused by Bothrops spp., and the rest by non-venomous snakes or dry bites. Snakebites tended to occur more often during the rainy season, coinciding with the period of greater reproductive activity of the snakes and greater availability of their prey. In addition, the increase in the water level of rivers and lakes during the rainy season tends to concentrate snakes in dry places and, thus, to increase encounters with humans. Information campaigns on prevention and first aid, specially among the most vulnerable groups (indigenous people, farmers, and children and teenagers in rural areas), and the importance of using protective equipment (boots, leggings, leather gloves) in certain high risk activities (e.g. agriculture and extractivism in forests) are fundamental for the reduction of snakebite morbidity. (AU)

Cerebrospinal fluid challenges for the diagnosis of herpes simplex infection in the central nervous system / Desafios do exame do líquido cefalorraquidiano para o diagnóstico de infecção por herpes simplex no sistema nervoso central

Abstract Herpes simplex virus (HSV) is a cause of a severe disease of the central nervous system (CNS) in humans. The demonstration of specific antibodies in the cerebrospinal fluid (CSF) may contribute to the retrospective neurological diagnosis. However, the commercial immunological tests for HSV infection are for use in serum samples. Objective: The aim of the present study was to adapt a commercial kit anti-HSV IgG used for serum samples to be performed with a CSF sample. Methods: Forty CSF specimens from 38 patients with suspected CNS HSV infection were serially diluted for detecting anti-HSV IgG by enzyme immunoassay (EIA). The same samples were also analyzed with the polymerase chain reaction (PCR). Results: The sensitivity of EIA test for HSV was 5% (dilution 1:40) and 65% (dilution 1:2) in CSF, and HSV DNA PCR was 15%. The combined analysis of EIA (dilution 1:2) and PCR increased the sensitivity up to 72.5%. The inflammatory CSF was associated with positive HSV PCR. Conclusions: We demonstrated the importance to adapt serological anti-HSV IgG EIA test for CSF assays to increase the accuracy of the analysis, considering the low concentration of specific antibodies in CSF.

Resumo O vírus herpes simples (HSV) é um dos agentes causadores de uma doença grave no sistema nervoso central (SNC) em humanos. A detecção de anticorpos específicos no líquido cefalorraquidiano (LCR) pode contribuir para o diagnóstico neurológico retrospectivo. Entretanto, os testes imunológicos comerciais são para uso em amostras de soro. Objetivo: Adaptar um kit comercial sorológico anti-HSV IgG para ser utilizado no de LCR. Metodos: Quarenta amostras de LCR de 38 pacientes com suspeita de infecção por HSV no SNC foram diluídas pesquisa de anticorpos anti-HSV IgG pelo método imunoenzimático (EIA). Além disso, as mesmas amostras também foram analisadas por reação em cadeia da polimerase (PCR). Resultados: A sensibilidade do teste EIA para o HSV consistiu em 5% (diluição 1:40) e 65% (diluição 1:2) no LCR, e o PCR do DNA do HSV, 15%. A análise combinada de EIA (diluição 1:2) e PCR aumentou a sensibilidade para 72,5%. Houve associação entre presença do LCR inflamatório e PCR positiva para HSV. Conclusões: Demonstramos a importância na adaptação previa do teste sorológico anti-HSV IgG EIA para ensaios do no LCR, a fim de aumentar a acuracia da análise, considerando a baixa concentração de anticorpos específicos no LCR.

Diagnosis of Cryptococcosis in Dogs by Latex Agglutination Test and Enzyme Immunoassay

Cryptococcus spp. is a pathogenic fungus which is an increasingly important cause of infection, particularly in the immunocompromised hosts. Diagnosis of cryptococcosis in animals can be carried out by isolation of the fungus but this requires several days to detect and identify the organism. Detection of cryptococcal antigen by latex agglutination test and enzyme immunoassay in serum is a rapid and easy method for diagnosis of cryptococcosis. In the present study, a total of 142 blood samples were collected from apparently healthy (n=89) and diseased dogs (n=53) for diagnosis of cryptococcosis. Latex agglutination test and enzyme immunosorbent assay (EIA) were carried out for the detection of cryptococcal antigen in serum. Of the 142 serum samples tested, six samples tested positive by Latex agglutination test while one sample tested positive by EIA. The sample which was positive by EIA was also positive by Latex agglutination test. The serum samples of dogs that tested positive for cryptococcal antigen were obtained from dogs suffering from symptoms like bloody faeces and vomit, emesis, chronic ear infection and discharge. Based on our findings, we conclude that the latex agglutination test in combination with the enzyme immunoassay can be used for the diagnosis of Cryptococcosis in dogs

Laboratory Diagnosis of Clostridium difficile Infection in Korea: The First National Survey

In May 2015, we conducted a voluntary online survey on laboratory diagnostic assays for Clostridium difficile infection (CDI) across clinical microbiology laboratories in Korea. Responses were obtained from 66 laboratories, including 61 hospitals and five commercial laboratories. Among them, nine laboratories reported having not conducted CDI assays. The toxin AB enzyme immunoassay (toxin AB EIA), nucleic acid amplification test (NAAT), and C. difficile culture, alone or in combination with other assays, were used in 51 (89.5%), 37 (64.9%), and 37 (64.9%) of the remaining 57 laboratories, respectively, and 23 (40.4%) of the laboratories performed all three assays. Only one laboratory used the glutamate dehydrogenase assay. Nine laboratories used the toxin AB EIA as a stand-alone assay. The median (range) of examined specimens in one month for the toxin AB EIA, NAAT, and C. difficile culture was 160 (50–2,060), 70 (7–720), and 130 (9–750), respectively. These findings serve as valuable basic data regarding the current status of laboratory diagnosis of CDI in Korea, offering guidance for improved implementation.

Performance Evaluation of Two Immunochromatographic Assays for Rotavirus Detection in Stool Specimens

BACKGROUND: Rotavirus is a major pathogen causing enteritis worldwide in children under five years of age. In recent years, immunochromatographic assay (ICA) has been widely used as a diagnostic test for rotavirus detection. This study aimed to compare and evaluate the performance of ICA-based rotavirus rapid test kits from two manufacturers. METHODS: Residual stool samples from a total of 130 children with acute enterocolitis from November 2017 to January 2018 were used. We compared the results of the two immunochromatographic methods (SD BIOLINE Rotavirus kit and GENEDIA Rotavirus Ag Rapid Test) with those of the currently used enzyme immunoassay method. RESULTS: Positive agreement, negative agreement, and total agreement rates between the SD BIOLINE rotavirus kit and the enzyme immunoassay were 98.0%, 100%, and 99.2%, respectively. Positive agreement, negative agreement, and total agreement rates between the GENEDIA Rotavirus Ag Rapid Test and the enzyme immunoassay were 96.0%, 100%, and 98.4%, respectively. CONCLUSIONS: Both rotavirus rapid test kits showed very good agreement with the conventional enzyme immunoassay. Therefore, it could be a useful test to detect rotavirus directly from stool samples in a short time.

The characteristics of recent HIV-1 infection and associated factors in Tianjin / 中华预防医学杂志

The newly reported HIV infected cases was collected, and HIV blood samples were detected to identify recent HIV infection in Tianjin during 2008-2015. Factors associated with HIV-1 infection were analyzed by the univariate and multivariate unconditional logistic regression. The recent HIV-1 infection proportion of homosexuals increased from 37.70% in 2008 to 83.68% in 2015. Those cases who aged ≤30 years (OR=1.53, 95%CI: 1.30-1.79), in han ethnic group (OR=1.40, 95%CI: 1.02-1.91), students (OR=1.79, 95%CI: 1.28-2.51) were more likely to be recent infected. The cases who had a high school education (OR=1.28, 95%CI: 1.05-1.56) or collage education (OR=1.23, 95%CI: 1.00-1.50) were more likely to be recent infected than those who had a primary school education. Compared with patients identified by hospitals, the recent HIV infections were more likely to be found through voluntary counseling and testing (VCT), STD outpatients, men who have sex with men (MSM) investigation and unpaid blood donors. Homosexual transmission has become the major route of HIV-1 recent infection in Tianjin.

Proteinograma e concentração sérica de IgG em potros, do nascimento aos trinta dias de vida, tratados com plasma / Proteinogram and serum IgG concentration in newborn foals up to thirty days of life treated with plasma

Este trabalho teve por objetivo avaliar o proteinograma e concentrações séricas de IgG (após a padronização de teste ELISA) em potros do nascimento aos trinta dias de idade, antes e depois de mamarem colostro e serem tratados com plasma por via intravenosa. Foram utilizados 20 potros e suas respectivas mães, além de quatro animais doadores de plasma. Foram colhidas amostras de sangue dos potros em cinco momentos, logo após o nascimento e antes de mamar colostro (M1), dez horas após nascimento (M2), 24 horas após nascimento e previamente administração do plasma sanguíneo (M3), 48 horas de vida e 24 horas após administração do plasma sanguíneo (M4), e 30 dias após nascimento (M5). Foram colhidos sangue e colostro das éguas progenitoras no momento do parto. A concentração de proteína total (PT) e albumina foram determinadas em analisador bioquímico, a concentração de PT também foi avaliada em refratômetro manual. O fracionamento proteico foi realizado utilizando eletroforese em gel de agarose. A densidade do colostro foi avaliada com colostrômetros de refração BRIX e de densidade específica. A concentração de IgG total de todas as amostras foi determinada por teste ELISA. Com o sistema de ELISA aqui proposto foi possível determinar concentrações de IgG em amostras de soro, plasma e colostro equino com adequada repetibilidade. A média ± desvio padrão da concentração sérica de IgG dos potros ao nascer, foi de 15±8mg/dL, com dez horas de vida foi de 2.408±608mg/dL, se manteve em níveis semelhantes até 48 horas (2.364±784mg/dL) e diminuíram significativamente aos 30 dias de vida (1.414±586mg/dL). A concentração sérica e colostral de IgG nas éguas foi de 1.746±505mg/dL e 7.714±2.619mg/dL, respectivamente. A concentração plasmática de IgG dos doadores de plasma foi de 2.026±148mg/dL. Houve correlação positiva entre as concentrações séricas de IgG e PT (r=0,69 para refratômetro e r=0,76 para bioquímico), GT (r=0,81) e gamaglobulina (r=0,85). Dez horas após o nascimento foi possível verificar a transferência de imunidade passiva, possibilitando adotar medidas profiláticas e/ou terapêuticas em haras de criação de cavalos. Considerando que a proteína total, globulinas totais e fração γ-globulina apresentam correlação com IgG, estas determinações são úteis para monitorar os potros após mamarem o colostro. Um litro de plasma administrado às 24 horas de vida não foi suficiente para aumentar as concentrações séricas de IgG, 24 horas após transfusão, em potros com adequada transferência de imunidade passiva.(AU)

The aim of this study was to evaluate serum protein and serum IgG concentrations (after a direct enzyme immunoassay test ELISA optimization) in newborns foals from birth to thirty days of life before and after colostrum consumption and intravenous treatment with plasma. Twenty foals and their respective progenitors as well as four plasma donor's horses were used. Blood samples were obtained from newborn foals at five time points, immediately after birth and before colostrum intake (M1), ten hours after birth (M2), 24 hours after birth and prior administration of blood plasma (M3), 48 hours after birth and 24 hours after plasma administration (M4), and 30 days after birth (M5). Blood and colostrum samples were collected from the progenitor mares immediately postpartum. Concentration of total protein (TP) and albumin were determined using a biochemical analyzer. The TP concentration was also measured by refractometer. Fractions of total serum protein were separated using agarose gel electrophoresis. Colostrum density was evaluated using BRIX refractometer and specific density colostrometer. Total IgG concentration was determined by an enzyme-linked immunosorbent assay. With the ELISA system proposed here it was possible to determine IgG concentrations in serum, plasma, and equine colostrum samples with adequate repeatability. Serum IgG concentration in foals at birth was 15±8mg/dL (mean ± standard deviation) raising at ten hours (2,408±608mg/dL) and remaining at similar levels up to 48 hours of life (2,364±784mg/dL), and decreasing significantly at 30 days of age (1,414±586mg/dL). Serum and colostrum IgG concentrations of mares were 1,746±505mg/dL and 7,714±2,619mg/dL, respectively. The plasma IgG concentrations from donor mares were 2,026±148mg/dL. Total protein, total globulins, and γ-globulin fraction showed correlation with IgG. Ten hours post birth was an adequate time to verify the transfer of passive immunity, allowing to adoption prophylactic and/or therapeutic measures in a horse farms. One liter of plasma administered at 24 hours of life was not sufficient to raise serum IgG concentrations in foals without passive immunity transfer failure.(AU)

Estimating HIV incidence among female sex workers and injection drug users in Dehong Prefecture, 2009-2017 / 中华预防医学杂志

Objective@#To obtain HIV incidence among injection drug users (IDU) and female sex workers (FSW) in Dehong Prefecture, Yunnan Province during 2009-2017.@*Methods@#We recruited drug users and female sex workers from all sentinel surveillance sites across Dehong Prefecture during 2009-2017. A total of 10 480 IDU and 18 126 FSW in Dehong Prefecture were recruited by fingerprint technique. Data about drug uses, commercial sexual behavior, sociodemographic characteristics was collected by structured questionnaire. HIV-positive patients who were long-term infected or with CD4+ T cell count was ≤200 were not included for further HIV incidence testing. Also, those who self-identified as on antiretroviral treatment (ART) or AIDS cases were also excluded. A total of 841 and 157 plasma specimens from IDU and FSW that met the inclusion criterion were finally included, respectively. Limiting antigen avidity enzyme immunoassay(LAg-Avidity EIA) were performed to calculate the HIV incidence among these two sub-populations.@*Results@#A total of 3 444 IDU were HIV-positive, among which 884 (25.7%) were Burmese with age of (30.4±7.7), and 2 560 were Chinese with age of (36.6±7.3). Among 228 HIV-positive FSW, 109 (47.8%) were Burmese with age of (27.1±6.3), 119 (52.5%) were Chinese with age of (29.9±11.1). For IDU, the estimated HIV incidence among Burmese in 2009-2010, 2011-2012, 2013-2014, 2015-2017 was 4.20% (95%CI: -0.55%-8.95%), 7.75% (95%CI: 2.95%-12.55%), 11.79% (95%CI: 5.38%-18.20%), 10.30% (95%CI: 5.67%-14.94%), respectively, while Chinese were 3.11% (95%CI: 1.59%-4.64%), 0.03% (95%CI: -0.03%-0.08%), 1.55% (95%CI: 0.54%-2.57%), 0.58% (95%CI: -0.06%-1.04%), respectively. In 2009-2011, 2012-2014, 2015-2017, estimated HIV incidence among Burmese FSW was 0.22% (95%CI: -0.21%-0.64%), 1.24%(95%CI: 0.15%-2.32%), 0.55%(95%CI: 0.01%-1.08%). Whereas, estimated HIV incidence among Chinese FSW was 0.62% (95%CI: 0.25%-0.98%), 0.11% (95%CI: -0.04%-0.26%), 0.22% (95%CI: 0-0.44%).@*Conclusion@#HIV incidences among Chinese IDU and FSW are on the downward trend, while Burmese IDU and FSW seem to be gaining momentum.

Progressive disseminated histoplasmosis: a systematic review on the performance of non-culture-based diagnostic tests

Abstract The diagnosis of progressive disseminated histoplasmosis is often a challenge to clinicians, especially due to the low sensitivity and long turnaround time of the classic diagnostic methods. In recent years, studies involving a variety of non-culture-based diagnostic tests have been published in the literature. We performed a systematic review by selecting studies evaluating non-culture-based diagnostic methods for progressive disseminated histoplasmosis. We searched for articles evaluating detection of antibody, antigens, as well as DNA-based diagnostic methods. A comprehensive PUBMED, Web of Science, and Cochrane Library search was performed between the years 1956 and 2016. Case reports, review articles, non-human models and series involving less than 10 patients were excluded. We found 278 articles and after initial review 18 articles were included: (12) involved antigen detection methods, (4) molecular methods, and (2) antibody detection methods. Here we demonstrate that the pursuit of new technologies is ultimately required for the early and accurate diagnosis of disseminated histoplasmosis. In particular, urinary antigen detection was the most accurate tool when compared with other diagnostic techniques.

The design of liquid level detection system by ultrasonic method on automatic enzyme ;immunoassay system / 中国医学装备

Objective:To design the liquid level detection system by ultrasonic method to meet the requirements of automatic enzyme immunoassay workstations and have a precise detection of liquid level. Methods:Because the existing sample needle should move down, liquid surface and the poor conductivity of liquid were invalid. So a method of ultrasonic liquid level detection system was designed. The control signal was launched by a microprocessor to control ultrasonic signal generator chip and the ultrasonic signal reflected when the liquid level in a test tube or microplate. Results: By receiving the reflection echo signal sent to the microprocessor for signal processing, the liquid level was detected. Conclusion: The ultrasonic method applied to the liquid level detection system to overcome the above shortcomings, improve pipetting accuracy and avoid cross pollution, so as to meet the requirement of the automatic enzyme immunoassay workstation test.

Chemiluminescence Immunoassay for Quantitative Analysis of Prostate Specific Antigen Complexed toα1-Antichymotrypsin in Human Serum / 分析化学

Eight mouse hybridoma cell lines which stably secreted monoclonal antibodies ( McAbs ) against human prostate-specific antigen-α1-antichymotrypsin complex ( PSA-ACT ) were obtained through hybridoma technique. After purification, the immunological characters of 8 McAbs were identified and classified by epitopes analysis through indirect enzyme-linked immunosorbent assay ( ELISA) . A pair of McAbs was chosen from above 8 McAbs, based on which a highly sensitive, simple and rapid chemiluminescence enzyme immunoassay ( CLEIA) was developed for determination of PSA-ACT in human serums using the lumino-H2 O2 reaction catalyzed by horseradish peroxidase ( HRP) as the chemiluminescence system. Several experiment factors such as coating buffer, coating concentration, dilution ratio of PSA-ACT-HRP complex, incubation time, immunoreaction protocol and chemiluminescence reaction time were optimized. The results showed that the linear range of the proposed method for PSA-ACT determination was 0-40 ng/mL (R2=0. 9943), with the detection limit of 0. 53 ng/mL. The inter-assay relative standard deviations (RSDs) were 4. 6%-6. 6%, and intra-assay RSDs were 5 . 7%-8 . 0%. The recoveries of PSA-ACT at three spiked levels in serum samples were 95. 4%-104. 2%. The proposed method exhibited a cross-reactivity of 0. 6% with free-PSA. The proposed method is stable, sensitive, rapid and simple, and provides a foundation for the development of PSA-ACT CLEIA kit and shows great value in clinical auxiliary diagnosis of prostate cancer.

Establishment of ELISA screening method in blood donors with IgA deficiency / 国际检验医学杂志

Objective To establish the enzyme‐linked immuno sorbent assay(ELISA) method for a large‐scale screening of IgA deficiency in blood donors .Methods The indirect ELISA was adopted .The goat anti‐human IgA antibody was coated in microwell plates and labeled by horse radish peroxidase (HRP) as enzyme‐labelled secondary antibody .Results The sensitivity of established ELISA detection method was 0 .1 μg/mL .The intraassay coefficients of variation (CV) for IgA concentrations of 0 .1 ,100 μg/mL were 1 .74% to 3 .49% .The median interassay CV was 3 .48% (range:1 .83% -6 .96% ) .The assay process was 80 min . Conclusion TheELISA detection method is successfully established with high sensitivity ,strong specificity ,timesaving and easy operating and can be used for a large scale screening of Ig A deficiency and establishment of blood donors bank of Ig A deficiency .

The analysis on dynamic change and severity of IL-1 beta, TNF alpha, cox-2 detected by automatic enzyme immunoassay analyzer on patients with acute gout / 中国医学装备

Objective:To analyze dynamic change and severity of IL-1 beta, TNF alpha, COX-2 detected by automatic enzyme immunoassay analyzer on patients with acute gout.Methods: Ninety cases of acute gout patients selected according to the different degrees of severity, and were divided into three observed groups. Thirty health people were treated as the control group. IL-1β, TNF-α, COX-2 levels of dynamic change were recorded and compared within the four groups of patients.Results: The serum IL-1β, TNF-α, COX-2 levels in the 1st, 3rd, 7th, 10th day in the observation group 1 and group 2 in acute gout attacks were significantly higher than the control group; The serum levels of IL-1β, TNF-α, COX-2 levels in the 7th 10th day in observation group 1 in acute gout was significantly higher than observation group 2; The serum IL-1β, TNF-α,COX-2 levels in observation group 3 have not significant difference with control group 2; The serum IL-1β, TNF-α, COX-2 levels in observation group1,2,3 in acute gout attacks in the 14th days have not significant difference with control group.Conclusion: After analgesic drugs for the treatment of acute gout patients from 10 to 14 days, the inflammatory cytokines can be completely back to normal. The results provided important evidence for the introduction of analgesic drugs in patients with acute gout treatment.

Establishment of chemiluminescent enzyme immunoassay for detecting antibodies against foot-and-mouth disease virus serotype O in swine / 生物工程学报

Recombinant structural protein VP1 of foot-and-mouth disease virus serotype O was expressed in Escherichia coli and then purified using Nickel affinity chromatography. A chemiluminescent enzyme immunoassay (CLEIA) method was established using the purified recombinant protein as coating antigen to detect antibody of foot-and-mouth disease virus serotype O in swine. The specificity of VP1-CLEIA method is 100%. The coefficients of variation in the plate and between plates are 1.10%-6.70% and 0.66%-4.80%, respectively. Comparing with the commercial indirect ELISA kit or liquid phase block ELISA kit, the calculated coincidence rate is 93.50% or 94.00%. The high specificity and stability suggested this detection method can be used to monitor the antibody level of foot-and-mouth disease virus serotype O in swine.

Fecal cortisol metabolites as indicators of stress in crab-eating-fox (Cerdocyoun thous) in captivity / Metabolitos fecais de cortisol como indicadores de estresse em cachorro-do-mato (Cerdocyoun thous) em cativeiro

Blood samples collection is a common method in biological research using domestic animals. However, most blood sampling techniques are complicated and highly invasive and may therefore not be appropriate for wildlife animals in research concerning stress. Thus, a non-invasive method to measure steroid hormones is critically needed. The first goal of this study was to determine how glucocorticoids concentrations are impacted by translocation and reproductive activity in crab-eating-fox (Cerdocyoun thous) in captivity. The physiological relevance of fecal glucocorticoid metabolites was further validated by demonstrating: (1) The translocation of a male to a females enclosure resulted in a 3.5-fold increase compared to baseline concentrations, (2) changes in adrenocortical activity, as reflected in concentrations of fecal cortisol metabolites during reproduction, gestation and lactation in females foxes, indicating that social interactions resulted in large increases of fecal glucocorticoids metabolites during the reproductive season. From these findings we conclude that fecal samples can be used for the non-invasive assessment of adrenocortical status in crab-eating-fox...

Coleta de sangue é um método comumente utilizado na pesquisa com animais domésticos. Entretanto, a técnica de coleta de sangue torna-se complicada e altamente invasiva em animais selvagens devido ao estresse, tornando-a inapropriada para pesquisa. Dessa maneira, métodos não invasivos utilizados na mensuração de hormonios tornam-se necessários. O principal objetivo deste estudo foi determinar como as concentrações de glucocorticoides atuam durante a translocação e a atividade reprodutiva de cachorro-do-mato (Cerdocyoun thous) em cativeiro. A relevância fisiológica da análise de metabolitos fecais de glucocorticoides pôde ser validada pela demonstração de que: (1) A translocação de machos para o recinto de fêmeas resultou em um aumento de 3.5 vezes comparado a concentrações basais; (2) mudanças na atividade adrenocortical, como reflexo das concentrações de metabolitos de cortisol fecal durante a reprodução, gestação e lactação em femeas de cachorro-do-mato (Cerdocyoun thous), indicaram que interações sociais resultaram em aumento de glucocorticoides fecais durante a época reprodutiva. Com estas constatações podemos concluir que amostras fecais podem ser usadas para acesso não invasivo da atividade adrenocortical em cachorro-do-mato (Cerdocyoun thous)...