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1.
Medicine (Baltimore) ; 99(5): e18864, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32000390

RESUMO

Nuclear factor-κB-inducing kinase (NIK) is a new regulator of nuclear factor-κB signaling, which plays an important role in tumorigenesis. This study aimed to examine the expression of NIK in gastric cancer and investigate its clinical significance.Tumor issues were collected from 80 gastric cancer patients who received surgery and the diagnosis was confirmed by postoperative pathological analysis. The expression of NIK in gastric cancer tissues and adjacent normal mucosa was detected by immunohistochemical analysis. The associations between NIK expression and clinicopathological features of the patients were further analyzed.NIK expression was significantly higher in gastric cancer tissues than in adjacent normal tissues (P < .05). Furthermore, NIK expression showed significant association with UICC stage, T status, and differentiation, but not with age and gender of gastric cancer patients.NIK is overexpressed in gastric cancer and is a potential diagnostic marker of gastric cancer.


Assuntos
Proteínas Serina-Treonina Quinases/metabolismo , Neoplasias Gástricas/enzimologia , Idoso , Biomarcadores Tumorais/metabolismo , Feminino , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia
2.
Allergol. immunopatol ; 48(1): 84-89, ene.-feb. 2020. tab, graf
Artigo em Inglês | IBECS | ID: ibc-186597

RESUMO

Background and objectives: Vitamin D status may be related to allergen sensitizations, but the evidence is inconsistent. The objective of this study was to assess whether serum 25-hydroxyvitamin D (25(OH)D) levels were associated with allergic sensitizations in early childhood. Methods: Data were collected from 2642 children who visited the Guangdong Women and Children's Hospital from January 2016 to May 2017 for routine health check-ups. Serum 25(OH)D levels were tested by electrochemiluminescence immunoassay. Allergic sensitizations including food and inhalant allergens were tested for specific IgE antibodies at one year (12 months 0 days through 12 months 30 days) and two years (24 months 0 days through 24 months 30 days) of age. Results: The mean level of serum 25(OH)D was 86.47 ± 27.55 nmol/L, with a high prevalence of vitamin D insufficiency (< 75 nmol/L) in children aged 0-2 years (36.8%). Lower 25(OH)D levels with serum total IgE of more than 200IU/mL (81.54 ± 25.53 nmol/L) compared with less than 100 IU/mL (87.92 ± 28.05 nmol/L). The common sensitization to allergens in children aged one and two years were milk (44.2%), cat epithelium (26.4%), egg (13.1%), dog epithelium (12.7%) and Dermatophagoides farinae (6.7%). After multivariate adjustment, data in 25(OH)D treated as a continuous variable or categories, no consistent associations were found between 25(OH)D levels and allergen-specific IgEs. Conclusions: Serum 25(OH)D level showed an inverse relationship with total IgE level in early childhood. However, there is lack of evidence to support associations between low 25(OH)D levels and allergic sensitization to various allergens


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Assuntos
Humanos , Masculino , Feminino , Lactente , Pré-Escolar , Vitamina D/análogos & derivados , Imunização/métodos , Imunoglobulina E/sangue , Hipersensibilidade/diagnóstico , Técnicas Imunoenzimáticas , Imunoglobulina E/imunologia , China
3.
Anal Bioanal Chem ; 412(8): 1723-1728, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32030492

RESUMO

Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM-5) assays are employed in routine clinical settings to diagnose tumor. We selected two nanobodies with high-affinity to CEACAM-5, termed Nb11C12 and Nb2D5, using phage-display technology. The Nb2D5 fused with calf intestinal alkaline phosphatase (CAP), human placental alkaline phosphatase (HAP), or Pyrococcus abyssi alkaline phosphatase (PAP) were expressed in human embryonic kidney (HEK293) cells. The enzymatic activity of Nb2D5-HAP fusion protein was the best and remained stable at 60 °C for 7 days. The affinity of Nb2D5-HAP fusion protein to CEACAM-5 reached 42 pM. A chemiluminescent enzyme immunoassay (CLEIA) based on Nb2D5-HAP fusion protein was established for quantitative CEACAM-5 assay in clinical settings. The CLEIA exhibited a wide linear range of 0.31-640 ng/mL toward CEACAM-5, with a limit of detection (LOD) of 0.85 ng/mL. No cross-reactivity occurred with CEACAM-1, CEACAM-3, CEACAM-6, or CEACAM-8, and no interference was observed with rheumatoid factors. The CLEIA based on Nb2D5-HAP fusion protein was stable for 8 weeks at 37 °C and 50% relative humidity. The CLEIA developed from Nb2D5-HAP fusion protein had much better stability and linearity with similar reproducibility compared with the enzyme-linked immunosorbent assay developed from conventional monoclonal antibodies, which have been widely used in clinics over the past several decades. Graphical abstract.


Assuntos
Fosfatase Alcalina/metabolismo , Antígeno Carcinoembrionário/metabolismo , Técnicas Imunoenzimáticas/métodos , Proteínas Recombinantes de Fusão/metabolismo , Anticorpos de Domínio Único , Antígeno Carcinoembrionário/imunologia , Proteínas Ligadas por GPI/imunologia , Células HEK293 , Humanos , Limite de Detecção , Luminescência , Reprodutibilidade dos Testes
4.
Pol J Vet Sci ; 22(4): 717-723, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31867929

RESUMO

Pseudorabies (PR) outbreaks have devastated many swine farms in several parts of China since late 2011. The outbreak-associated pseudorabies virus (PRV) variant strains exhibited some typical amino acid changes in glycoprotein E (gE), a diagnostic antigen used for discriminating between PRV-infected and vaccinated animals (DIVA). To counteract the potential impact of epitope variations on current serological diagnostics of PRV, we produced monoclonal antibodies (mAbs) against gE protein of one representative PRV variant strain and developed a blocking immunoperoxidase monolayer assay (b-IPMA) for DIVA. The b-IPMA was based on the inhibition of binding between PRV-infected cells and mAb by PRV-specific antibodies present in clinical swine sera and was validated by comparison with a commercial PRV gpI Antibody Test Kit (IDEXX Laboratories, USA). The diagnostic sensitivity, diagnostic specificity and agreement were determined to be 99.25%, 98.18% and 99.02% respectively upon testing 509 serum samples. b-IPMA detected only PRV-specific antibodies and showed no cross- -reactivity with antibodies elicited by gE-deleted vaccine or other common swine pathogens. Thus, b-IPMA has the potential to be used for high-throughput screening of PRV-infected animals in veterinary clinics.


Assuntos
Herpesvirus Suídeo 1/imunologia , Técnicas Imunoenzimáticas/veterinária , Vacinas contra Pseudorraiva/imunologia , Pseudorraiva/prevenção & controle , Doenças dos Suínos/virologia , Animais , Anticorpos Monoclonais , Anticorpos Antivirais/sangue , China/epidemiologia , Surtos de Doenças/veterinária , Epitopos , Ligação Proteica , Pseudorraiva/diagnóstico , Pseudorraiva/virologia , Suínos , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/epidemiologia
5.
Sensors (Basel) ; 19(19)2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31575036

RESUMO

Hepatitis C virus (HCV) accounts for 15%-20% of cases of acute infection, and chronic HCV infection is developed in about 50%-80% of HCV patients. Unfortunately, due to the lack of proper medical care, difficulty in screening for HCV infection, and lack of awareness resulted in chronic HCV infection in 71 million people on a global scale, and about 399,000 deaths in 2016. It is crucial to recognize that the effective use of antiviral medicines can cure more than 95% of HCV infected people. The Global Health Sector Strategy (GHSS) aim is to reduce the new HCV infections and the HCV associated mortality by 90% and 65%, respectively. Therefore, the methods that are simple, yet powerful enough to detect HCV infections with high sensitivity, specificity, and a shorter window period are crucial to restrain the global burden of HCV healthcare. This article focuses on the technologies used for the detection of HCV in clinical specimens.


Assuntos
Hepacivirus/imunologia , Hepatite C/diagnóstico , Immunoblotting/métodos , Técnicas Imunoenzimáticas/métodos , Anticorpos Antivirais/análise , Antígenos Virais/análise , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Hepacivirus/genética , Hepatite C/imunologia , Humanos , Luminescência , Proteínas Virais/metabolismo
6.
Parasit Vectors ; 12(1): 470, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31604476

RESUMO

BACKGROUND: Transmission of vector-borne virus by insects is a complex mechanism consisting of many different processes; viremia in the host, uptake, infection and dissemination in the vector, and delivery of virus during blood-feeding leading to infection of the susceptible host. Bluetongue virus (BTV) is the prototype vector-borne orbivirus (family Reoviridae). BTV serotypes 1-24 (typical BTVs) are transmitted by competent biting Culicoides midges and replicate in mammalian (BSR) and midge (KC) cells. Previously, we showed that genome segment 10 (S10) encoding NS3/NS3a protein is required for virus propagation in midges. BTV serotypes 25-27 (atypical BTVs) do not replicate in KC cells. Several distinct BTV26 genome segments cause this so-called 'differential virus replication' in vitro. METHODS: Virus strains were generated using reverse genetics and their growth was examined in vitro. The midge feeding model has been developed to study infection, replication and disseminations of virus in vivo. A laboratory colony of C. sonorensis, a known competent BTV vector, was fed or injected with BTV variants and propagation in the midge was examined using PCR testing. Crossing of the midgut infection barrier was examined by separate testing of midge heads and bodies. RESULTS: A 100 nl blood meal containing ±105.3 TCID50/ml of BTV11 which corresponds to ±20 TCID50 infected 50% of fully engorged midges, and is named one Midge Alimentary Infective Dose (MAID50). BTV11 with a small in-frame deletion in S10 infected blood-fed midge midguts but virus release from the midgut into the haemolymph was blocked. BTV11 with S1[VP1] of BTV26 could be adapted to virus growth in KC cells, and contained mutations subdivided into 'corrections' of the chimeric genome constellation and mutations associated with adaptation to KC cells. In particular one amino acid mutation in outer shell protein VP2 overcomes differential virus replication in vitro and in vivo. CONCLUSION: Small changes in NS3/NS3a or in the outer shell protein VP2 strongly affect virus propagation in midges and thus vector competence. Therefore, spread of disease by competent Culicoides midges can strongly differ for very closely related viruses.


Assuntos
Vírus Bluetongue/fisiologia , Ceratopogonidae/virologia , Deleção de Genes , Insetos Vetores/virologia , Mutação Puntual , Animais , Vírus Bluetongue/genética , Linhagem Celular , Embrião de Galinha , Cricetinae , Cervos , Feminino , Técnicas Imunoenzimáticas , Genética Reversa , Replicação Viral , Sequenciamento Completo do Genoma
7.
MMWR Morb Mortal Wkly Rep ; 68(42): 947-952, 2019 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-31652252

RESUMO

A 2014 report evaluating accuracy of serologic testing for transfusion-transmissible viruses at African blood center laboratories found sensitivities of 92%, 87%, and 90% for detecting infections with human immunodeficiency virus (HIV), hepatitis B virus (HBV), and hepatitis C virus (HCV), respectively (1). Following substantial investments in national blood transfusion service (NBTS) laboratories, in 2017 investigators tested proficiency at 84 blood center laboratories (29 NBTS and 55 non-NBTS) in seven African countries. A blinded panel of 25 plasma samples was shipped to each participating laboratory for testing with their usual protocols based on rapid diagnostic tests (RDTs) (2) and third and fourth generation enzyme immunoassays (EIA-3 and EIA-4). Sensitivity and specificity were estimated using separate regression models that clustered assays by laboratory and adjusted for assay type and NBTS laboratory status. Mean specificities were ≥95% for all three viruses; however, mean sensitivities were 97% for HIV-positive, 76% for HBV-positive, and 80% for HCV-positive samples. Testing sensitivities for all viruses were high when EIA-3 assays were used (≥97%). Lower sensitivities for HBV-positive samples and HCV-positive samples were associated with assay types other than EIA-3, used primarily by non-NBTS laboratories. Proficiency for HIV testing has improved following international investments, but proficiency remains suboptimal for HBV and HCV testing. In sub-Saharan African blood centers, the quality of rapid tests used for HBV and HCV screening needs to be improved or their use discouraged in favor of EIA-3 tests.


Assuntos
Bancos de Sangue , HIV/isolamento & purificação , Hepacivirus/isolamento & purificação , Vírus da Hepatite B/isolamento & purificação , Programas de Rastreamento/normas , África , Testes Diagnósticos de Rotina , Humanos , Técnicas Imunoenzimáticas , Programas de Rastreamento/métodos , Sensibilidade e Especificidade , Testes Sorológicos
8.
Turkiye Parazitol Derg ; 43(3): 106-110, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31502771

RESUMO

Objective: The aim of this study was to determine the frequency of toxoplasmosis in pregnant women who were admitted to our hospital in their first trimester and to contribute to screening and management strategies by evaluating clinical outcomes. Methods: In this retrospective study, women in their first trimester of pregnancy who were aged between 15-49 years, admitted to the Mersin University Medical Faculty Gynecology and Obstetrics outpatient clinic between 2012-2017, and screened for Toxoplasma gondii immunoglobulin M (IgM) antibodies were included. The data was obtained from the hospital's digital records. First, the high-risk patients were identified who had anti-T. gondii IgM seropositivity and subsequently underwent anti-T. gondii immunoglobulin G (IgG) and anti-T. gondii IgG avidity tests. Next, the invasive procedures and medical treatments performed for diagnosis and treatment, as well as the clinical course and results for each patient were evaluated. Cases were then analyzed according to the admittance year and patient's age. Results: Anti-T. gondii IgM positivity was found in 266 (7.66%) of 3474 pregnant women meeting the study's criteria. The frequency of the Toxoplasma IgM seropositivity was the highest in the 15-25 age group and this frequency decreased gradually as the age of the patients increased. Congenital toxoplasmosis was detected in 1 of 61 patients who had a positive polymerase chain reaction for T. gondii performed in the amniotic fluid. Conclusion: In our province, the prevalence of anti-T. gondii IgM was found to be 7.66% in pregnant women who were admitted to a tertiary health institution in their first trimester of pregnancy. This rate is much higher than the average in Turkey; therefore, we suggest that routine screening of pregnant women for T. gondii may be recommended in this region.


Assuntos
Anticorpos Antiprotozoários/sangue , Complicações Parasitárias na Gravidez/epidemiologia , Toxoplasma/imunologia , Toxoplasmose/epidemiologia , Adolescente , Adulto , Afinidade de Anticorpos , Feminino , Hospitalização , Humanos , Técnicas Imunoenzimáticas , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Pessoa de Meia-Idade , Gravidez , Primeiro Trimestre da Gravidez , Prevalência , Estudos Retrospectivos , Estudos Soroepidemiológicos , Centros de Atenção Terciária , Toxoplasmose Congênita/epidemiologia , Turquia/epidemiologia , Adulto Jovem
9.
Exp Eye Res ; 188: 107807, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31539543

RESUMO

In order to study the mechanisms involved in the development of posterior capsule opacification (PCO) we compared in vivo developed PCO with PCO formed in tissue culture with focus on the periphery of the lens capsule to evaluate lens regeneration potential. We studied three human tissue groups: Cultured lens capsules after mock cataract surgery (n = 6, 30 days), lens capsules from donors that had previously undergone cataract surgery (IOL capsules) (n = 12) and intact lenses (n = 6). All samples were stained with Vimentin, alpha Smooth Muscle Actin, Picro Sirius Red (for collagen) and Paired box protein (Pax6). We found that cultured capsules and less developed IOL capsules consisted mainly of monolayers of mesenchymal cells, while more developed IOL capsules, contained lens epithelial cells (LECs), globular cells and lens fiber cells. Many IOL capsule samples expressed collagen I and III in areas where cells were in contact with the IOL. Pax6 had a similar dispersed distribution in less developed IOL capsules and cultured capsules, while more developed IOL capsules and intact lenses, concentrated Pax6 in LECs at the equatorial lens bow. The similarities between cultured capsules and less developed IOL capsules indicate that our in vitro developed PCO is comparable to early in vivo developed PCO. The similar morphology of more developed IOL capsules and intact lenses seems to indicate an attempt at lens regeneration.


Assuntos
Opacificação da Cápsula/patologia , Cápsula Posterior do Cristalino/patologia , Actinas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/metabolismo , Opacificação da Cápsula/metabolismo , Extração de Catarata , Feminino , Humanos , Técnicas Imunoenzimáticas , Implante de Lente Intraocular , Masculino , Pessoa de Meia-Idade , Técnicas de Cultura de Órgãos , Cápsula Posterior do Cristalino/metabolismo , Doadores de Tecidos , Vimentina/metabolismo
10.
Infez Med ; 27(3): 316-321, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31545776

RESUMO

Transmission of infections through blood and blood product transfusion is a serious healthcare problem. There are insufficient up-to-date data about seroprevalence of HBsAg, anti-HCV and anti-HIV ½ among healthy blood donors in Turkey. We aimed to investigate the seroprevalence of HBsAg, anti-HCV and anti-HIV ½ in Southeastern Anatolia, Turkey. HBsAg, anti-HCV, and anti-HIV ½ analysis results among blood donors who applied to Dicle University Faculty of Medicine, Diyarbakir District Blood Centre, between January 1, 2011 and December 31, 2015 were retrospectively evaluated. HBsAg, anti-HCV, and anti-HIV 1/2 screenings were performed using a fully automated device with the microparticle enzyme immunoassay method (MEIA). The chi-square (χ2) test was applied to variables. Among the donors, 1607 (1.73%) were HBsAg-positive, 255 (0.27%) were anti-HCV-positive and two (0.0021%) were positive for anti-HIV 1/2. HBsAg positivity rates by years were 2.50% in 2011, 1.92% in 2012, 1.74% in 2013, 1.53% in 2014 and 1.27% in 2015 (p<0.001). HBsAg-positivity was 0.78% for the donors between 18-24 years of age, 1.90% for those between 25-49 years of age and 3.92% for donors over the age of 49 (p<0.001). Anti-HCV positivity rates were as follows: 0.35% in 2011, 0.34% in 2012, 0.29% in 2013, 0.23% in 2014 and 0.16% in 2015 (p<0.001). Verified anti-HIV 1/2 positivity was observed for only two donors (0.0021%) within five years. HBsAg and anti-HCV positivity were observed to decrease significantly over the years and were significantly lower among younger donors.


Assuntos
Doadores de Sangue/estatística & dados numéricos , Anticorpos Anti-HIV/sangue , HIV-1/imunologia , HIV-2/imunologia , Antígenos de Superfície da Hepatite B/sangue , Anticorpos Anti-Hepatite C/sangue , Adolescente , Adulto , Distribuição por Idade , Distribuição de Qui-Quadrado , Feminino , Soroprevalência de HIV , Nível de Saúde , Hepatite C/imunologia , Humanos , Técnicas Imunoenzimáticas/métodos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Estudos Soroepidemiológicos , Distribuição por Sexo , Fatores de Tempo , Turquia , Adulto Jovem
11.
Fish Shellfish Immunol ; 94: 199-210, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499199

RESUMO

The poor understanding of nutrition needed has become a significant obstruction to artificial conservation of Yangtze sturgeon (Acipenser dabryanus) and the relationship between ployunsaturated fatty acid nutrition and the immune response of Yangtze sturgeon is remains unclear. To explore this relationship, the immune response was determined by the activities of serum immune-related enzymes and the transcriptome pattern in the spleen after feeding different fat source diets for 7 weeks. In addition, the gene expression pattern after a lipopolysaccharide (LPS) challenge was investigated in the presence of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). Long-term feeding of the fish oil diets increased the serum immune-related enzyme activities, including lysozyme, acid phosphatase, and alkaline phosphatase of Yangtze sturgeon. More than 653,999 transcripts with an N50 length of 1047 bp were obtained and a final set of 280,408 unigenes was generated. After annotating the unigenes, 3549 genes were assigned to the immune system and 2839 were identified to participate in the response to the different fat sources. A transcriptome assay showed the fish oil diets moderately upregulated immune-related signaling pathways in the spleen of Yangtze sturgeon, including NLR signaling, platelet activation, Fc gamma R-mediated phagocytosis, Th17 cell differentiation, and Th1 and Th2 cell differentiation. The quantitative polymerase chain reaction (qPCR) results of candidate genes for these pathways showed similar results. The LPS challenge study revealed that DHA and EPA moderately upregulated the candidate immune-related genes and modulated excessive activation of the immune pathway by the pathogen. This study confirmed the immunomodulatory function of unsaturated fatty acids in Yangtze sturgeon. This research will provide a reference for the preparation of artificial diets for Yangtze sturgeon.


Assuntos
Gorduras Insaturadas na Dieta/metabolismo , Ácidos Graxos Insaturados/metabolismo , Óleos de Peixe/metabolismo , Peixes/imunologia , Imunidade Inata , Animais , Gorduras Insaturadas na Dieta/administração & dosagem , Espécies em Perigo de Extinção , Ácidos Graxos Insaturados/administração & dosagem , Óleos de Peixe/administração & dosagem , Peixes/metabolismo , Perfilação da Expressão Gênica/veterinária , Técnicas Imunoenzimáticas/veterinária , Distribuição Aleatória , Reação em Cadeia da Polimerase em Tempo Real/veterinária
12.
Intervirology ; 62(3-4): 145-155, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31533104

RESUMO

BACKGROUND: When infected with the chikungunya virus (CHIKV), 3% to 28% of CHIKV-infected individuals remain asymptomatic, necessitating the development of improved high-throughput screening methods to overcome the limitations of molecular diagnostics or enzyme-linked immunosorbent assays (ELISAs). OBJECTIVE: In this study, two novel monoclonal antibodies (mAbs) targeting envelope 1 (E1) of CHIKV were developed and applied in a fluorescence-linked immunosorbent assay (FLISA) using coumarin-derived dendrimer as the fluorophore. METHODS: The performance of the FLISA was compared with that of ELISA. RESULTS: Using the two novel mAbs (2B5 and 2C8), FLISA could detect 1 × 105 PFU/mL of CHIKV, exhibiting a 2-fold lower limit of detection (LOD) compared to ELISA. The LOD of FICT corresponded to a comparative threshold value of 23.95 and 4 × 106 of RNA copy number/µL. In the presence of human sera and blood, virus detection by FLISA was 3-fold better than ELISA, with an LOD of 2 × 105 PFU/mL. Sera and blood interfered with the ELISA, resulting in 6 × 105 PFU/mL as the LOD. CONCLUSIONS: FLISA using two novel mAbs and coumarin-derived dendrimer is a superior diagnostic assay for detecting CHIKV in human sera and blood, compared to conventional ELISA.


Assuntos
Antígenos Virais/análise , Febre de Chikungunya/diagnóstico , Vírus Chikungunya/isolamento & purificação , Testes Diagnósticos de Rotina/métodos , Fluorometria/métodos , Técnicas Imunoenzimáticas/métodos , Proteínas do Envelope Viral/análise , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/isolamento & purificação , Vírus Chikungunya/imunologia , Humanos , Sensibilidade e Especificidade
13.
Sci Total Environ ; 687: 849-857, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31412488

RESUMO

Increased awareness of phthalic acid esters (PAEs) toxicity has given rise to a dramatic increase in concern about the determination of these contaminations in the environment. In this paper, a sensitive, selective and rapid enzyme immunoassay of ELISA based on polyclonal antibody for detecting butyl benzyl phthalate (BBP) was developed and applied in the environmental water and soil samples. The hapten of BBP was synthesized, then applied to prepare artificial antigen and produce polyclonal antibody capable of specific recognizing BBP. From the optimal standard curve of ELISA for BBP, the values of LOD (limit of detection, IC10) and IC50 were 2.5 and 79.4 ng/mL, respectively. The ELISA showed high specificity, with the cross-reactivity toward BBP analogs < 9.6%. The satisfactory accuracy and precision were demonstrated by the recoveries of 76-116% and coefficient of variations (CVs) of 4.7-13.7%. Furthermore, BBP contamination was investigated at 3.1-25.2 ng/mL in real water samples and 4.2-76.4 ng/g in real soil samples (with the detection rate of 55% in 20 samples) by the developed ELISA, which also had shown a good correlation with that the results obtained by HPLC. All of this indicated that the developed enzyme immunoassay could be applied for sensitive and selective determination of BBP contamination in the environmental samples. Furthermore, the strategy of BBP hapten synthesis and an alternative method of BBP determination could be provided.


Assuntos
Poluentes Ambientais/análise , Técnicas Imunoenzimáticas , Ácidos Ftálicos/análise , Cromatografia Líquida de Alta Pressão , Ensaio de Imunoadsorção Enzimática
14.
Clin Lab ; 65(8)2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31414736

RESUMO

BACKGROUND: Clostridioides (formerly Clostridium) difficile infection (CDI) is linked to misuse of antimicrobials. The prevalence of disease varies with difficulties of establishing the diagnosis because of the lack of sensitivity and specificity of laboratory tests. The clinical impact of upgrading CDI testing from routine to molecular based-algorithm is still unclear. The aim of this study is to assess the impact of upgrading CDI testing from routine to molecular based-algorithm on the management of CDI and evaluate the role of antimicrobials on the course of CDI. METHODS: This is an observational case-study. A total of 564 patients were included from whom stool samples were tested by enzyme immunoassay (EIA) and Xpert for C. difficile. Data on the number and results of tests ordered, antimicrobial exposure, comorbidities, and treatment with metronidazole or vancomycin were collected. The main outcome measures were C. difficile tests (EIA and Xpert C. difficile Assay) and prevalence of CDI. RESULTS: CDI was found in 9 and 10 cases out of 313 and 254 patients tested by the EIA and Xpert C. difficile assay, respectively, giving an overall incidence of 0.03 per 1,000 patient tested. Reduction was noted in the number of tests ordered per patient for presumptive CDI after shifting to the Xpert C. difficile assay which was not statistically significant (p-value 0.2). Also, there was less metronidazole and vancomycin therapy initiated for patients with a negative C. difficile test (p-value 0.2) observed with molecular testing. CONCLUSIONS: Xpert C. difficile testing is a supportive tool for diagnosing CDI with rapid turnaround time that is helpful for patient management and initiating effective infection control measures. The clinical accuracy of the assay is still to be determined in the context of low carriage rate in the local patient population.


Assuntos
Algoritmos , Infecções por Clostridium/diagnóstico , Clostridium difficile/isolamento & purificação , Fezes/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Técnicas de Laboratório Clínico/métodos , Infecções por Clostridium/epidemiologia , Infecções por Clostridium/microbiologia , Clostridium difficile/efeitos dos fármacos , Clostridium difficile/fisiologia , Humanos , Técnicas Imunoenzimáticas/métodos , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular/métodos , Prevalência , Estudos Prospectivos , Arábia Saudita/epidemiologia , Sensibilidade e Especificidade , Adulto Jovem
15.
Am J Case Rep ; 20: 1248-1252, 2019 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-31444319

RESUMO

BACKGROUND Few cases of falsely undetectable PSA due to the presence of an inhibitory serum factor have been reported in the world literature. We present a case of falsely low-to-undetectable PSA with data from a serum dilution series, the current literature on biochemical assay interference, and the implications for prostate cancer salvage treatment. CASE REPORT A 63-year-old man was treated with prostatectomy for high-risk prostate cancer and was found to have a rising PSA after approximately 3 years following surgery. He subsequently transferred his care to a different health system and was found to have an undetectable PSA. He was eventually found to have an elevated PSA once again after the particular assay at this institution was changed. He thus received salvage prostate radiotherapy and androgen deprivation therapy. CONCLUSIONS While falsely low PSA results cannot be explained by the presence of serum heterophile antibodies, competitive antibody interference against the immunoassay reagents or anti-PSA antibodies are possible explanations for the results of the dilution experiments performed in this case study. We suggest that unexpected PSA testing results should raise concern for assay interference and warrant further clinical workup.


Assuntos
Antígeno Prostático Específico/sangue , Reações Falso-Negativas , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/terapia , Terapia de Salvação
16.
Exp Parasitol ; 205: 107739, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31437436

RESUMO

Intestinal protozoans found in ancient human samples have been studied primarily by microscopy and immunodiagnostic assays. However, such methods are not suitable for the detection of zoonotic genotypes. The objectives of the present study were to utilize immunoenzimatic assays for coproantigen detection of Cryptosporidium sp., Giardia duodenalis, and Entamoeba histolytica/Entamoeba dispar in sixty ancient human and animal samples collected from 14 archaeological sites in South America, and to carry out a critical analysis of G. duodenalis according to results obtained from three diagnostic methodologies: microscopy, immunodiagnostic tests (immunoenzymatic and immunofluorescence), and molecular biology (PCR and sequencing). More than half (31/60) of the samples analyzed using immunoenzymatic tests were positive for at least one of the intestinal protozoans, with 46.6% (28/60) corresponding to G. duodenalis, 26.6% (16/60) to Cryptosporidium sp., and 5% (3/60) to E. histolytica/E. dispar. Cryptosporidium sp. and G. duodenalis coinfection was observed in 15% (9/60) of the samples, whereas all three protozoans were found in 5% (3/60) of samples. In the Northeast Region of Brazil, by immunoenzymatic tests there is evidence that G. duodenlais and Cryptosporidium sp. have infected humans and rodents for at least 7150 years. However, for G. duodenalis, the results from the three diagnostic tests were discordant. Specifically, despite the efficiency of the molecular biology assay in the experimental models, G. duodenalis DNA could not be amplified from the ancient samples. These results raise the following question: Are all ancient samples positive for coproantigen of G. duodenalis by immunoenzymatic tests truly positive? This scenario highlights the importance of further studies to evaluate the sensitivity and specificity of the immunoenzymatic method in the archaeological context.


Assuntos
Arqueologia/métodos , Cryptosporidium/isolamento & purificação , Entamoeba/isolamento & purificação , Fezes/parasitologia , Giardia lamblia/isolamento & purificação , Técnicas Imunoenzimáticas/normas , Animais , Antígenos de Protozoários/análise , Antígenos de Protozoários/genética , Cryptosporidium/genética , Cryptosporidium/imunologia , Entamoeba/genética , Entamoeba/imunologia , Entamoeba histolytica/genética , Entamoeba histolytica/imunologia , Entamoeba histolytica/isolamento & purificação , Giardia lamblia/genética , Giardia lamblia/imunologia , Humanos , Enteropatias Parasitárias/parasitologia , Roedores , Sensibilidade e Especificidade , América do Sul
17.
Analyst ; 144(16): 4813-4819, 2019 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-31281909

RESUMO

In the clinical diagnosis of tumor, the immunological detection of single tumor markers may lead to errors and missed inspection. Therefore, it is necessary to establish an accurate and effective method for the simultaneous detection of multiple tumor markers. Thus, we developed a time-resolved chemiluminescence immunoassay (TRCLIA) to simultaneously detect carcinoembryonic antigen (CEA) and neuron-specific enolase (NSE) in human serum. Horseradish peroxidase (HRP) and alkaline phosphatase (ALP) were used as the detection probes to label the monoclonal antibodies of CEA and NSE by strain-promoted azide-alkyne cycloaddition (SPAAC), respectively. Based on a sandwich immunoassay, the targets in the samples were captured by antibodies immobilized on the surface of carboxylate-modified polystyrene microspheres (CPSMS) and sandwiched by other antibodies labeled with HRP and ALP. Since HRP and ALP had different dynamic characteristics, the CEA and NSE signals were recorded at 0.5 s and 20 min, respectively, and cross-interference could be avoided effectively. The whole signal detection processes could be completed in 20 min. The linear ranges of CEA and NSE were 0.1-64 ng mL-1 and 0.05-64 ng mL-1 and the limits of detection were 0.085 ng mL-1 and 0.044 ng mL-1 (S/N = 2), respectively. Also, 45 human serum samples obtained from patients having lung disease were tested by TRCLIA and commercial chemiluminescence enzyme-linked immunoassay (CLEIA) kits with good correlation. The correlation coefficients of CEA and NSE were 0.985 and 0.970, respectively. The results demonstrated a novel, effective, reliable and convenient TRCLIA method for the clinical diagnosis of CEA and NSE. The TRCLIA method has the potential to be an effective clinical tool for the early screening of lung cancer and can be applied in clinical diagnosis.


Assuntos
Antígeno Carcinoembrionário/sangue , Técnicas Imunoenzimáticas/métodos , Fosfopiruvato Hidratase/sangue , Fosfatase Alcalina/química , Anticorpos Imobilizados/imunologia , Anticorpos Monoclonais/imunologia , Armoracia/enzimologia , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/imunologia , Antígeno Carcinoembrionário/imunologia , Peroxidase do Rábano Silvestre/química , Humanos , Limite de Detecção , Luminescência , Substâncias Luminescentes/química , Medições Luminescentes/métodos , Luminol/química , Pneumopatias/sangue , Fosfopiruvato Hidratase/imunologia
18.
Anatol J Cardiol ; 22(1): 13-20, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31264652

RESUMO

OBJECTIVE: Left ventricle (LV) geometry and dyssynchrony are associated with LV remodeling after acute myocardial infarction (AMI). The aim of this prospective study was to assess the diagnostic value of new three-dimensional echocardiography (3DE) parameters [sphericity (SI) and systolic dyssynchrony indexes (SDI)] for the prediction of LV remodeling after AMI and to compare them with two-dimensional echocardiography (2DE) parameters. METHODS: 2DE and 3DE were performed in 75 patients with AMI within 3 days from the onset of MI and 6 months later. LV remodeling was defined as a ≥15% increase in the LV end-diastolic volume (EDV) at follow-up. 3D SI was calculated by dividing EDV by the volume of a sphere whose diameter was derived from the major end-diastolic LV long axis. SDI was considered as a standard deviation of the time from cardiac cycle onset to minimum systolic volume in 16 LV segments. RESULTS: LV remodeling was identified in 34 (45%) patients using the 2DE method and in 22 (29%) patients using the 3DE method. Evaluated 3DE parameters, such as EDV [area under the receiver operating characteristic (ROC) curve (AUC) 0.742, sensitivity 71%, specificity 79%], end-systolic volume (AUC 0.729, sensitivity 69%, specificity 78%), SDI (AUC 0.777, sensitivity 73%, specificity 77%), and SI, had significant prognostic value for LV remodeling. According to the AUC, the highest predictive value had 3D SI (AUC 0.957, sensitivity 90%, specificity 91%). CONCLUSION: 3DE parameters, especially 3D SI and SDI, play important roles in the prediction of LV remodeling after AMI and can be used in clinical practice.


Assuntos
Ecocardiografia Tridimensional/normas , Infarto do Miocárdio/diagnóstico por imagem , Disfunção Ventricular Esquerda/prevenção & controle , Remodelação Ventricular , Idoso , Área Sob a Curva , Proteína C-Reativa/análise , Estudos de Coortes , Feminino , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/complicações , Revascularização Miocárdica , Nefelometria e Turbidimetria , Variações Dependentes do Observador , Intervenção Coronária Percutânea , Estudos Prospectivos , Curva ROC , Sensibilidade e Especificidade , Fumar , Terapia Trombolítica , Troponina I/análise , Função Ventricular Esquerda/fisiologia
19.
Vox Sang ; 114(7): 740-748, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31321786

RESUMO

BACKGROUND AND OBJECTIVES: The purpose of the study was to evaluate a lyophilized anti-D immunoglobulin preparation to serve as a replacement WHO International Standard for the calibration of potency assays of anti-D immunoglobulin products. Such products are used to prevent haemolytic disease of the foetus and newborn due to maternal alloanti-D. MATERIALS AND METHODS: The candidate 3rd International Standard for anti-D immunoglobulin (16/332) was evaluated and calibrated against the 2nd International Standard for anti-D immunoglobulin (01/572), along with a coded duplicate, a second candidate preparation (16/278) and a comparability sample (16/272) in an international collaborative study. Twenty of 21 laboratories in 15 countries performed one or more of the three European Pharmacopoeia reference methods. RESULTS: The overall geometric mean potency (from all methods) of the candidate 3rd International Standard, 16/332, was 296·6 IU/ampoule, with inter-laboratory variability, expressed as % GCV, of 4·7%. SE-HPLC of the immunoglobulin preparations demonstrated combined monomeric and dimeric IgG peak areas of >95% for all samples. Accelerated stability studies have shown both 16/332 and 16/278 to be very stable for long-term storage at -20°C. CONCLUSIONS: Preparation 16/332 was established by the World Health Organisation Expert Committee on Biological Standardization as the 3rd International Standard for anti-D immunoglobulin with an assigned potency of 297 IU/ampoule.


Assuntos
Eritroblastose Fetal/sangue , Técnicas Imunoenzimáticas/normas , Técnicas de Diagnóstico Molecular/normas , Imunoglobulina rho(D)/imunologia , Eritroblastose Fetal/imunologia , Humanos , Imunoglobulina D/imunologia , Indicadores e Reagentes/normas , Padrões de Referência , Organização Mundial da Saúde
20.
BMC Infect Dis ; 19(1): 632, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31315573

RESUMO

BACKGROUND: Hepatitis B virus (HBV) testing in oral fluid samples may provide advantages in diagnosis, screening or prevalence studies, especially among individuals with venous access difficulties. This study aims to optimize one commercially available assay for detecting total anti-HBc marker in oral fluid samples and to evaluate its utility under real life conditions in different settings for the purposes of prevalence and diagnostic studies. METHODS: Oral fluid was collected using a Salivette device and some parameters were initially evaluated: type of elution buffer and sample volume. Thereafter, the utility of oral fluid samples for detection of anti-HBc was evaluated in real life conditions in which, 1296 individuals gave serum and oral fluid samples. All serum samples were submitted to commercial EIAs to detect total anti-HBc, according to the manufacturer's instructions and oral fluid samples according to previous optimization. RESULTS: In optimization evaluation, PBS/BSA 0.5% and 100 µL of oral fluid (volume was two-fold increased compared to serum in EIA) were chosen as transport buffer and sample volume. In the field study, anti-HBc was detected in 211 out of 1296 serum samples giving overall oral fluid sensitivity of 52.6% and specificity of 96%. Concordance was higher in ambulatory setting (67.7) compared to general population (31.8). Mean ± standard deviation values of optical density/cutoff (OD/CO) in serum samples were higher in false-negative oral fluid samples than those seen in true positive samples. Sensitivity was higher in those presenting active infection compared to anti-HBc isolate and past infection. Sensitivity also increased in the ambulatory group when HCV individuals were excluded. CONCLUSIONS: It was possible to optimize a commercial EIA for detecting anti-HBc in oral fluid samples and where the highest concordance was found in ambulatory settings and among individuals with active infection.


Assuntos
Anticorpos Anti-Hepatite B/análise , Hepatite B/diagnóstico , Técnicas Imunoenzimáticas/métodos , Saliva/virologia , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade
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