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1.
Rev Inst Med Trop Sao Paulo ; 61: e48, 2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31531626

RESUMO

School-age children are a social group in which blood collection for laboratory testing can be perceived as an invasive procedure, with low acceptance and tolerance of stakeholders. This problem could be circumvented by replacing serum samples with saliva. For this purpose, and to make the collection of saliva samples playful and instructive for children, educational activities on hygiene and toxoplasmosis transmission and prevention were performed using toys and audiovisual tools. The target audience consisted of 7-10 year-old children from low-income families who attended public schools in the city of São Paulo. Saliva samples were used in a previously described in-house Enzyme-Linked Immunosorbent Assays (ELISA) to detect anti- Toxoplasma gondii IgG antibodies and establish the immunological status of each of the participants. One year later, children's memory and fixation of concepts regarding hygiene habits, as well as transmission and prevention of toxoplasmosis were tested in the same schools, by means of a questionnaire application, using students who did not participate in the first intervention as controls. The prevalence of positive anti- T. gondii IgG among students was 50% (82/164). One year later, 45 children had more knowledge on toxoplasmosis (28/45 vs 29/147) and they drew the cat's involvement in the transmission of toxoplasmosis more often than controls (28/45 vs 29/147). Sorted according to the presence of specific IgG in saliva, recovered positive students presented worse memory of the above cited knowledge as did saliva-negative IgG students, but both groups had isolated higher frequency of fixed knowledge than non-intervened students. Our data show that there is a high prevalence of T. gondii infection in school-children from low-income areas; saliva is an alternative to blood for anti- T. gondii IgG detection; and a one-day educational intervention in school-children was effective in promoting knowledge fixation on hygiene and toxoplasmosis transmission and prevention after one year.


Assuntos
Anticorpos Antiprotozoários/análise , Educação em Saúde/métodos , Conhecimentos, Atitudes e Prática em Saúde , Higiene , Imunoglobulina G/análise , Saliva/parasitologia , Toxoplasma/imunologia , Criança , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Pobreza , Inquéritos e Questionários
2.
BMC Infect Dis ; 19(1): 576, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31272486

RESUMO

BACKGROUND: Pleural parasitic infestation (PPI) is a disease prevalent in certain parts of the world. It is frequently misdiagnosed due to its lack of standardized diagnostic criteria. The purpose of this study was to evaluate the clinical characteristics of PPI patients and develop a practical diagnostic approach for PPI. METHODS: A retrospective study was conducted by reviewing the medical records of 11 patients with PPI. A practical diagnostic approach was proposed based on the unique laboratory findings. RESULTS: All patients demonstrated respiratory symptoms, including shortness of breath, cough, fever, chest pain, excessive sputum and hemoptysis. Leukocytosis (> 10,000/µL) and eosinophilia (> 500/µL) of peripheral blood were present in 45.5 and 36.4% patients, respectively. The mean concentrations of pleural effusion lactate dehydrogenase (LDH), adenosine deaminase (ADA), protein and carcinoembryonic antigen (CEA) were 338.2 U/L (range, 61-667 U/L), 11.6 U/L (range, 0.1-28.2 U/L), 43.7 g/dL (range, 21.9-88.1 g/dL), and 1.84 mg/mL (range, 0.28-4.8 mg/mL), respectively. The mean percentage of eosinophils in the pleural effusion was 19.5% (10.5-41%). Blood test was positive for parasite-specific IgG antibody in 9 patients, including 4 for Paragonimus westermani, 3 for Taenia solium, 1 for Clonorchis sinensis and 1 for Echinococcus granulosus. Eggs of Clonorchis sinensis were detected in the stool of two patients. Sparganum was found in the pleural effusion of one patient. Respiratory symptoms and abnormal appearances in pulmonary radiographic examination were disappeared in all patients who received anti-parasitic treatment. CONCLUSIONS: In patients with unexplained pleural effusion, parasite-specific IgG antibody tests should be performed when pleural fluid testing shows eosinophilic pleural effusion. It is preferable to consider the diagnosis of PPI in clinical practice when serum parasite-specific IgG antibody test is positive.


Assuntos
Imunoglobulina G/análise , Doenças Parasitárias/diagnóstico , Idoso , Dor no Peito , Tosse , Eosinófilos/patologia , Feminino , Febre , Hemoptise , Humanos , Masculino , Pessoa de Meia-Idade , Doenças Parasitárias/parasitologia , Doenças Parasitárias/patologia , Derrame Pleural/metabolismo , Derrame Pleural/parasitologia , Estudos Retrospectivos , Escarro
3.
J Dairy Sci ; 102(8): 7459-7463, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31202644

RESUMO

Transition milk is a source of immunoglobulin G (IgG) and could potentially be used to provide calves with passive immunity, when the IgG concentration is ≥50 g/L. Assessment of IgG concentrations in transition milk would be required before feeding and could be conducted using cow-side tests such as refractometers. Currently, limited information is available on the ability of refractometers to assess transition milk quality. We hypothesized that digital refractometry could be used to provide an accurate cow-side assessment of IgG concentrations in colostrum and transition milk, and IgG concentration in colostrum and one or more transition milking in an Irish herd is >50 g/L. The objectives of this study were to determine the IgG concentrations in colostrum and first, second, third, fourth, and fifth transition milk, and determine the utility of a digital refractometer in assessing quality of colostrum and transition milk produced by cows in a pasture-based dairy production system. A convenient sample of 75 dairy cows were enrolled. Colostrum and transition milk IgG concentrations were determined by radial immunodiffusion and refractometry. Sensitivity and specificity of the refractometer were determined and cut-off points that maximized sensitivity and specificity were determined using receiver operating characteristic curves. Median (range) IgG concentrations in colostrum and first, second, third, fourth, and fifth milking were 99.6, 43.5, 12.5, 5.3, 1.9, and 1.8 g/L, respectively. The sensitivity (0.8-1) of digital refractometry in identifying samples with low IgG concentrations in colostrum, first, second, and third transition milk was acceptable. In contrast, digital refractometry was not useful for assessing IgG concentrations in the fourth and fifth milking due to low IgG concentrations.


Assuntos
Colostro/imunologia , Imunoglobulina G/análise , Refratometria/veterinária , Animais , Bovinos , Feminino , Imunodifusão/veterinária , Leite , Gravidez , Curva ROC , Refratometria/instrumentação , Sensibilidade e Especificidade
4.
Schweiz Arch Tierheilkd ; 161(5): 287-297, 2019 May.
Artigo em Alemão | MEDLINE | ID: mdl-31064735

RESUMO

INTRODUCTION: Good quality colostrum intake is essential for passive immunity in neonatal calf and foals. The quality of colostrum depends on its IgG content. Since in practice the possibilities for a direct determination of these parameter are limited, an estimation of colostrum quality is often based on its biophysical properties. The simple methodology allows to perform measurements directly on site and immediately after birth. The biophysical properties included in the current review are density, refractive index, relative density, viscosity, pH, and color. The determination of the density of bovine colostrum is a common method for the evaluation of the IgG content in practice since it correlates with IgG. Density of bovine colostrum should be 1047 g/l. Different colostrometers are available for the measurement. This method is also used in the horses. Density of equine colostrum should be 1060 g/l. Refractive index or the relative density by refractometer has been proven to correlate with the IgG and is an established method in cattle and horses. Good bovine colostrum should have an index of 22% brix and equine colostrum 23% brix. Although not yet established in practice an additional methodology would be the determination viscosity of bovine colostrum. Viscosity could be measured with an outlet cup. A relationship between viscosity and IgG has demonstrated. Sufficient data for this method is missing for equine colostrum. Color and pH are not suitable for estimating the IgG content.


Assuntos
Bovinos , Colostro/química , Cavalos , Imunoglobulina G/análise , Animais , Refratometria , Viscosidade
5.
PLoS Negl Trop Dis ; 13(5): e0007353, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31059497

RESUMO

BACKGROUND: The search for diagnostic biomarkers has been profiting from a growing number of high quality sequenced genomes and freely available bioinformatic tools. These can be combined with wet lab experiments for a rational search. Improved, point-of-care diagnostic tests for visceral leishmaniasis (VL), early case detection and surveillance are required. Previous investigations demonstrated the potential of IgG1 as a biomarker for monitoring clinical status in rapid diagnostic tests (RDTs), although using a crude lysate antigen (CLA) as capturing antigen. Replacing the CLA by specific antigens would lead to more robust RDTs. METHODOLOGY: Immunoblots revealed L. donovani protein bands detected by IgG1 from VL patients. Upon confident identification of these antigens by mass spectrometry (MS), we searched for evidence of constitutive protein expression and presence of antigenic domains or high accessibility to B-cells. Selected candidates had their linear epitopes mapped with in silico algorithms. Multiple high-scoring predicted epitopes from the shortlisted proteins were screened in peptide arrays. The most promising candidate was tested in RDT prototypes using VL and nonendemic healthy control (NEHC) patient sera. RESULTS: Over 90% of the proteins identified from the immunoblots did not satisfy the selection criteria and were excluded from the downstream epitope mapping. Screening of predicted epitope peptides from the shortlisted proteins identified the most reactive, for which the sensitivity for IgG1 was 84% (95% CI 60-97%) with Sudanese VL sera on RDT prototypes. None of the sera from NEHCs were positive. CONCLUSION: We employed in silico searches to reduce drastically the output of wet lab experiments, focusing on promising candidates containing selected protein features. By predicting epitopes in silico we screened a large number of peptides using arrays, identifying the most promising one, for which IgG1 sensitivity and specificity, with limited sample size, supported this proof of concept strategy for diagnostics discovery, which can be applied to the development of more robust IgG1 RDTs for monitoring clinical status in VL.


Assuntos
Testes Diagnósticos de Rotina/métodos , Leishmaniose Visceral/diagnóstico , Anticorpos Antiprotozoários/análise , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/análise , Antígenos de Protozoários/imunologia , Simulação por Computador , Humanos , Imunoglobulina G/análise , Imunoglobulina G/imunologia , Leishmania donovani/genética , Leishmania donovani/imunologia , Leishmania donovani/isolamento & purificação , Leishmaniose Visceral/parasitologia , Peptídeos/análise , Sensibilidade e Especificidade
6.
Exp Parasitol ; 202: 1-6, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31077732

RESUMO

Neospora caninum is an apicomplexan parasite distributed worldwide. Although a positive association between the presence of birds and abortions in cattle associated to N. caninum has been reported, the role of the birds in the epidemiologic cycle of the parasite is unknown. To the best knowledge, no experimental studies have evaluated N. caninum in the eared dove, Zenaida auriculata. Therefore, we aimed to determine whether Z. auriculat can act as intermediate host for N. caninum. Eighteen birds were divided into four groups, G1, G2, G3, and G4 (control); G1, G2 and G3 received 2 × 106 tachyzoites of NC-1 strain via different routes: subcutaneous, intramuscular, and intraperitoneal, respectively. G4 composed of three birds. Serum samples were collected weekly, and one bird each from G1, G2 and G3 was euthanized on the 7th and 14th day post-inoculation (dpi). The remaining birds were euthanized after the 28th dpi. Tissues from the doves were evaluated using histopathological analysis, PCR and dog bioassay to detect the parasite. Dogs were fed with tissues from the birds and monitored for 30 days. Serum samples were collected weekly from the dogs for serological analysis, and feces samples were collected daily until the end of the experiment for coproparasitological examinations. No dove showed clinical signs of the infection; however, all of them seroconverted after the inoculation, with stronger immunological response in the G3 birds. The lung tissue of one G3 bird showed positive PCR results; it was euthanized on the 7th dpi, and an inflammatory infiltrate was observed in the lung and kidney from this dove. The dogs did not shed oocysts or seroconverted. Our results indicate that the intraperitoneal route induced infection in the doves; however, the parasite may have been eliminated by the host, and the doves may be resistant to chronic infection.


Assuntos
Doenças das Aves/parasitologia , Coccidiose/veterinária , Columbidae/parasitologia , Neospora/isolamento & purificação , Animais , Anticorpos Antiprotozoários/análise , Bioensaio/métodos , Bioensaio/veterinária , Coccidiose/parasitologia , DNA de Protozoário/química , DNA de Protozoário/isolamento & purificação , Cães , Feminino , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Imunoglobulina G/análise , Imuno-Histoquímica/veterinária , Rim/patologia , Fígado/patologia , Pulmão/patologia , Masculino , Músculo Esquelético/patologia , Neospora/genética , Neospora/imunologia , Reação em Cadeia da Polimerase/veterinária
7.
Transplant Proc ; 51(4): 1024-1026, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31101164

RESUMO

PURPOSE: The single antigen bead (SAB) test contributes to conventional cellular and solid phase crossmatch tests in renal transplantation. However, the determination of anti-HLA antibodies of the patients may not reflect the pathologic features of these antibodies. Highly sensitized patients produce antibodies against a number of HLAs; therefore, their transplantation chance decreases. In this study, we aimed to evaluate SAB and C1q test results of highly sensitized patients. METHOD: In this study, 33 end-stage renal failure patients with >80% panel reactive antibody were included. Of the patients, 58% (n = 19) were female, and 42% (n = 14) were male. The mean age was 46.2 ± 12.4. All of the serum samples were inactivated by heat before use. SAB and C1q tests were performed according to the manufacturer's instructions. RESULTS: We obtained statistically significant results between the positive bead counts and raw mean fluorescence intensity (MFI) values of 2 tests (P < .01 for class I and II). There was a statistically significant difference between the 2 tests in terms HLA-A, -C, -DR, and -DP MFI values, whereas HLA-B and -DQ MFI values were similar for the 2 tests. CONCLUSION: The difference of raw MFI values between the 2 tests may be due to the fact that the C1q test detects only IgG1 and IgG3 antibodies, whereas the SAB test can detect all IgG subtypes. We considered that anti-HLA-B and -DQ antibodies have high complement-fixing features; these antibodies should be investigated selectively due to the similarity of anti-HLA-B and -DQ antibody MFI values in the 2 tests.


Assuntos
Tipagem e Reações Cruzadas Sanguíneas/métodos , Complemento C1q/imunologia , Antígenos HLA/imunologia , Isoanticorpos/análise , Adulto , Feminino , Imunofluorescência , Humanos , Imunoglobulina G/análise , Imunoglobulina G/imunologia , Isoanticorpos/imunologia , Falência Renal Crônica , Transplante de Rim , Masculino , Pessoa de Meia-Idade
8.
Anal Chim Acta ; 1070: 112-122, 2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31103164

RESUMO

Capillary electrophoresis-systematic evolution of ligands by exponential enrichment (CE-SELEX) has proven to be an effective technique for aptamers selection. In this study, we present an online reaction based convenient single-step CE-SELEX (ssCE-SELEX) mode with human thrombin (H-Thr) as a model target. The selection progress was monitored through bulk Kd analysis, which showed more than a 1000-fold improvement over the initial library after two rounds of selection. Three selected candidate sequences presented high binding affinities against H-Thr with nanomolar (nM) Kd determined by nonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM, 56.4-177.1 nM) and CE based non-linear fitting (CE-NLF, 98.2-199.7 nM). They also exhibited high specificities towards H-Thr compared with bovine thrombin, IgG, lysozyme, and lactoferrin. Meanwhile, the Kd results by isothermal titration calorimetry (ITC) confirmed the effective CE in measuring the aptamer affinity. In addition, three candidates were applied as aptasensors in the AuNPs based colorimetric assay, which showed visible color change and good linear relationships (R2 > 0.93) with H-Thr concentration. Furthermore, molecular dynamics (MD) simulation was performed to validate the binding of the three candidates with H-Thr by binding sites and binding free energy. The ssCE-SELEX method avoids off-line incubation, saves time and sample, and may provide a universal and convenient method for aptamers selection.


Assuntos
Aptâmeros de Nucleotídeos/química , DNA de Cadeia Simples/química , Sistemas On-Line , Técnica de Seleção de Aptâmeros/métodos , Animais , Calorimetria , Bovinos , Eletroforese Capilar , Humanos , Imunoglobulina G/análise , Lactoferrina/análise , Ligantes , Muramidase/análise , Muramidase/metabolismo , Trombina/análise
9.
J Anim Sci ; 97(6): 2515-2523, 2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31004130

RESUMO

The aim of this study was to determine the effects of dietary grape seed polyphenols (GSP) supplementation during the late gestation and lactation period on reproductive performance, antioxidative status in serum, nutrient composition, and Ig content in colostrum of multiparous sows. On day 80 of gestation, a total of 64 sows with similar body condition were allocated to a completely randomized block design with 4 dietary treatments (n = 16 sows per treatment): 1) basal diet (CON, control group); 2) basal diet supplemented with 200 IU/kg vitamin E (200VE, positive control group); 3) basal diet supplemented with 200 mg/kg GSP (200GSP); and 4) basal diet supplemented with 300 mg/kg GSP (300GSP). The trial lasted 56 d until the piglets were weaned on day 21 of lactation. Reproductive performance, parameters of antioxidative status, and levels of progesterone (P4) and estradiol (E2) in serum, nutrient composition, and Ig content in colostrum of sows were determined. The number of dead fetuses was reduced, and farrowing survival was significantly improved in the litters from 300GSP-fed (P < 0.05). Preweaning survivability significantly increased in the litters from sows fed 200GSP and 200VE (P < 0.05). The activity of superoxide dismutase and glutathione peroxidase (GSH-Px) in the serum was significantly increased in sows fed 200GSP and 300GSP (P < 0.05). The activity of GSH-Px in the serum also significantly increased in sows fed 200VE (P < 0.05). Sows fed 300GSP had the greatest levels of P4 and E2 in the serum, which was significantly greater than sows fed 200VE and CON (P < 0.05). No significant differences were found among treatments for the content of solids-not-fat, fat, protein, and lactose in colostrum (P > 0.05). However, sows fed GSP had greater IgM and IgG content in colostrum compared with sows fed 200VE and CON (P < 0.05). In conclusion, dietary GSP supplementation during late gestation and lactation improved the farrowing survival and preweaning survivability, enhanced the antioxidant status and hormone levels in serum, and increased the IgM and IgG content in colostrum of sows.


Assuntos
Antioxidantes/análise , Colostro/química , Suplementos Nutricionais , Polifenóis/farmacologia , Reprodução/efeitos dos fármacos , Vitis/química , Ração Animal/análise , Animais , Líquidos Corporais/metabolismo , Dieta/veterinária , Feminino , Glutationa Peroxidase/sangue , Imunoglobulina G/análise , Lactação/efeitos dos fármacos , Paridade , Gravidez , Distribuição Aleatória , Suínos , Vitamina E/farmacologia
10.
Transbound Emerg Dis ; 66(4): 1701-1708, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30985075

RESUMO

Tick-borne encephalitis virus (TBEV) and West Nile virus (WNV) are important arthropod-borne zoonotic flaviviruses. Due to the emergence of WNV in TBEV-endemic regions co-circulation of both viruses is increasing. Flaviviruses are structurally highly similar, which leads to cross-reacting antibodies upon infection. Currently available serological assays for TBEV and WNV infections are therefore compromised by false-positive results, especially in IgG measurements. In order to discriminate both infections novel diagnostic methods are needed. We describe an ELISA to measure IgG antibodies specific for TBEV and WNV, applicable to human and horse sera. Mutant envelope proteins were generated, that lack conserved parts of the fusion loop domain, a predominant target for cross-reacting antibodies. These were incubated with equine and human sera with known TBEV, WNV or other flavivirus infections. For WNV IgG, specificities and sensitivities were 100% and 87.9%, respectively, for horse sera, and 94.4% and 92.5%, respectively, for human sera. TBEV IgG was detected with specificities and sensitivities of 95% and 96.7%, respectively, in horses, and 98.9% and 100%, respectively, in humans. Specificities increased to 100% by comparing individual samples on both antigens. The antigens could form the basis for serological TBEV- and WNV-assays with improved specificities.


Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/veterinária , Doenças dos Cavalos/diagnóstico , Febre do Nilo Ocidental/diagnóstico , Febre do Nilo Ocidental/veterinária , Vírus do Nilo Ocidental/isolamento & purificação , Animais , Anticorpos Antivirais/análise , Cavalos , Humanos , Imunoglobulina G/análise
11.
Rev Sci Instrum ; 90(3): 035004, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30927803

RESUMO

Microring resonators have shown promising potential for highly sensitive, label-free, real-time detection of biomolecules. Accurate quantitative detection of target molecules through use of photonic integrated circuits has been demonstrated for environmental monitoring and medical diagnostics. Here, we described the design, fabrication, and characterization of a highly sensitive, label-free microring optical resonator integrated with poly-(dimethylsiloxane) microfluidic channels, which consumes only 30 µl of sample solution. The resonance wavelength shifts resulting from the change in the effective refraction index can be measured in situ, and thus the binding events on the resonator surface, including antibody immobilization, blocking of the resonator surface, and the specific binding of antibody and antigen, can be recorded throughout the entire experimental process in real time. We measured the binding events for the detection of human immunoglobulin G. The system had a detection limit of 0.5 µg/ml, a value substantially (14 times) lower than that of a previously reported microring resonator. To verify the usefulness and adaptability of this technique, human epidermal growth factor receptor 2 was used for the detection. The microring optical resonator was able to monitor reactions between biological molecules in real time and thus can be used in quantitative detection and biological sensing with little sample consumption.


Assuntos
Técnicas Biossensoriais/instrumentação , Dimetilpolisiloxanos , Dispositivos Lab-On-A-Chip , Humanos , Imunoglobulina G/análise
12.
J Chromatogr B Analyt Technol Biomed Life Sci ; 1114-1115: 93-99, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30939413

RESUMO

Characterization of free thiol variants in antibody therapeutics is important for biopharmaceutical development, as the presence of free thiols may have an impact on aggregate formation, structural and thermal stability, as well as antigen-binding potency of monoclonal antibodies. Most current methods for free thiol quantification involve labeling of free thiol groups by different tagging molecules followed by UV, fluorescence or mass spectrometry (MS) detection. Here, we optimized a label-free liquid chromatography (LC)-UV/MS method for free thiol quantification at a subunit level and compared this method with two orthogonal and conventional approaches, Ellman's assay and peptide mapping with differential alkylation. This subunit unit approach was demonstrated to be able to provide domain-specific free thiol quantification and comparable results with labeling approaches, using a relatively simple and efficient workflow.


Assuntos
Anticorpos Monoclonais/análise , Anticorpos Monoclonais/química , Cromatografia Líquida/métodos , Compostos de Sulfidrila/análise , Compostos de Sulfidrila/química , Animais , Ácido Ditionitrobenzoico , Imunoglobulina G/análise , Imunoglobulina G/química , Espectrometria de Massas/métodos , Mapeamento de Peptídeos , Reprodutibilidade dos Testes
13.
J Dairy Sci ; 102(6): 5542-5549, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30904298

RESUMO

Colostrum represents the sole source to acquire humoral immunity and is an important energy source for newborn lambs and goat kids. However, colostrum composition (i.e., the contents of IgG, fat, protein, and lactose) is affected by various factors such as parity and litter size and, potentially, by breed. In the present study, we examined the colostrum composition of different goat and sheep breeds raised for milk and meat production in Switzerland and Germany. Ten goat breeds (Anglo-Nubian, Appenzell, Boer, Bunte Deutsche Edelziege, Chamois-colored, Grisons Striped, Peacock, Saanen, Toggenburg, and Valais Blackneck) and 10 sheep breeds (Brown-Headed Meat, East Friesian Milk, German Blackheaded Mutton, Gray Horned Heath, Lacaune Dairy, Merino Land, Swiss Black-Brown Mountain, Swiss Charollais, Swiss White Alpine, and Valais Blacknose) were involved in this study. First colostrum samples were obtained from ewes (n = 100) and goats (n = 116) between 10 and 390 min after parturition and analyzed for total IgG, fat, protein, and lactose contents. Colostral IgG concentrations varied between 4.8 and 75.0 mg/mL in goats, and between 6.2 and 65.4 mg/mL in ewes, and the time interval between milking and parturition did not affect colostral IgG concentrations. In goats, the highest IgG concentrations were found in Boer (meat-type; 61.0 ± 10.3 mg/mL; mean ± SD) and the lowest concentrations were observed in Bunte Deutsche Edelziege (milk-type; 26.5 ± 12.5 mg/mL). In sheep, East Friesian Milk and Lacaune Dairy showed the lowest colostral IgG concentrations (17.9 ± 7.3 and 20.2 ± 8.0 mg/mL, respectively), and the highest values were observed in the Merino Land breed (44.2 ± 15.7 mg/mL). The lowest fat and protein concentrations and concomitantly highest lactose concentrations were observed in colostrum of East Friesian Milk and Lacaune Dairy sheep. Parity number did not affect colostrum composition in sheep or goats. In contrast, colostral fat content was higher in ewes bearing twins and triplets than in those carrying singletons. Increasing litter size tended to be associated with higher protein and lower lactose concentrations in ovine (i.e., singletons vs. twins vs. triplets) and caprine colostrum (i.e., singletons vs. twins), whereas colostral IgG concentrations were not affected by litter size. In conclusion, IgG and concentrations of other colostrum constituents showed a wide range in goats and ewes and were mainly affected by the type of breed.


Assuntos
Colostro/química , Cabras/imunologia , Imunoglobulina G/análise , Leite/metabolismo , Ovinos/imunologia , Animais , Colostro/imunologia , Ácidos Graxos/análise , Feminino , Alemanha , Lactose/análise , Tamanho da Ninhada de Vivíparos , Paridade , Parto , Gravidez , Proteínas/análise , Especificidade da Espécie , Suíça
14.
Biosens Bioelectron ; 132: 196-202, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30875631

RESUMO

Photonic crystals and plasmonic nanohole arrays are the conventional substrates for label-free biodetection applications. In this article, we readdressed these systems in terms of their sensing capability and provided a broad picture for a selection mechanism of optimum parameters providing strong sensing signals. We first investigated the physical origin of the transmission resonances supported by the two systems, which is the core of the label-free sensing mechanism, relying on strong light-matter interactions. We conducted an extensive theoretical study on optical and sensing properties of the two systems, e.g., linewidth of the optical modes, refractive index sensitivity and figure-of-merit capacities. Our theoretical analyses provided a rule-of-thumb method for the selection of geometrical device parameters of the two systems. In order to experimentally investigate the sensing properties, we fabricated the two systems via a lift-off free fabrication method based on electron beam lithography, where the plasmonic nanohole arrays are realized by covering the phonic crystal surface with a thin metal. As an example, we demonstrated the sensing strength of two systems with identical dimensions by monitoring the spectral variations within their optical responses. We also performed label-free sensing experiments through detection of protein mono- and bilayers, where the geometrical parameters favor the plasmonic sensor system. Integrating a high-resolution optical read-out scheme with a multi-spectral data tracking technique, we achieved an experimentally minimum detectable protein concentration as low as 200 pg/mL for the plasmonic nanohole array and 1 ng/mL for the photonic crystal-based sensing platform.


Assuntos
Técnicas Biossensoriais/instrumentação , Nanoestruturas/química , Nanotecnologia/instrumentação , Animais , Desenho de Equipamento , Humanos , Imunoglobulina G/análise , Nanoestruturas/ultraestrutura , Fótons , Refratometria
15.
Biosens Bioelectron ; 132: 368-374, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30901726

RESUMO

A simple, convenient, and inexpensive method to fabricate optical fiber based biosensors which utilize periodic hole arrays in gold films for signal transduction is reported. The process of hole array formation mainly relies on self-assembly of hydrogel microgels in combination with chemical gold film deposition and subsequent transfer of the perforated film onto an optical fiber tip. In the fabrication process solely chemical wet lab techniques are used, avoiding cost-intensive instrumentation or clean room facilities. The presented method for preparing fiber optic plasmonic sensors provides high throughput and is perfectly suited for commercialization using batch processing. The transfer of the perforated gold film onto an optical fiber tip does not affect the sensitivity of the biosensor ((420 ±â€¯83) nm/refractive index unit (RIU)), which is comparable to sensitivities of sensor platforms based on periodic hole arrays in gold films prepared by significantly more complex methods. Furthermore, real-time and in-line immunoassay studies with a specially designed 3D printed flow cell are presented exploiting the presented optical fiber based biosensors.


Assuntos
Tecnologia de Fibra Óptica/instrumentação , Imunoensaio/instrumentação , Imunoglobulina G/análise , Ressonância de Plasmônio de Superfície/instrumentação , Animais , Desenho de Equipamento , Cabras , Ouro/química , Fibras Ópticas , Impressão Tridimensional , Coelhos , Refratometria/instrumentação
16.
Talanta ; 198: 105-110, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30876537

RESUMO

N-glycans influence the activity of antibody drugs such as antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Thus, glycan profiling is considered a critical quality attribute (CQA) and requires routine and comprehensive monitoring. In this report, we validate the new glycan profiling method called Rapi-Fluor method, which reduced the sample preparation time and increased the FLR and MS intensities compared with conventional 2-AB method. Optimized glycan release, labeling, hydrophilic interaction liquid chromatography (HILIC) enrichment, and HILIC separation resulted in low variation and short preparation time. The method evaluated for human IgG standard varied from 100 µg/mL to 4000 µg/mL in 25 µL of water. The determination of coefficient (r2 > 0.9992), recovery (88.992% ~ 111.198%), limit of detection (LOD < 193.274 µg/mL), limit of quantification (LOQ < 585.679 µg/mL), and precision (Intra-day < 2.317%RSD and Inter-day < 4.287%RSD) were evaluated with four major glycans from antibody drugs. In addition, the method was used for glycan profiling of five different commercial antibodies. The method yielded precise results for IgG glycan analysis and demonstrated effective glycan profiling of commercial antibody drugs.


Assuntos
Anticorpos Monoclonais/análise , Imunoglobulina G/análise , Polissacarídeos/análise , Adalimumab/análise , Bevacizumab/análise , Cromatografia Líquida de Alta Pressão , Humanos , Infliximab/análise , Espectrometria de Massas , Rituximab/análise , Trastuzumab/análise
17.
Anal Bioanal Chem ; 411(11): 2425-2437, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30880351

RESUMO

The baseline instability for capillary electrophoretic analysis is an intrinsic feature of the technique, which has not been thoroughly examined for its impact on therapeutic protein purity analysis with the capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) applications. For the particular CE-SDS application, this phenomenon was manifested through peak migration time shifts and sliding of the superimposed baseline profile. These dual phenomena are closely associated so that experimental assessment alone may not shed enough light to the underlying drivers. In the current study, both experimental and simulation approaches were employed to assess the systematic drifts. Computer simulation was used to decipher the two underlying factors and test their contributions toward purity and impurity peak determination inaccuracies. The data generated in this study demonstrated that the electrophoretic baseline disturbance had more pronounced impact on the purity data than the migration time shift. In addition, the potential contributing factors to the baseline disturbances were assessed experimentally which indicated that the source is related to thermal disruption during a sample run and the unique baseline patterns came from the background electrolytes. To improve data reproducibility for drug purity testing in the industrial setting and quality control (QC) environment, it is recommended to run shorter injection sequences including fewer samples and closely monitor the baseline drift for accurate integration. Those methods would help reduce the impact of systematic drift and disturbances. Graphical abstract.


Assuntos
Anticorpos Monoclonais/análise , Eletroforese Capilar/métodos , Imunoglobulina G/análise , Anticorpos Monoclonais/isolamento & purificação , Simulação por Computador , Contaminação de Medicamentos , Imunoglobulina G/isolamento & purificação , Modelos Químicos , Dodecilsulfato de Sódio/química
20.
Eur J Clin Microbiol Infect Dis ; 38(5): 883-890, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30701339

RESUMO

Sampling the blood compartment by an invasive procedure such as phlebotomy is the most common approach used for diagnostic purposes. However, phlebotomy has several drawbacks including pain, vasovagal reactions, and anxiety. Therefore, alternative approaches should be tested to minimize patient's discomfort. Saliva is a reasonable compartment; when obtained, it generates little or no anxiety. We setup a multiplexed serology assay for detection of Toxoplasma gondii IgG and IgM, rubella IgG, and CMV IgG, in serum, whole blood, and saliva using novel plasmonic gold (pGOLD) chips. pGOLD test results in serum, whole blood, and saliva were compared with commercial kits test results in serum. One hundred twenty serum/saliva sets (Lyon) and 28 serum/whole blood/saliva sets (Nice) from France were tested. In serum and whole blood, sensitivity and specificity of multiplex T. gondii, CMV, and rubella IgG were 100% in pGOLD when compared to commercial test results in serum. In saliva, sensitivity and specificity for T. gondii and rubella IgG were 100%, and for CMV IgG, sensitivity and specificity were 92.9% and 100%, respectively, when compared to commercial test results in serum. We were also able to detect T. gondii IgM in saliva with sensitivity and specificity of 100% and 95.4%, respectively, when compared to serum test results. Serological testing by multiplex pGOLD assay for T. gondii, rubella, and CMV in saliva is reliable and likely to be more acceptable for systematic screening of pregnant women, newborn, and immunocompromised patients.


Assuntos
Infecções por Citomegalovirus/diagnóstico , Ouro/química , Análise Serial de Proteínas/normas , Rubéola (Sarampo Alemão)/diagnóstico , Saliva/imunologia , Testes Sorológicos/normas , Toxoplasmose/diagnóstico , Adolescente , Adulto , Anticorpos Antiprotozoários/análise , Anticorpos Antivirais/análise , Antígenos de Protozoários/química , Antígenos Virais/química , Criança , Pré-Escolar , Citomegalovirus/imunologia , Citomegalovirus/isolamento & purificação , Humanos , Imunoglobulina G/análise , Imunoglobulina M/análise , Lactente , Recém-Nascido , Pessoa de Meia-Idade , Vírus da Rubéola/imunologia , Vírus da Rubéola/isolamento & purificação , Sensibilidade e Especificidade , Toxoplasma/imunologia , Toxoplasma/isolamento & purificação , Adulto Jovem
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