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1.
Food Chem ; 359: 129878, 2021 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-33934031

RESUMO

The allergenic potency of the cricket Acheta domesticus, a promising edible insect, has never been assessed. This work aims to study the immunoreactivity of Acheta domesticus, and its cross-reactivity with the shrimp Litopenaeus vannamei, assessing the effect of cooking and gastrointestinal digestion on their allergenic properties. Different cricket proteins were detected by immunoblotting with shrimp-allergic patients' sera. Tropomyosin was identified as the most relevant IgE-binding protein, and its cross-reactivity with shrimp tropomyosin was demonstrated by ELISA. While shrimp tropomyosin showed scarce stability to gastric digestion, cricket tropomyosin withstood the whole digestion process. The sarcoplasmic calcium-binding protein, specifically detected in shrimp, showed exceptional stability to gastrointestinal digestion. IgE-binding proteins in a model of enriched baked products were partially protected from proteolysis. In conclusion, the ingestion of A. domesticus proteins poses serious concerns to the Crustacean-allergic population. The high stability of tropomyosin may represent a risk of primary sensitization and clinical cross-reactivity.


Assuntos
Alérgenos/análise , Hipersensibilidade Alimentar , Gryllidae/imunologia , Imunoglobulina E/análise , Penaeidae/química , Frutos do Mar/análise , Animais , Proteínas de Ligação ao Cálcio/imunologia , Reações Cruzadas , Digestão , Ensaio de Imunoadsorção Enzimática , Manipulação de Alimentos , Gryllidae/química , Humanos , Immunoblotting , Tropomiosina/imunologia
2.
Cutan Ocul Toxicol ; 37(4): 309-318, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29508626

RESUMO

BACKGROUND: Drug hypersensitivity reactions of immediate type pose a challenging problem, especially, if standard diagnostic procedures do not lead to conclusive results. The aim of this investigation is to identify, whether basophil activation test (BAT) is able to provide additional benefit in the diagnostic evaluation of immediate type drug hypersensitivity reactions to antibiotics in comparison with the routine allergological diagnostic methods. MATERIALS AND METHODS: We investigated patients, who presented to the Department of Dermatology and Allergology of the University Hospital of RWTH Aachen in Germany for diagnostic workup of type I allergic reactions to antibiotics during the period from 2009 to 2012. The analysis was performed retrospectively based on patient records. The inclusion criteria were performed standard allergological in vivo diagnostic and a BAT as a part of diagnostic workup. RESULTS: Eighty-two diagnostic investigations were performed in 52 patients. BAT was positive in 9 of 12 cases with a positive clinical history but negative skin test results. Furthermore, all patients who reported severe drug hypersensitivity reactions (anaphylactic reaction grade 2 and above) showed positive BAT (5/5), while only three of these five cases demonstrated a positive skin testing that led to the conclusion of possible immediate type drug hypersensitivity. CONCLUSIONS: Although skin tests remain the most important part of the primary diagnostic investigation, BAT is an additional valuable and sensitive in vitro test in the diagnostic procedure of immediate type allergic reactions to antibiotics. However, further standardized investigations are needed in order to calculate exact sensitivity and specificity of this diagnostic tool in both, adult and pediatric populations.


Assuntos
Antibacterianos/efeitos adversos , Basófilos , Hipersensibilidade a Drogas/diagnóstico , beta-Lactamas/efeitos adversos , Adolescente , Adulto , Idoso , Anafilaxia/diagnóstico , Anafilaxia/etiologia , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Hipersensibilidade Imediata/diagnóstico , Imunoglobulina E/análise , Ativação de Macrófagos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Estudos Retrospectivos , Medição de Risco , Testes Cutâneos , Adulto Jovem
3.
Allergy ; 73(8): 1707-1714, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29601091

RESUMO

BACKGROUND: Early peanut introduction (EPI) in the first year of life is associated with reduced risk of developing peanut allergy in children with either severe eczema and/or egg allergy. However, EPI recommendations differ among countries with formal guidelines. METHODS: Using simulation and Markov modeling over a 20-year horizon to attempt to explore optimal EPI strategies applied to the US population, we compared high-risk infant-specific IgE peanut screening (US/Canadian) with the Australiasian Society for Clinical Immunology and Allergy (Australia/New Zealand) (ASCIA) and the United Kingdom Department of Health (UKDOH)-published EPI approaches. RESULTS: Screening peanut skin testing of all children with early-onset eczema and/or egg allergy before in-office peanut introduction was dominated by a no screening approach, in terms of number of cases of peanut allergy prevented, quality-adjusted life years (QALY), and healthcare costs, although screening resulted in a slightly lower rate of allergic reactions to peanut per patient in high-risk children. Considering costs of peanut allergy in high-risk children, the per-patient cost of early introduction without screening over the model horizon was $6556.69 (95%CI, $6512.76-$6600.62), compared with a cost of $7576.32 (95%CI, $7531.38-$7621.26) for skin test screening prior to introduction. From a US societal perspective, screening prior to introduction cost $654 115 322 and resulted in 3208 additional peanut allergy diagnoses. Both screening and nonscreening approaches dominated deliberately delayed peanut introduction. CONCLUSIONS: A no-screening approach for EPI has superior health and economic benefits in terms of number of peanut allergy cases prevented, QALY, and total healthcare costs compared to screening and in-office peanut introduction.


Assuntos
Arachis/imunologia , Imunomodulação , Triagem Multifásica/economia , Hipersensibilidade a Amendoim/economia , Hipersensibilidade a Amendoim/prevenção & controle , Testes Cutâneos/economia , Fatores Etários , Austrália , Canadá , Criança , Pré-Escolar , Intervenção Médica Precoce , Eczema , Hipersensibilidade a Ovo , Diretrizes para o Planejamento em Saúde , Humanos , Imunoglobulina E/análise , Lactente , Cadeias de Markov , Nova Zelândia , Hipersensibilidade a Amendoim/imunologia , Risco , Reino Unido , Estados Unidos
4.
Soc Sci Med ; 165: 19-27, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27485729

RESUMO

Understanding how human environments affect our health by "getting under the skin" and penetrating the cells, organs and physiological systems of our bodies is a key tenet in public health research. Here, we examine the idea that early life socioeconomic position (SEP) can be biologically embodied, potentially leading to the production of health inequalities across population groups. Allostatic load (AL), a composite measure of overall physiological wear-and-tear, could allow for a better understanding of the potential biological pathways playing a role in the construction of the social gradient in adult health. We investigate the factors mediating the link between two components of parental SEP, maternal education (ME) and parental occupation (PO), and AL at 44 years. Data was used from 7573 members of the 1958 British birth cohort follow-up to age 44. AL was constructed using 14 biomarkers representing four physiological systems. We assessed the contribution of financial/materialist, psychological/psychosocial, educational, and health behaviors/BMI pathways over the life course, in mediating the associations between ME, PO and AL. ME and PO were mediated by three pathways: educational, material/financial, and health behaviors, for both men and women. A better understanding of embodiment processes leading to disease development may contribute to developing adapted public policies aiming to reduce health inequalities.


Assuntos
Alostase/fisiologia , Biomarcadores/análise , Classe Social , Adulto , Pressão Sanguínea , Índice de Massa Corporal , Proteína C-Reativa/análise , Estudos de Coortes , Escolaridade , Inglaterra , Etnicidade/estatística & dados numéricos , Feminino , Fibrinogênio/análise , Hemoglobinas Glicadas/análise , Frequência Cardíaca , Humanos , Hidrocortisona/análise , Imunoglobulina E/análise , Imunoglobulina E/sangue , Lipoproteínas HDL/análise , Lipoproteínas HDL/sangue , Masculino , Mães/estatística & dados numéricos , Pico do Fluxo Expiratório , Autorrelato , Inquéritos e Questionários , Triglicerídeos/análise , Triglicerídeos/sangue
5.
Biomed Microdevices ; 18(2): 33, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26973054

RESUMO

Herein, we report a simple, sensitive, rapid and low-cost ultraminiaturized assay technique for quantitative detection of 1 µl of clinical or biochemical sample on a novel ultraminiaturized assay plate (UAP). UAP is prepared by making tiny cavities on a polypropylene sheet. As UAP cannot immobilize a biomolecule through absorption, we have activated the tiny cavities of UAP by 1-fluoro-2-nitro-4-azidobenzene in a photochemical reaction. Activated UAP (AUAP) can covalently immobilize any biomolecule having an active nucleophilic group such as amino group. Efficacy of AUAP is demonstrated by detecting human IgE, antibody of hepatitis C virus core antigen and oligonucleotides. Quantification is performed by capturing the image of the colored assay solution and digitally quantifying the image by color saturation without using costly NanoDrop spectrophotometer. Image - based detection of human IgE and an oligonucleotide shows an excellent correlation with absorbance - based assay (recorded in a NanoDrop spectrophotometer); it is validated by Pearson's product-moment correlation with correlation coefficient of r = 0.9545088 and r = 0.9947444 respectively. AUAP is further checked by detecting hepatitis C virus Ab where strong correlation of color saturation with absorbance with respect to concentration is observed. Ultraminiaturized assay successfully detects target oligonucleotides by perfectly hybridizing with their respective complementary oligonucleotide probes but not with a random oligonucleotide. Ultraminiaturized assay technique has substantially reduced the requirement of reagents by 100 times and assay timing by 50 times making it a potential alternative to conventional method.


Assuntos
Custos e Análise de Custo , Ensaio de Imunoadsorção Enzimática/economia , Ensaio de Imunoadsorção Enzimática/métodos , Miniaturização/métodos , Animais , Antígenos Virais/imunologia , Azidas/química , Cor , Hepacivirus/imunologia , Humanos , Imunoglobulina E/análise , Imunoglobulina E/imunologia , Nitrobenzenos/química , Oligonucleotídeos/análise , Processos Fotoquímicos , Fatores de Tempo
6.
Food Chem Toxicol ; 84: 181-7, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26321724

RESUMO

Rice is cultivated as a staple grain crop in many countries, especially in Asia. In the present study, recombinant rice chitinase was expressed, purified and characterized by in silico and immunobiochemical methods. Rice chitinase was affinity purified and it resolved at 24 kDa on SDS-PAGE. Purified protein was analyzed for pepsin resistance, heat stability, and IgE binding using atopic patients' sera. Chitinase was resistant to pepsin digestion and heat treatment at 90 °C for 1 h. It showed significant IgE binding with 7 of 110 patients' sera positive to different food allergens. Homology modeled 3D structure of rice chitinase was used for B cell epitope prediction. In silico predicted B cell peptides were assessed for IgE binding by ELISA using food allergic patients' sera, epitope RC2 showed IgE binding comparable to chitinase. In conclusion, chitinase was identified as a potential allergen and may share cross reactive epitopes with food allergens.


Assuntos
Alérgenos/química , Quitinases/química , Proteínas Alimentares/química , Imunoglobulina E/química , Modelos Moleculares , Oryza/enzimologia , Proteínas de Plantas/química , Alérgenos/efeitos adversos , Alérgenos/genética , Alérgenos/metabolismo , Sítios de Ligação , Sítios de Ligação de Anticorpos , Quitinases/efeitos adversos , Quitinases/genética , Quitinases/metabolismo , Biologia Computacional , Reações Cruzadas , Proteínas Alimentares/efeitos adversos , Proteínas Alimentares/metabolismo , Digestão , Estabilidade Enzimática , Mapeamento de Epitopos , Sistemas Inteligentes , Hipersensibilidade Alimentar/sangue , Hipersensibilidade Alimentar/imunologia , Temperatura Alta/efeitos adversos , Humanos , Imunoglobulina E/análise , Imunoglobulina E/metabolismo , Índia , Oryza/efeitos adversos , Fragmentos de Peptídeos/efeitos adversos , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Proteínas de Plantas/efeitos adversos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Recombinantes/efeitos adversos , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Homologia Estrutural de Proteína
7.
Ann Allergy Asthma Immunol ; 115(1): 10-6.e1, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26123420

RESUMO

BACKGROUND: Despite the significant burden of childhood asthma, little is known about prevention-oriented management before and after hospitalizations for asthma exacerbation. OBJECTIVE: To investigate the proportion and characteristics of children admitted to the intensive care unit (ICU) for asthma exacerbation and the frequency of guideline-recommended outpatient management before and after the hospitalization. METHODS: A 14-center medical record review study of children aged 2 to 17 years hospitalized for asthma exacerbation during 2012-2013. Primary outcome was admission to the ICU; secondary outcomes were 2 preventive factors: inhaled corticosteroid (ICS) use and evaluation by asthma specialists in the pre- and posthospitalization periods. RESULTS: Among 385 children hospitalized for asthma, 130 (34%) were admitted to the ICU. Risk factors for ICU admission were female sex, having public insurance, a marker of chronic asthma severity (ICS use), and no prior evaluation by an asthma specialist. Among children with ICU admission, guideline-recommended outpatient management was suboptimal (eg, 65% were taking ICSs at the time of index hospitalization, and 19% had evidence of a prior evaluation by specialist). At hospital discharge, among children with ICU admission who had not previously used controller medications, 85% were prescribed ICSs. Furthermore, 62% of all children with ICU admission were referred to an asthma specialist during the 3-month posthospitalization period. CONCLUSION: In this multicenter study of US children hospitalized with asthma exacerbation, one-third of children were admitted to the ICU. In this high-risk group, we observed suboptimal pre- and posthospitalization asthma care. These findings underscore the importance of continued efforts to improve prevention-oriented asthma care at all clinical encounters.


Assuntos
Antiasmáticos/uso terapêutico , Asma/tratamento farmacológico , Fidelidade a Diretrizes , Guias de Prática Clínica como Assunto , Adolescente , Alergia e Imunologia , Criança , Pré-Escolar , Gerenciamento Clínico , Feminino , Humanos , Imunoglobulina E/análise , Pacientes Internados/estatística & dados numéricos , Seguro Saúde/estatística & dados numéricos , Unidades de Terapia Intensiva , Tempo de Internação/estatística & dados numéricos , Masculino , Encaminhamento e Consulta , Estudos Retrospectivos , Índice de Gravidade de Doença , Distribuição por Sexo , Estados Unidos
8.
Otolaryngol Head Neck Surg ; 152(1 Suppl): S1-43, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25644617

RESUMO

OBJECTIVE: Allergic rhinitis (AR) is one of the most common diseases affecting adults. It is the most common chronic disease in children in the United States today and the fifth most common chronic disease in the United States overall. AR is estimated to affect nearly 1 in every 6 Americans and generates $2 to $5 billion in direct health expenditures annually. It can impair quality of life and, through loss of work and school attendance, is responsible for as much as $2 to $4 billion in lost productivity annually. Not surprisingly, myriad diagnostic tests and treatments are used in managing this disorder, yet there is considerable variation in their use. This clinical practice guideline was undertaken to optimize the care of patients with AR by addressing quality improvement opportunities through an evaluation of the available evidence and an assessment of the harm-benefit balance of various diagnostic and management options. PURPOSE: The primary purpose of this guideline is to address quality improvement opportunities for all clinicians, in any setting, who are likely to manage patients with AR as well as to optimize patient care, promote effective diagnosis and therapy, and reduce harmful or unnecessary variations in care. The guideline is intended to be applicable for both pediatric and adult patients with AR. Children under the age of 2 years were excluded from the clinical practice guideline because rhinitis in this population may be different than in older patients and is not informed by the same evidence base. The guideline is intended to focus on a limited number of quality improvement opportunities deemed most important by the working group and is not intended to be a comprehensive reference for diagnosing and managing AR. The recommendations outlined in the guideline are not intended to represent the standard of care for patient management, nor are the recommendations intended to limit treatment or care provided to individual patients. ACTION STATEMENTS: The development group made a strong recommendation that clinicians recommend intranasal steroids for patients with a clinical diagnosis of AR whose symptoms affect their quality of life. The development group also made a strong recommendation that clinicians recommend oral second-generation/less sedating antihistamines for patients with AR and primary complaints of sneezing and itching. The panel made the following recommendations: (1) Clinicians should make the clinical diagnosis of AR when patients present with a history and physical examination consistent with an allergic cause and 1 or more of the following symptoms: nasal congestion, runny nose, itchy nose, or sneezing. Findings of AR consistent with an allergic cause include, but are not limited to, clear rhinorrhea, nasal congestion, pale discoloration of the nasal mucosa, and red and watery eyes. (2) Clinicians should perform and interpret, or refer to a clinician who can perform and interpret, specific IgE (skin or blood) allergy testing for patients with a clinical diagnosis of AR who do not respond to empiric treatment, or when the diagnosis is uncertain, or when knowledge of the specific causative allergen is needed to target therapy. (3) Clinicians should assess patients with a clinical diagnosis of AR for, and document in the medical record, the presence of associated conditions such as asthma, atopic dermatitis, sleep-disordered breathing, conjunctivitis, rhinosinusitis, and otitis media. (4) Clinicians should offer, or refer to a clinician who can offer, immunotherapy (sublingual or subcutaneous) for patients with AR who have inadequate response to symptoms with pharmacologic therapy with or without environmental controls. The panel recommended against (1) clinicians routinely performing sinonasal imaging in patients presenting with symptoms consistent with a diagnosis of AR and (2) clinicians offering oral leukotriene receptor antagonists as primary therapy for patients with AR. The panel group made the following options: (1) Clinicians may advise avoidance of known allergens or may advise environmental controls (ie, removal of pets; the use of air filtration systems, bed covers, and acaricides [chemical agents formulated to kill dust mites]) in patients with AR who have identified allergens that correlate with clinical symptoms. (2) Clinicians may offer intranasal antihistamines for patients with seasonal, perennial, or episodic AR. (3) Clinicians may offer combination pharmacologic therapy in patients with AR who have inadequate response to pharmacologic monotherapy. (4) Clinicians may offer, or refer to a surgeon who can offer, inferior turbinate reduction in patients with AR with nasal airway obstruction and enlarged inferior turbinates who have failed medical management. (5) Clinicians may offer acupuncture, or refer to a clinician who can offer acupuncture, for patients with AR who are interested in nonpharmacologic therapy. The development group provided no recommendation regarding the use of herbal therapy for patients with AR.


Assuntos
Antialérgicos/uso terapêutico , Terapias Complementares/métodos , Rinite Alérgica/diagnóstico , Rinite Alérgica/terapia , Terapia por Acupuntura/métodos , Administração Intranasal , Adolescente , Adulto , Criança , Pré-Escolar , Doença Crônica , Comorbidade , Efeitos Psicossociais da Doença , Análise Custo-Benefício , Diagnóstico Diferencial , Quimioterapia Combinada , Medicina Baseada em Evidências , Feminino , Glucocorticoides/administração & dosagem , Antagonistas dos Receptores Histamínicos/uso terapêutico , Humanos , Imunoglobulina E/análise , Imunoterapia/métodos , Comunicação Interdisciplinar , Antagonistas de Leucotrienos/uso terapêutico , Masculino , Procedimentos Cirúrgicos Nasais/métodos , Fitoterapia/métodos , Prevalência , Qualidade de Vida , Encaminhamento e Consulta , Rinite Alérgica/tratamento farmacológico , Rinite Alérgica/economia , Rinite Alérgica/epidemiologia , Rinite Alérgica/imunologia , Conchas Nasais/cirurgia , Estados Unidos/epidemiologia
9.
Vestn Ross Akad Med Nauk ; (6): 748-55, 2015.
Artigo em Russo | MEDLINE | ID: mdl-27093804

RESUMO

UNLABELLED: High morbidity rate of atopic diseases among children, including high importance of grass pollen as a sensitizing agent, determine the relevance ofstudies on diagnostic examination systems for appointment of adequate therapy. The research of the most relevant allergens for patients to excludeof duplicating and uninformative tests became urgent after development of a new type of diagnostic tests that does not require expensive equipment. The objective of this research was to evaluate the results of in vitro- and in vivo-diagnostic examinations of children with various forms of atopic disease caused by pollen of meadow grasses, and to choose the most significant prognostic parameters for the diagnosis. METHODS: 277 children aged 4-16 years with various forms of atopic disease were included in the study. There were performed skin prick tests and determination of IgE-antibodies levels to allergen extracts of cocksfoot (g3), meadow fescue (g4), timothy grass (g6). RESULTS: In the studied group of patients 32-50% of children have antibodies to grass allergens. There was a close correlation of antibody response on the investigated allergens, quantitative coincidence of IgE-antibodies to g3 andg4 allergens levels. IgE (g6) concentration was close to the IgE(g3) and IgE(g4) levels (85.0 ± 21.6%). Analysis of the skin tests results showed that 44% of patients have a positive response to grass allergens, and in vivo-tests results coincide with serologicaltests results, mostly in a qualitative sense. The most significant relationship was noted between in vivo and in vitro-tests in the results of testing the response to meadow fescue pollen. CONCLUSION: Based on these data IgE concentration index to meadow fescue allergens can be used as a prognostic marker to determine the sensitization of patients with different nosology forms of allergy and can help to improve allergic diagnostics.


Assuntos
Alérgenos , Dermatite Atópica , Imunoglobulina E , Pólen , Rinite Alérgica Sazonal , Adolescente , Alérgenos/classificação , Alérgenos/imunologia , Criança , Pré-Escolar , Redução de Custos/métodos , Dermatite Atópica/sangue , Dermatite Atópica/diagnóstico , Dermatite Atópica/economia , Dermatite Atópica/etiologia , Feminino , Humanos , Imunoglobulina E/análise , Imunoglobulina E/sangue , Técnicas Imunológicas/economia , Técnicas Imunológicas/métodos , Masculino , Avaliação das Necessidades , Pólen/classificação , Pólen/imunologia , Rinite Alérgica Sazonal/sangue , Rinite Alérgica Sazonal/diagnóstico , Rinite Alérgica Sazonal/economia , Rinite Alérgica Sazonal/etiologia , Federação Russa
12.
Bioanalysis ; 6(14): 2005-18, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25158969

RESUMO

Allergies occur when a person's immune system mounts an abnormal response with or without IgE to a normally harmless substance called an allergen. The standard skin-prick test introduces suspected allergens into the skin with lancets in order to trigger allergic reactions. This test is annoying and sometimes life threatening. New tools such as lab-on-a-chip and lab-on-a-disc, which rely on microfabrication, are designed for allergy testing. These systems provide benefits such as short analysis times, enhanced sensitivity, simplified procedures, minimal consumption of sample and reagents and low cost. This article gives a summary of these systems. In particular, a cell-based assay detecting both the IgE- and non-IgE-type triggers through the study of degranulation in a centrifugal microfluidic system is highlighted.


Assuntos
Alérgenos/análise , Hipersensibilidade/diagnóstico , Imunoglobulina E/análise , Dispositivos Lab-On-A-Chip , Técnicas Analíticas Microfluídicas/instrumentação , Alérgenos/imunologia , Animais , Teste de Degranulação de Basófilos/economia , Teste de Degranulação de Basófilos/instrumentação , Teste de Degranulação de Basófilos/métodos , Basófilos/imunologia , Basófilos/fisiologia , Degranulação Celular , Desenho de Equipamento , Humanos , Hipersensibilidade/imunologia , Imunoglobulina E/imunologia , Dispositivos Lab-On-A-Chip/economia , Mastócitos/imunologia , Mastócitos/fisiologia , Técnicas Analíticas Microfluídicas/economia , Técnicas Analíticas Microfluídicas/métodos
13.
Ann Biol Clin (Paris) ; 71(3): 325-32, 2013.
Artigo em Francês | MEDLINE | ID: mdl-23747670

RESUMO

French medical laboratories must be accredited before November 2016 according to NF/EN/ISO 15189 standard. However, technical accreditation guidelines cannot be applied literally for the determination of specific IgE for several reasons: more than 600 allergen tests, lack of international gold standard, limited external quality controls. Furthermore, the technique for determination of specific IgE is CE DM-IVD marked, common to all specificities, automatised, standardized according to a single calibration curve. Thus, we propose an efficient but reasonable solution conform to the idea of the accreditation by validating the process. We recommend: a flexible extend type A; choice of only one representative allergen (Dermatophagoides pteronyssinus) for repeatability and precision (20 tests, 2 levels 0.5-1 and 8-12 kUA/L) performed on patients sera, reproducibility (30 consecutive determinations using an Internal Quality Control/IQC), accuracy (IQC and rare External Quality Controls) compared with peers. Sensitivity, specificity, dynamic range, detection threshold are determinated by the provider. Linearity may be checked if the laboratory practices sample dilution for values higher than the upper limit guaranteed by the provider. In the absence of international gold standard, the uncertainty is not measurable. In case of change of instrument, the results obtained by the systems must be compared through 35 tests of different specificities distributed across the range of calibration and including 5 values close to the detection limit. This methodology allows a scientifically effective verification, technically and financially reasonable, to ensure the excellence of the performance of the laboratory with regard to peers and users (allergologists and patients).


Assuntos
Acreditação/normas , Alérgenos/imunologia , Técnicas de Laboratório Clínico/normas , Imunoglobulina E/análise , Laboratórios/legislação & jurisprudência , Técnicas de Laboratório Clínico/economia , Técnicas de Laboratório Clínico/métodos , Análise Custo-Benefício , Humanos , Hipersensibilidade/sangue , Hipersensibilidade/diagnóstico , Hipersensibilidade/imunologia , Imunoglobulina E/imunologia , Laboratórios/economia , Laboratórios/normas , Guias de Prática Clínica como Assunto , Controle de Qualidade , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Incerteza , Estudos de Validação como Assunto
14.
Biosens Bioelectron ; 48: 287-92, 2013 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-23708751

RESUMO

In this communication, we report ELISA technique on an activated polypropylene microtest plate (APPµTP) as an illustrative example of a low cost diagnostic assay. Activated test zone in APPµTP binds a capture biomolecule through covalent linkage thereby, eliminating non-specific binding often prevalent in absorption based techniques. Efficacy of APPµTP is demonstrated by detecting human immunoglobulin G (IgG), human immunoglobulin E (IgE) and Aspergillus fumigatus antibody in patient's sera. Detection is done by taking the image of the assay solution by a desktop scanner and analyzing the color of the image. Human IgE quantification by color saturation in the image-based assay shows excellent correlation with absorbance-based assay (Pearson correlation coefficient, r=0.992). Significance of the relationship is seen from its p value which is 4.087e-11. Performance of APPµTP is also checked with respect to microtiter plate and paper-based ELISA. APPµTP can quantify an analyte as precisely as in microtiter plate with insignificant non-specific binding, a necessary prerequisite for ELISA assay. In contrast, paper-ELISA shows high non-specific binding in control sera (false positive). Finally, we have carried out ELISA steps on APPµTP by ultrasound waves on a sonicator bath and the results show that even in 8 min, it can convincingly differentiate a test sample from a control sample. In short, spectrophotometer-free image-based miniaturized ELISA on APPµTP is precise, reliable, rapid, and sensitive and could be a good substitute for conventional immunoassay procedures widely used in clinical and research laboratories.


Assuntos
Anticorpos Antifúngicos/sangue , Aspergilose/sangue , Aspergillus fumigatus/imunologia , Ensaio de Imunoadsorção Enzimática/instrumentação , Imunoglobulina E/análise , Imunoglobulina G/análise , Aspergilose/imunologia , Ensaio de Imunoadsorção Enzimática/economia , Humanos , Polipropilenos/química , Sensibilidade e Especificidade , Fatores de Tempo
15.
Intern Med J ; 43(1): 46-52, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21790924

RESUMO

BACKGROUND: There is overwhelming evidence that asthma guidelines aimed at reducing airway inflammation are superior to those based on clinical symptoms alone. This involves targeting eosinophilic inflammation with inhaled corticosteroids. AIM: Because induced sputum is not readily available, our study set out to investigate whether the collective or singular use of routine asthma investigations can predict sputum eosinophilia. METHODS: Eighty patients underwent skin prick testing, blood tests (IgE, full blood count), spirometry, exhaled fraction nitric oxide (FeNO), PD15 to hypertonic saline, and induced sputum testing at first assessment. A predictive model for sputum eosinophilia (defined as ≥3% eosinophils) was sought using routinely available tests. RESULTS: Fifty-four subjects underwent both induced sputum and FeNO testing. Seventeen (30%) revealed eosinophilic inflammation, nine (16%) neutrophilic, four (7%) mixed granulocytic and 26 (46%) paucigranulocytic. Positive predictors for sputum eosinophilia included low forced expiratory volume in 1 s (FEV(1))% predicted, raised serum eosinophil, positive smoking history, Polynesian ethnicity and negative asthma family history. There was a non-statistically significant trend for FeNO predicting sputum eosinophilia. The best combination of predictors was low FEV(1)% predicted, raised serum eosinophil, positive smoking history and negative family history of asthma. CONCLUSION: This study demonstrates that the serum eosinophil count and FEV(1) combined with aspects of a clinical history may provide a simple and practical alternative to assessment of airway (sputum) eosinophilia in the clinical setting. A full blood count can be performed at a substantially lesser cost and with greater accessibility than induced sputum. We feel the time has come for the clinical utility of the serum eosinophil count to be revisited.


Assuntos
Asma/patologia , Eosinofilia Pulmonar/diagnóstico , Escarro/citologia , Corticosteroides/uso terapêutico , Adulto , Antiasmáticos/uso terapêutico , Asma/complicações , Asma/tratamento farmacológico , Asma/imunologia , Testes Respiratórios , Estudos Transversais , Eosinófilos , Feminino , Volume Expiratório Forçado , Humanos , Imunoglobulina E/análise , Masculino , Pessoa de Meia-Idade , Óxido Nítrico/análise , Guias de Prática Clínica como Assunto , Valor Preditivo dos Testes , Estudos Prospectivos , Eosinofilia Pulmonar/etiologia , Eosinofilia Pulmonar/patologia , Solução Salina Hipertônica , Salivação/efeitos dos fármacos , Testes Cutâneos
16.
J Nutr Sci Vitaminol (Tokyo) ; 59(5): 462-9, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24418881

RESUMO

Miso paste (miso), a fermented soybean food, is popular in Japan and other Asian countries. However, the soybean is known to induce an allergenic response in some individuals. In the present study, we evaluated the allergenicity of various kinds of miso available in Japan. Total proteins were extracted from Amakuti-kome miso, Karakuti-kome miso, Mugi-miso and Mame-miso, and the protein profiles were analyzed. The major protein bands detected in the intact soybean extract were not present in any of the miso samples, which instead showed various low molecular weight protein bands of approximately 10-25 kDa. The existence levels of six major soybean allergens were determined by Western blotting using specific antibodies. We found that the allergen levels varied among miso and allergen types; however, allergen levels were consistently lower in miso than in the soybean extract. We obtained similar results for IgE-ELISA experiments using serum IgE from soybean allergy patients. Taken together, these results indicate that compared to soybean extract, various types of miso contain small quantities of intact soybean allergens. Additionally, several lines of evidence indicated that the allergen levels were exceptionally low in the dark-colored Karakuti-kome miso and Mame-miso, which are produced with relatively long fermentation periods, suggesting that the duration of fermentation might be a key factor in the hypoallergenicity of miso.


Assuntos
Alérgenos/análise , Antígenos de Plantas/análise , Dieta , Hipersensibilidade Alimentar/imunologia , Fragmentos de Peptídeos/efeitos adversos , Alimentos de Soja/efeitos adversos , Proteínas de Soja/efeitos adversos , Alérgenos/efeitos adversos , Alérgenos/química , Alérgenos/metabolismo , Antígenos de Plantas/efeitos adversos , Antígenos de Plantas/química , Antígenos de Plantas/metabolismo , Aspergillus/metabolismo , Bacillus/metabolismo , Western Blotting , Dieta/etnologia , Ensaio de Imunoadsorção Enzimática , Fermentação , Hipersensibilidade Alimentar/sangue , Hipersensibilidade Alimentar/dietoterapia , Hipersensibilidade Alimentar/etiologia , Humanos , Imunoglobulina E/análise , Imunoglobulina E/metabolismo , Japão , Peso Molecular , Fragmentos de Peptídeos/análise , Fragmentos de Peptídeos/isolamento & purificação , Fragmentos de Peptídeos/metabolismo , Pigmentação , Extratos Vegetais/efeitos adversos , Extratos Vegetais/química , Proteólise , Alimentos de Soja/análise , Alimentos de Soja/economia , Alimentos de Soja/microbiologia , Proteínas de Soja/análise , Proteínas de Soja/química , Proteínas de Soja/metabolismo , Fatores de Tempo
17.
Mol Nutr Food Res ; 56(6): 986-95, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22707273

RESUMO

SCOPE: Ara h 6 has recently been recognized as an important peanut allergen. Recombinant allergens have been used for analysis of IgE binding, but have not been used to analyze the allergic effector activity that is more relevant to allergic reactions. METHODS AND RESULTS: Ara h 6 was expressed as a recombinant protein in both Escherichia coli and Pichia pastoris (rAra h 6-E. coli and rAra h 6-Pichia, respectively). Effector activity was assayed by measuring degranulation of RBL SX-38 cells sensitized with IgE from patients with severe peanut allergy. Compared to native Ara h 6 (nAra h 6), rAra h 6-Pichia had intact effector function whereas rAra h 6-E. coli had significantly reduced function. The lower effector activity in rAra h 6-E. coli compared to nAra h 6 and rAra h 6-Pichia did not appear to be due to differences in posttranslational modifications (analyzed by mass spectrometry and staining for carbohydrates) and may be due to subtle alteration(s) of folding seen on CD analysis and on nonreduced gels. Finally, we introduced point mutations in four important IgE-binding linear epitopes of Ara h 6 and found dramatically reduced allergic effector activity. CONCLUSION: Our studies demonstrate the utility of fully functional rAra h 6-Pichia as a starting point for analysis of specific mutations that adversely affect allergic effector function.


Assuntos
Albuminas 2S de Plantas/biossíntese , Antígenos de Plantas/biossíntese , Basófilos/imunologia , Escherichia coli/metabolismo , Proteínas Mutantes/biossíntese , Hipersensibilidade a Amendoim/imunologia , Pichia/metabolismo , Albuminas 2S de Plantas/química , Albuminas 2S de Plantas/genética , Animais , Antígenos de Plantas/química , Antígenos de Plantas/genética , Teste de Degranulação de Basófilos , Linhagem Celular , Dicroísmo Circular , Células Clonais , Epitopos/química , Humanos , Imunoglobulina E/análise , Proteínas Mutantes/química , Proteínas Mutantes/genética , Hipersensibilidade a Amendoim/sangue , Mutação Puntual , Conformação Proteica , Dobramento de Proteína , Ratos , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo
18.
Ann Fr Anesth Reanim ; 30(3): 294-304, 2011 Mar.
Artigo em Francês | MEDLINE | ID: mdl-21392931
19.
Analyst ; 136(17): 3461-8, 2011 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-21293790

RESUMO

Automated microchip electrophoresis was used as a simple and rapid method to measure effective dissociation constants (K(d,eff)) of aptamers against both large and small molecule targets. Human thrombin, immunoglobulin E (IgE), and adenosine triphosphate (ATP) were selected as model analytes to validate the method, with four ligands including two DNA aptamers for thrombin (two distinct epitopes), an IgE aptamer, and an ATP aptamer. The approach is based on a microchip version of a DNA mobility shift assay. Non-denaturing microchip gel electrophoresis separations of DNA could resolve and quantify unbound from target-bound aptamers when using large molecules as targets. To extend the technique to small molecule targets such as ATP, an aptamer/competitor strategy was used, in which a DNA competitor complementary to the aptamer could be displaced by ATP and electrophoretically resolved. Using an automated microchip electrophoresis platform, parallel separations of 11 titration samples were completed in ~0.5 h. Analytical performance comparisons show that our approach provides significant advantages in minimized reagent consumption (typically tens of pmol of aptamer and target), reduced analysis time, and minimized user interaction when compared to previously reported methods for aptamer K(d) measurement. Moreover, the flexibility and ease of K(d,eff) measurement for aptamers against large and small targets make this a unique and valuable approach that should find widespread use. Finally, the feasibility of using this method during aptamer selection processes (e.g. SELEX) was shown by accurate bulk K(d,eff) measurement of a known thrombin aptamer (THRaptA) spiked into a random-sequence DNA pool at as low as 5.0% (molar %) of the total pool; only ~825 fmol of total binding sequences were needed for an 11-point titration curve.


Assuntos
Trifosfato de Adenosina/análise , Aptâmeros de Nucleotídeos/química , Eletroforese em Microchip/métodos , Imunoglobulina E/análise , Trombina/análise , Eletroforese em Microchip/economia , Humanos , Técnica de Seleção de Aptâmeros , Sensibilidade e Especificidade , Fatores de Tempo
20.
Ital J Pediatr ; 36: 61, 2010 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-20836868

RESUMO

BACKGROUND: Allergy places a considerable cost burden on society. Specific immunoglobulin E (spIgE) testing may improve the management of allergy patients. There is therefore a reason to quantify the economic consequences of the use of spIgE testing in the diagnosis of allergic conditions. METHODS: The expected costs of spIgE testing versus no-testing were calculated using a clinical decision model based on a prospective clinical trial performed in primary care. RESULTS: The expected costs per patient over 2 years decreased from 802 euros in the "no-test strategy" to 560 euros in the spIgE "test strategy". Cost savings persisted even after assumptions about the prevalence of allergy and the prices of medications were changed. The "test strategy" increased the percentage of patients correctly diagnosed from 54 to 87%. CONCLUSIONS: spIgE testing of children with respiratory and/or skin problems in primary care in Italy reduces overall costs to society. These cost savings mostly result from a reduction in the use of medications, particularly corticosteroids. The study indicates that spIgE testing of all children with respiratory and/or skin symptoms would be a cost-effective strategy.


Assuntos
Custos de Cuidados de Saúde/tendências , Hipersensibilidade/diagnóstico , Imunoglobulina E/análise , Atenção Primária à Saúde/economia , Adolescente , Criança , Pré-Escolar , Análise Custo-Benefício , Feminino , Humanos , Hipersensibilidade/economia , Hipersensibilidade/imunologia , Lactente , Recém-Nascido , Itália , Masculino , Estudos Prospectivos
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