Quantification assay for the major allergen of cupressus sempervirens pollen, cup s 1, by sandwich elisa

التفاصيل البيبلوغرافية
العنوان: Quantification assay for the major allergen of cupressus sempervirens pollen, cup s 1, by sandwich elisa
المؤلفون: Alberto Martínez, M. C. Arilla, I. Ibarrola, Juan A. Asturias
المصدر: Allergologia et Immunopathologia. 32:319-325
بيانات النشر: Codon Publications, 2004.
سنة النشر: 2004
مصطلحات موضوعية: Pulmonary and Respiratory Medicine, Streptavidin, Low protein, Immunology, Enzyme-Linked Immunosorbent Assay, medicine.disease_cause, Sensitivity and Specificity, chemistry.chemical_compound, Allergen, Pollen, Cupressus sempervirens, Botany, medicine, Immunology and Allergy, Plant Proteins, Polysaccharide-Lyases, Detection limit, Chromatography, biology, Reproducibility of Results, General Medicine, Allergens, Antigens, Plant, Cupressus, Reference Standards, biology.organism_classification, Standard curve, chemistry, Pectate lyase
الوصف: Background: The Cupressaceae are an important cause of pollinosis, particularly in Mediterranean countries. Cypress pollen allergenic extracts are difficult to produce since they have a low protein and a high carbohydrate content and consequently accurate standardization of these extracts is essential for diagnosis and immunotherapy. Method: Natural Cup s 1 was purified by a combination of hydrophobic interaction, gel filtration and ion exchange chromatographies and its enzymatic activity was analyzed. The allergen was used as reference material in the ELISA standard curve. The assay was based on a specific monoclonal antibody (3D2) immobilized on ELISA plates and used to capture Cup s 1. Bound proteins were detected by a combination of biotinylated specific antiserum and peroxidase-conjugated streptavidin. Results: Purified Cup s 1 is a functional pectate lyase enzyme with a specific activity of 750 U/mg protein. The developed ELISA measured Cup s 1 concentrations ranging from 31.25 to 250 ng/ml in the lineal portion of the standard curve. The intra-assay and inter-assay variation coefficients in the working range were less than 8.1 % and 16 %, respectively. The assay was highly sensitive, with a detection limit of 3.8 ng/ml. The dose-response curves obtained with C. sempervirens pollen extracts and extracts belonging to other species from the Cupressaceae family showed a good parallelism compared with those obtained using the purified allergen, indicating that the same protein was measured. Conclusions: The assay described is sensitive, specific and reproducible for the quantification of Cup s 1 in C. sempervirens pollen extracts for clinical use. This ELISA could also be useful for other Cupressaceae-related pollen extracts.
تدمد: 0301-0546
URL الوصول: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::122863036e9e6a1cc557dd754da428b1
https://doi.org/10.1016/s0301-0546(04)79263-0
حقوق: CLOSED
رقم الأكسشن: edsair.doi.dedup.....122863036e9e6a1cc557dd754da428b1
قاعدة البيانات: OpenAIRE