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Quantification of Der f 1 in houses of patients allergic to house dust mite, Dermatophagoides farinae, using a locally produced detection reagents.

AbstractBACKGROUND:
House dust mite (HDM) allergen quantification in house dust samples before and after the allergen elimination is one means of convincing the target population about the health benefits of allergen removal from their environment.
OBJECTIVE:
To produce local reagents for quantification of Der f 1 (major allergen of Dermatophagoides farinae) in dust samples from houses of HDM allergic Thai patients.
METHODS:
Recombinant Der f 1 was used for immunization of a BALB/c mouse for hybridoma production. Polyclonal antibody (PAb) to whole body extract of D. farinae was prepared from an immunized rabbit. A sandwich ELISA (MAb-allergen-PAb) was used, in comparison with the commercialized reagents (Indoor Biotechnology, UK), to quantify Der f 1 in dust samples.
RESULTS:
Two hybridoma clones, Dfl-1 and Dfl-2, were established. Their secreted MAbs (MAbDfl-1 and MAbDfl-2, respectively) bound to the homologous antigen as well as native Der f 1 and a crude extract of D. farinae. Epitopes of MAbDfI-1 and MAbDfl-2 were located at amino acid residues 206NSQHYGISNYCQ217 and 283DYW---NSWD-WGDSG298 of Der f 1. MAbDf-1 had higher affinity to Der f 1 than the MAbDfl-2. A sandwich ELISA (MAbDfl-1-allergen-PAb) and commercialized reagents (MAbl-allergen-MAb2 sandwich ELISA) were used in comparison for quantification of Der f 1 in 42 dust samples collected from bedrooms and living rooms of 21 houses of the HDM allergic patients. All of the 42 dust samples measured by both ELISAs had the Der f 1 levels higher than 2 mg per gram of fine dust which is the HDM allergy sensitizing level. In addition, Der f 1 levels in 41 samples (except 1 sample from a living room) measured by the MAbDfI-1-PAb and MAbl-MAb2 sandwich ELISAs were higher than 10 mg per g of dust which is the morbidity level of HDM allergen. The local sandwich ELISA showed a high coefficient correlation (r = 0.91) in measuring known amounts of recombinant and native Der f 1. The results indicate that the reagents produced in the present study can be used for measuring the environmental levels of HDM Der f 1. The assay can also be used for standardization of the HDM extract for monitoring patient's allergenic status or for immunotherapeutic purpose.
AuthorsNitat Sookrung, Thanyarat Kamlanghan, Nitaya Indrawattana, Anchalee Tungtrongchitr, Pongsakorn Tantilipikorn, Chaweewan Bunnag, Kovit Pattanapanyasat, Wanpen Chaicumpa
JournalAsian Pacific journal of allergy and immunology / launched by the Allergy and Immunology Society of Thailand (Asian Pac J Allergy Immunol) Vol. 29 Issue 1 Pg. 78-85 (Mar 2011) ISSN: 0125-877X [Print] Thailand
PMID21560492 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References
  • Antigens, Dermatophagoides
  • Arthropod Proteins
  • Epitopes
  • Indicators and Reagents
  • Cysteine Endopeptidases
  • Dermatophagoides farinae antigen f 1
Topics
  • Amino Acid Sequence
  • Animals
  • Antigens, Dermatophagoides (analysis, immunology)
  • Arthropod Proteins
  • Cysteine Endopeptidases
  • Dermatophagoides farinae (immunology)
  • Environmental Monitoring (methods)
  • Enzyme-Linked Immunosorbent Assay
  • Epitopes (immunology)
  • Humans
  • Hybridomas
  • Hypersensitivity
  • Indicators and Reagents
  • Mice
  • Mice, Inbred BALB C
  • Molecular Sequence Data
  • Rabbits
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Sequence Alignment

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