title: Cockroach allergen detection and cockroach allergens of allergic Thai patients.
abstract: Hybridomas secreting monoclonal antibodies (MAb) specific to American cockroach (Periplaneta americana) were produced through a fusion of immune splenocytes of a BALB/c mouse immunized with crude cockroach (CR) extract and mouse myeloma cells. Two hybridomas namely 38G6 and 3C2 were established. These specific hybridomas secreted IgG1 monoclonal immunoglobulins with antigenic specificities to CR protein components of over 207 to 72 kDa and 45 to 40 kDa, respectively. The monoclonal antibodies were applied to select their specific epitopes out of the crude CR extract using affinity chromatography. A Prausnitz-Kustner test revealed that these epitopes were allergens which caused wheals and flares of the skin of a guinea-pig previously sensitized with a pool of serum samples from CR allergic patients. The monoclonal antibodies were also used in a capture ELISA to detect specific IgE in serum samples of allergic Thai patients. It was found that 72% and 76% of the patients had IgE antibodies to the epitopes of MAb 38G6 and MAb 3C2, respectively, indicating that the two epitopes are major CR allergens among the CR allergic Thai patients. An antibody-sandwich ELISA was developed for quantitative detection of CR allergens using the two monoclonal antibodies as a capture reagent and rabbit polyclonal antibodies to crude CR extract as a detection reagent. The assay could detect allergenic epitopes contained in as little as 122 pg of crude cockroach extract, and has high potential for direct measurement of the marker allergens in extracts of environmental samples.
authors.affiliation: Faculty of Allied Health Sciences, Thammasat University, Rangsit Center, Pathum-thani, Thailand.
authors.forename: Nitat
authors.initials: N
authors.lastname: Sookrung
authors.forename: Pornphan
authors.initials: P
authors.lastname: Diraphat
authors.forename: Wanpen
authors.initials: W
authors.lastname: Chaicumpa
authors.forename: Pongsri
authors.initials: P
authors.lastname: Tongtawe
authors.forename: Yuwaporn
authors.initials: Y
authors.lastname: Sakolvaree
authors.forename: Pramuan
authors.initials: P
authors.lastname: Tapchaisri
authors.forename: Vanna
authors.initials: V
authors.lastname: Mahakittikun
authors.forename: Anchalee
authors.initials: A
authors.lastname: Tungtrongchitr
authors.forename: Pakit
authors.initials: P
authors.lastname: Vichyanond
authors.forename: Chaweewan
authors.initials: C
authors.lastname: Bunnag
mesh_terms: D000485:Allergens / Q000276:immunology* / Q000302:isolation & purification*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002648:Child; D003058:Cockroaches / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006168:Guinea Pigs; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D006967:Hypersensitivity / Q000276:immunology*; D007428:Intradermal Tests / Q000379:methods; D051379:Mice; D011817:Rabbits; D013785:Thailand
chemical_list: D000485:Allergens; D000911:Antibodies, Monoclonal