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110 Partial characterization of Fraxinus chinensis (Urapan) allergens and cross-reactivity among different Fraxinus species and Olea europea (OLIVE)
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110
Abstracts
J ALLERGYCLIN IMMUNOL JANUARY 1996
Partial characterization of the silk allergens in mulberry silk e x t r a c t . W Zaoming*, MD, R Codina, MS, E Fernandez-Caldas, Ph D, R F Lockey MD and S C 8ukanz, MD. University of South Florida and V.A. Hospital, Tampa, FI and *Peking Union Medical College, Beijing, China. Although there have been reports of allergic reactions, asthma and hypersensitivity pneumonitis to silk allergens, especially in oriental countries, these allergens have not been characterized. We report an in vivo and in vitro study of subjects allergic to silk, as well as a partial characterization of the allergens involved. Forty one subjects with a clinical history of silk allergy who had asthma, allergic rhinitis or both and who had a positive intradermal skin test to mulberry silk extract (MSE) (wheal response to MSE larger than the 1 histamine equivalent test) were evaluated. Four asthmatic subjects with a negative skin test and RAST to MSE were used as controls. Twenty two of the 41 subjects had conjunctival provocation with a MSE. Twenty one of 22 subjects (96%) had a positive test. RAST specific IgE was present in 37 of 41 (90%) subjects. SDS-PAGE of MSE demonstrated 13 protein bands of less than 21 to 95 KD. IgE wich bound to protein bands was present in 22 of 32 (68%) sera. Fifteen of wich reacted to a protein band with a MW between 35.1 and 50 KD. Allergic silk patients have in vivo and in vitro evidence of sensitivity to MSE but some sera do not have antibodies which react with the MSE immunoblot protein bands. The low avidity of the IgE is probably responsible for this dicrepancy between in vivo and in vitro results.
111
PARTIALCHARAC'IERIZATION OF Frax/nu$ chinensis (URAPAN)ALLERGENSAND CRO6S-REACTIVITYAMONG DIFFERENTFraxlnus SPECIES AND 0/ea europea (OLIVEI.
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A Rodriguez*, R Codina, E Femandez-Caldas, F C Leal*and R F Lockey. Unlverslty of South Florida and VA Hospital, Tampa, Fl and *Hospital Lorencita Vfllegas de Santos. Bogota, Colombia. Fraxlnus chlnertsts (FcL a common tree in Bogota, is in the
Oleaceae family. Whether Its pollen is an aeroallergen has not been Investigated, but genuses of the same family, such Olea europea (Oct are sources of important aeroaUergens. The purpose of this study was to characterize the Fc pollen allergens by SDS-PAGE and IEF/Western blot and to ascertain whether there is cross-reacthrlty among Fc, Oe, F. velutlna (Fv), F. petxnsylvan[ea (Fp), F. latlfolia (Fl), and F. americana {Fa) (Greer Laboratories, Lcnoir, NC). Sera were obtained from 6 subjects who live In Bogota and who reacted positively to a prick skin test to Fc extract (wheal > 3 ram); 2 had asthma and 4 allergic rhinitis. SDS-PAGE of the Fc extract demontrated 12 protein bands wlth MWs from less than 21.9 to 142.9 KD. The serum from subject # I recognized 4 bands ranging from less than 21.9 to 50 KD and the sera from subject #2 recognized 4 bands ranging from 50 to 142.9 KD. The sera from the other 4 subjects did not react on the SDS-PAGEIWesteru blot. IEF of the FC extract demonstrated 12 bands in the pH range of 3,5 to 7.35, 7 of which reacted with IgE in tile serum of subject # 1. Cross. inhibition studies indicate there are similar allergens in P~, Fv, Fl and r a extracts and a partial cross-reaetlvlty with and Oe. This study demonstrates that Fo pollen allergens contains 7 or more IgE binding proteins. These allergens are present in smaller amounts in other Fraxinus species and some share common epltopes with the Oe allergens.
Allergens of horse dander. Comparison between breeds and individual animals using immunoblotting. Drebor~ S MD PhD. Felix K PhD. Ferr~ndiz R PhD. ~inarsson R PhD. Linkt~ping, Sweden Some patients report allergic symptoms when exposed to certain animals or breeds and not to other breeds or individuals but there are no reports on breed specific allergens in horse dander. Bashkir horses have been claimed to be non-allergenic.We investigated the variation in allergeniccomposition of dander extracts from various horse breeds and individual horses. Twentyfour extracts were prepared,23 from individual animals from seven breeds and one extract from a mixture of dander from four animals of the Bashkir breed. The allergeniccompositionwas compared by SDS-PAGE and immunoblotting.Nineteen allergenic components with molecular weights ranging between 14 and 130 kD were detected. The 14 kD, 19 kI), 27 kD, and 39 kD proteins were present in all extracts and bound IgE from more than 50 % of sera tested. There was marked variation in allergenic composition, both within and betweenbreeds, but no breed-specificallergens were identified. We did not find any breed-specificallergensin horse dander. Dander from all horse breeds investigatedcontainedthe most important allergens.We bypothesise that differences in scale production could explain the differences in sensitivityto breeds and individuals noted by patients.
Study of B e r m u d a Pollen I g E - B i n d i n g Protein, BG60: Characterization o f Its Physicochemical Property. S___NN S___uu.PhD, Taipei, TWN; SW Huang MD, Gaineville FL; Y_._C_C Lee PhD, Bahimore M D Bermuda grass pollen contains multiple lgE-binding protein molecules. One o f the components with molecular weight o f 60 KD, designated BG60, has been shown to contain isoforms. To investigate its physicoehemieal nature, we purified BG60 from pollen by a combination o f ion-exchange chromatography, gel filtration chromatography, Blue gel chromatography and reverse-phase HPLC. EL1SA assay and immunoblot techniques were used to determine antibody binding. Enzyme digestion and compositional analysis were used to study the chemical structure. The result showed: 1). both native and denatured antigen react with human IgE and lgG, 2). the antigen contains most o f the natural amino acids and is rich in lysine and arginine, 3). the antigen is bound by plant iectins, 4). some oligosaccharides could be cleaved off' from the native molecules and 5). carbohydrate moiety contains mostly mannose, N-acetylglucosamine and fucose. We conclude: 1). aUergenicity and antigenicity o f the BG60 do not totally depend on its conformationai structure, 2). BG60 is a glycoprotein, 3). some carbohydrate is bound to peptide backbone through N-linkage.
109
110
Abstracts
J ALLERGYCLIN IMMUNOL JANUARY 1996
Partial characterization of the silk allergens in mulberry silk e x t r a c t . W Zaoming*, MD, R Codina, MS, E Fernandez-Caldas, Ph D, R F Lockey MD and S C 8ukanz, MD. University of South Florida and V.A. Hospital, Tampa, FI and *Peking Union Medical College, Beijing, China. Although there have been reports of allergic reactions, asthma and hypersensitivity pneumonitis to silk allergens, especially in oriental countries, these allergens have not been characterized. We report an in vivo and in vitro study of subjects allergic to silk, as well as a partial characterization of the allergens involved. Forty one subjects with a clinical history of silk allergy who had asthma, allergic rhinitis or both and who had a positive intradermal skin test to mulberry silk extract (MSE) (wheal response to MSE larger than the 1 histamine equivalent test) were evaluated. Four asthmatic subjects with a negative skin test and RAST to MSE were used as controls. Twenty two of the 41 subjects had conjunctival provocation with a MSE. Twenty one of 22 subjects (96%) had a positive test. RAST specific IgE was present in 37 of 41 (90%) subjects. SDS-PAGE of MSE demonstrated 13 protein bands of less than 21 to 95 KD. IgE wich bound to protein bands was present in 22 of 32 (68%) sera. Fifteen of wich reacted to a protein band with a MW between 35.1 and 50 KD. Allergic silk patients have in vivo and in vitro evidence of sensitivity to MSE but some sera do not have antibodies which react with the MSE immunoblot protein bands. The low avidity of the IgE is probably responsible for this dicrepancy between in vivo and in vitro results.
111
PARTIALCHARAC'IERIZATION OF Frax/nu$ chinensis (URAPAN)ALLERGENSAND CRO6S-REACTIVITYAMONG DIFFERENTFraxlnus SPECIES AND 0/ea europea (OLIVEI.
112
A Rodriguez*, R Codina, E Femandez-Caldas, F C Leal*and R F Lockey. Unlverslty of South Florida and VA Hospital, Tampa, Fl and *Hospital Lorencita Vfllegas de Santos. Bogota, Colombia. Fraxlnus chlnertsts (FcL a common tree in Bogota, is in the
Oleaceae family. Whether Its pollen is an aeroallergen has not been Investigated, but genuses of the same family, such Olea europea (Oct are sources of important aeroaUergens. The purpose of this study was to characterize the Fc pollen allergens by SDS-PAGE and IEF/Western blot and to ascertain whether there is cross-reacthrlty among Fc, Oe, F. velutlna (Fv), F. petxnsylvan[ea (Fp), F. latlfolia (Fl), and F. americana {Fa) (Greer Laboratories, Lcnoir, NC). Sera were obtained from 6 subjects who live In Bogota and who reacted positively to a prick skin test to Fc extract (wheal > 3 ram); 2 had asthma and 4 allergic rhinitis. SDS-PAGE of the Fc extract demontrated 12 protein bands wlth MWs from less than 21.9 to 142.9 KD. The serum from subject # I recognized 4 bands ranging from less than 21.9 to 50 KD and the sera from subject #2 recognized 4 bands ranging from 50 to 142.9 KD. The sera from the other 4 subjects did not react on the SDS-PAGEIWesteru blot. IEF of the FC extract demonstrated 12 bands in the pH range of 3,5 to 7.35, 7 of which reacted with IgE in tile serum of subject # 1. Cross. inhibition studies indicate there are similar allergens in P~, Fv, Fl and r a extracts and a partial cross-reaetlvlty with and Oe. This study demonstrates that Fo pollen allergens contains 7 or more IgE binding proteins. These allergens are present in smaller amounts in other Fraxinus species and some share common epltopes with the Oe allergens.
Allergens of horse dander. Comparison between breeds and individual animals using immunoblotting. Drebor~ S MD PhD. Felix K PhD. Ferr~ndiz R PhD. ~inarsson R PhD. Linkt~ping, Sweden Some patients report allergic symptoms when exposed to certain animals or breeds and not to other breeds or individuals but there are no reports on breed specific allergens in horse dander. Bashkir horses have been claimed to be non-allergenic.We investigated the variation in allergeniccomposition of dander extracts from various horse breeds and individual horses. Twentyfour extracts were prepared,23 from individual animals from seven breeds and one extract from a mixture of dander from four animals of the Bashkir breed. The allergeniccompositionwas compared by SDS-PAGE and immunoblotting.Nineteen allergenic components with molecular weights ranging between 14 and 130 kD were detected. The 14 kD, 19 kI), 27 kD, and 39 kD proteins were present in all extracts and bound IgE from more than 50 % of sera tested. There was marked variation in allergenic composition, both within and betweenbreeds, but no breed-specificallergens were identified. We did not find any breed-specificallergensin horse dander. Dander from all horse breeds investigatedcontainedthe most important allergens.We bypothesise that differences in scale production could explain the differences in sensitivityto breeds and individuals noted by patients.
Study of B e r m u d a Pollen I g E - B i n d i n g Protein, BG60: Characterization o f Its Physicochemical Property. S___NN S___uu.PhD, Taipei, TWN; SW Huang MD, Gaineville FL; Y_._C_C Lee PhD, Bahimore M D Bermuda grass pollen contains multiple lgE-binding protein molecules. One o f the components with molecular weight o f 60 KD, designated BG60, has been shown to contain isoforms. To investigate its physicoehemieal nature, we purified BG60 from pollen by a combination o f ion-exchange chromatography, gel filtration chromatography, Blue gel chromatography and reverse-phase HPLC. EL1SA assay and immunoblot techniques were used to determine antibody binding. Enzyme digestion and compositional analysis were used to study the chemical structure. The result showed: 1). both native and denatured antigen react with human IgE and lgG, 2). the antigen contains most o f the natural amino acids and is rich in lysine and arginine, 3). the antigen is bound by plant iectins, 4). some oligosaccharides could be cleaved off' from the native molecules and 5). carbohydrate moiety contains mostly mannose, N-acetylglucosamine and fucose. We conclude: 1). aUergenicity and antigenicity o f the BG60 do not totally depend on its conformationai structure, 2). BG60 is a glycoprotein, 3). some carbohydrate is bound to peptide backbone through N-linkage.