Identification and analysis of basidiospore allergens from puffballs

Ibáñez, María Dolores; Horner, W. Elliott; Liengswangswong, V.; Sastre, J.; Lehrer, Samuel B.

doi:10.1016/0091-6749(88)90080-2

Cited by 15 publications

(8 citation statements)

References 17 publications

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“…In both previous reports of C.cyathiformis aller gens [10,11], it was unclear whether Calc Bd9.3 was lost by fractionation or due to lability. The presence of Calc Bd9.3 in the HIC fraction at zero time indi cates that the allergen does persist through both fractionation steps, although reduced in amount.…”

Section: Discussionmentioning

confidence: 99%

“…A previous report from our laboratory established that crude and fractionated C. cyalhiformis basidio- spore extracts were capable of binding IgE from pooled sera of atopic individuals [10]. We also charac terized the reactivity of these allergens with individual sera [II], Some variability existed in these studies re garding allergen presence at different stages of frac tionation, notably allergen Calc Bd9.3 [11].…”

Section: Introductionmentioning

confidence: 99%

“…HIC was conducted with the radioallergosorbent test (RAST)-active G-75 pool, corresponding to 'pool 3' of Ibanez et al [10]. Lyophilized material from this pool was resuspended in 0.50 M Tris buffer.…”

Section: Chromatographymentioning

confidence: 99%

“…Chromatographic separations were performed as described pre viously [10], Briefly, spore extract was fractionated by gel filtration and hydrophobic interaction chromatography [HIC). Gel filtration (GF) was performed in a 5x 100 cm column with Sephadex G-75 (Pharmacia.…”

Section: Chromatographymentioning

confidence: 99%

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Stability Studies of Calvatia cyathiformis Basidiospore Allergens

Horner

Ibáñez²,

Sb³

1989

Int Arch Allergy Immunol

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Calvatia cyathiformis allergens in unfractionated extract (crude), and in extract sequentially fractionated by gel filtration (GF) and hydrophobic interaction chromatography (HIC) were tested for stability. C. cyathiformis allergen sources (crude, GF, HIC) were sampled immediately (0 h), or incubated at 4, 24 or 37 °C and then sampled at 8, 24 or 96 h. Polyacrylamide gel-isoelectric focusing immunoprints revealed 3 allergen(s) groups, or bands (Bds) with respective pi of 3.6–4.6, 6.6 and 9.3. Only Bds 3.6–4.6 were stable at 37 °C. At 24°C, Bds3.6–4.6 persisted to 96 h, Bd6.6 persisted 24 h, and Bd9.3 waned in 8 h. At 4°C all 3 allergens in HIC were stable for 8 and 24 h; Bd9.3 was reduced at 96 h. All allergen activity was labile to low pH conditions except for Bds3.6–4.6. Proteinase K degraded Bd9.3 more rapidly than Bd6.6. Immunoprint patterns corresponded to the stained gels and were consistent among different sources. Bd9.3 is very labile, but reactive with 63% of sera tested. Since 10–15% of C. cyathiformis reactors bind IgE only to Bd9.3, this is notable variability, and significant for diagnosis and treatment.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Chromatographymentioning

confidence: 99%

Section: Chromatographymentioning

confidence: 99%

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Stability Studies of Calvatia cyathiformis Basidiospore Allergens

Horner

Ibáñez²,

Sb³

1989

Int Arch Allergy Immunol

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“…The cath odic fraction eluates were collected from several runs, pooled and refocused, again in a gel slurry, final pH 5.4-10.4. The eluants of 9.3 containing fractions were then passed over a phenyl-Sepharose col- umn (HIC) to remove ampholytes [5]. Column fractions containing protein were dialyzed, aliquoted and held frozen at -70°C .…”

Section: Methodsmentioning

confidence: 99%

Basidiomycete Allergy: Identification and Characterization of an Important Allergen from Calvatia cyathiformis

Horner

Lopez²,

Salvaggio³

et al. 1991

Int Arch Allergy Immunol

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Basidiomycetes were not considered as major aeroallergen sources until spore traps revealed the prevalence of basidiospores, which were in some cases associated with epidemic outbreaks of asthma. More recently, we established that approximately one third of subjects with respiratory allergic disease were skin prick test positive to basidiospore extracts. Bronchial challenge with these extracts induced reversible immediate and late phase bronchospasm in sensitive subjects. Screening individual RAST-positive sera for reactivity to immunoprinted Calvatia cyathiformis spore extract indicated that a basic component with pI 9.3 reacted with 63% of test sera. Further analysis by RAST and immunoprint (IP) inhibition showed that this component, Calc Bd9.3, appeared to cross-react with spore extracts from 3 of 5 other species tested. To isolate this component for additional analysis, a protocol was used with two sequential stages of preparative isoelectric focusing (IEF) followed by hydrophobic interaction chromatography. This yielded purified Cal c Bd9.3 that retained IgE-binding activity by IP, was a single band by IEF and SDS-PAGE (16 kD) analysis when stained with Coomassie blue. Double diffusion in gel with rabbit antiserum to Cal c BD9.3 demonstrated a single precipitin band in crude extract that was identical to that obtained with purified Cal c Bd9.3. By IP this antiserum recognized pure Cal c Bd9.3 and the 9.3 band in crude extracts from C. cyathiformis and 3 other species, corroborating the cross-reactivity. However, other bands were also recognized. These studies indicate that basidiospores are major fungal aeroallergens, many of which contain a common basic allergen to which most sensitized subjects react.

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