Identification of <i>Parietaria judaica</i> Pollen Allergens

Abstract

Parietaria judaica pollen allergens, fractionated by SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes, were identified using 52 sera collected in Australia and Sicily from P. judaica pollen-allergic patients. IgE-binding pollen components transferred to nitrocellulose were detected by reaction with ¹²⁵I-anti-human IgE and autoradiography. Nine pollen components, ranging in molecular weight (MW) from approximately 10,000 to 80,000 daltons, bound IgE antibodies but the two fastest migrating components sometimes each separated into two very closely migrating bands. The faster of the two components exhibiting doublet formation (MW approximately 10,000 daltons) showed by far the highest frequency of IgE binding, being recognised by 50 of the 52 sera examined. Although patients’ IgE reaction patterns to P. judaica allergens were heterogeneous, the degree of heterogeneity was much less than that observed with house dust mite and other pollen extracts studied by electrophoretic transfer analysis. Results with gradient gel-nitrocellulose transfer experiments which showed no IgE-binding components with MWs less than 70,000 daltons, and comparisons of our electroblotting results with crossed radioimmunoelectrophoresis results of others, suggested that the doublet proteins with MWs of approximately 10,000 probably bind to higher MW proteins in P. judaica pollen extracts.

cDNA cloning, expression and primary structure of Par j I, a major allergen of Parietaria judaica pollen

“…The P5 clone also showed a 185 bp 3' untraslated region with a canonical poly(A)' tail at the end. The molecular weight of the P5 protein is of 14,866 kDa which corresponds to the previously detected molecular weight of the native Par j I allergen [16]. To confirm that the P5 clone represented a full length clone, a Northern blot from total and poly(A)' RNAs from Pj flowers has been performed.…”

“…Lane B shows a total cell lysates from recombinant P5 clone without induction, while on Lane A a pool of protein markers (range 45-200 kDa) was used. At least 9 of them were capable of binding specific human IgE and 4 were classified as major allergens [15][16]. To evaluate which of them we had isolated, we performed an immunoblot screened with two monoclonal antibodies specific for the Par j I allergen [17,25] and the result is reported in Fig.…”

Detection of IgE Antibodies to a Wide Range of Insect Species in Subjects with Suspected Inhalant Allergies to Insects

“…Sodium dodecyl sulphate (SDS) polyacrylamide gel electro phoresis (PAGE) was carried out as previously describd [Tovey and Baldo, 1984;Ford et al, 1986] using 9-27% gradient acrylamide gels. High and low MW markers (Bio-Rad or Pharmacia) were in cluded in each electrophoretic separation.…”

A Re-examination of Ryegrass <i>(Lolium perenne) </i>Pollen Allergens

“…Because of this uncertainty about the identity or otherwise of the various allergenic components iden tified in, or isolated from, ryegrass pollen, and in view of the recent success of applying electroblotting to the identification of allergens in complex mixtures Baldo, 1984, 1985;Ford et al" 1985Ford et al" , 1986, we have applied the method to critically re-examine the allergenic spectrum of pollen from this species. Central to this study were the aims of defining a standard reference pattern that can readily be repro duced and interpreted in different laboratories and to apply the method to defining individual patient al lergen recognition patterns.…”