title: A recombinant Sal k 1 isoform as an alternative to the polymorphic allergen from Salsola kali pollen for allergy diagnosis.
abstract: BACKGROUND
  The incidence of Amaranthaceae pollen allergy has increased due to the desertification occurring in many countries. In some regions of Spain, Salsola kali is the main cause of pollinosis, at almost the same level as olive and grass pollen. Sal k 1 - the sensitization marker of S. kali pollinosis - is used in clinical diagnosis, but is purified at a low yield from pollen. We aimed to produce a recombinant (r)Sal k 1 able to span the structural and immunological properties of the natural isoforms from pollen, and validate its potential use for diagnosis.
  
  
  METHODS
  Specific cDNA was amplified by PCR, cloned into the pET41b vector and used to transform BL21 (DE3) Escherichia coli cells. Immunoblotting, ELISA, basophil activation and skin-prick tests were used to validate the recombinant protein against Sal k 1 isolated from pollen. Sera and blood cells from S. kali pollen-sensitized patients and specific monoclonal and polyclonal antisera were used.
  
  
  RESULTS
  rSal k 1 was produced in bacteria with a yield of 7.5 mg/l of cell culture. The protein was purified to homogeneity and structural and immunologically validated against the natural form. rSal k 1 exhibited a higher IgE cross-reactivity with plant-derived food extracts such as peanut, almond or tomato than with pollen sources such as Platanus acerifolia and Oleaceae members.
  
  
  CONCLUSIONS
  rSal k 1 expressed in bacteria retains intact structural and immunological properties in comparison to the pollen-derived allergen. It spans the immunological properties of most of the isoforms found in pollen, and it might substitute natural Sal k 1 in clinical diagnosis.
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Universidad Complutense de Madrid, Madrid, Spain.
authors.forename: Salvador
authors.initials: S
authors.lastname: Mas
authors.forename: Patrice
authors.initials: P
authors.lastname: Boissy
authors.forename: Rafael I
authors.initials: RI
authors.lastname: Monsalve
authors.forename: Javier
authors.initials: J
authors.lastname: Cuesta-Herranz
authors.forename: Araceli
authors.initials: A
authors.lastname: Díaz-Perales
authors.forename: Javier
authors.initials: J
authors.lastname: Fernández
authors.forename: Carlos
authors.initials: C
authors.lastname: Colás
authors.forename: Rosalía
authors.initials: R
authors.lastname: Rodríguez
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Barderas
authors.forename: Mayte
authors.initials: M
authors.lastname: Villalba
mesh_terms: D000485:Allergens / Q000235:genetics / Q000302:isolation & purification; D052179:Antigens, Plant / Q000235:genetics / Q000302:isolation & purification; D015549:Basophil Degranulation Test; D003001:Cloning, Molecular; D003429:Cross Reactions; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism; D011058:Pollen / Q000235:genetics / Q000276:immunology*; D020033:Protein Isoforms / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D006255:Rhinitis, Allergic, Seasonal / Q000175:diagnosis* / Q000276:immunology*; D031248:Salsola / Q000235:genetics / Q000276:immunology*; D013030:Spain
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D020033:Protein Isoforms; D011994:Recombinant Proteins; D007073:Immunoglobulin E