Identification and characterisation of cat allergens

[Truncated abstract] Background While cats are known to produce at least 10 allergenic proteins, only Fel d 1 has been extensively characterised. Recent studies have shown that some cat allergic subjects have high IgE binding to Fel d 1 but for many patients the titre is low. The purpose of this study is to identify and characterise new cat allergens, and determine their clinical significance. Methods Several methods were used to characterise three potential allergens. The allergen named as Fel d 8 was identified and produced using an immunoassay to screen a cat mandibular cDNA library with human cat allergic sera. Natural haptoglobin was isolated from cat sera by a procedure based on ammonium sulphate precipitation. cDNA clones encoding Fel d 7 were isolated from cat cDNA libraries by DNA hybridisation using a probe made by PCR with primers designed to amplify the homologue of a known dog allergen, Can f 1. Recombinant Fel d 8 was expressed in Escherichia coli and purified by chromatography. The IgE binding activities of Fel d 8 and haptoglobin for a panel of sera from allergic and non-allergic subjects were determined by dissociation enhanced lanthanide fluorescence immunoassay (DELFIA®) and compared to the binding by five allergens; Fel d 1, Fel d 2, Fel d 3, Fel d 4 and IgE binding S100A12. Results Three prospective cat allergens were isolated and characterised to various degrees. A cDNA from an IgE-reactive clone isolated from the mandibular library encoded a protein with a predicted molecular mass of 26 kDa. It had high sequence identity with breast cancer and salivary gland expression gene protein and horse latherin (an allergenic horse sweat protein). The recombinant polypeptide, Fel d 8, bound IgE in over 45% of 78 sera from subjects with cat allergy. The natural protein of haptoglobin was isolated from cat serum and bound IgE in 12% of the cat allergic subjects. The importance of these allergens was demonstrated by the finding that the DELFIA® as