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Objective: To prepare anti-human IgE nanobody by phage display technology, and to establish a method for hypersensitivity detection of cat dander specific IgE antibody. Methods: Allergen bio-information of cat was searched in WHO/IUIS Allergen Database. After synthesizing sequence, recombinant cat dander allergenic protein Fel d 1 was expressed and purified in prokaryotic expression system. Human IgE was used to immunize Bactrian camel and RNA were extracted from lymphocyte to construct phage display library. Library capacity, diversity and insertion rate were analyzed, anti-human IgE nanobody were obtained by screening and identification. A magnetic particle chemical method for cat dander specific IgE antibody detection was established using recombinant allergen-coupled magnetic particles and acridine ester-labeled nanobodies. Results: Capacity of phage display library was 1.88×108 cfu/ml, insertion rate was 93.6%, and purity of nanobody was >95%. Linear range of the method based on nanobody was 0.1~100 U/ml, who was consistent with ImmunoCAP detection system by clinical data. Conclusion: Nanobody-based cat dander specific IgE antibody hypersensitivity assay is successfully prepared, providing a technical basis for auxiliary diagnosis of cat allergic diseases. [ABSTRACT FROM AUTHOR]