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Egg is one of the major allergens with specific labeling requirements. α-livetin, which also called chicken serum albumin (CSA), is one of the major allergens in egg need to be detected in food matrix. A monoclonal antibody (mA) specific to α-livetin was developed, but the property of this mAb is not clear. The relationship between matrix effect, extractability of α-livetin and their thermostability during in vitro study need to be elaborated. Our research aims (1) to characterize this mAb that is specific for α-livetin; (2) to develop a novel extraction buffer for α-livetin (CSA) in egg yolk and chicken blood; (3) we hypothesis matrix-induced thermal instability of α-livetin (CSA) because of hydrophobic effect (hydrophobic effect) and chemical interaction (thiol-disulfide interchange) interaction. mAb was purified from the supernatant using immunoaffinity. Indirect non-competitive ELISA was performed to study the selectivity of mAb. Two-dimensional gel electrophoresis was performed to further investigate the isoelectric point (pI). SDS PAGE was performed to study the molecular integrity and solubility of the target protein with different pH conditions. BCA assay was performed to study the solubility of the target protein with different heating conditions. Western blot was performed to study the mAb selectivity; to verify the target protein’s molecular weight; to optimized the extractability of the extraction buffer; to investigate antigenicity of the target protein under extraction buffer with different pH conditions; to test the antigenicity of the target protein under different heat treatment conditions. As to the results of this study, the target protein of the mAb is α-livetin (chicken serum albumin) (70 kDa). As for buffer selection, on one hand, detergent increased the solubility of the target protein, on the other hand, based on the property of this mAb reducing reagent was required to cleave the disulfide bond of α- livetin to enhance antigenicity intensity. As for pH effect, α-livetin remained its antigenicity under neutral condition after heat treatment in the novel extraction buffer, and its immunoreactivity did not change significantly (P > 0.05) after heat treatment. Finally, a novel extraction buffer (10mM DTT with 0.1% SDS in PBS under neutral pH) was developed. The target protein was successfully isolated. The target’s antigenicity reaction with the mAb decreased after heat treatment was confirmed.