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To develop the enzyme-linked immunosorbent assay(ELISA) for the analysis of N-acetylchitooligosaccharides(NACOS) and chitooligosaccharides (COS), specific antibodies(Abs) were produced, and their properties were compared.N-acetylchitohexaose (NACOS6), chitohexaose (COS6), andCOS mixture (COSM) conjugated to bovine serum albumin(BSA) were used to immunize rabbits. By the use of specificAbs and NACOS6-horseradish peroxidase (HRP), COS6-HRP, and COSM-HRP conjugates, competitive direct ELISA(cdELISA) was developed. The detection limits of NACOS6by the anti-NACOS6 Ab and COS6 by the anti-COS6 and theanti-COSM Abs in the cdELISAs were about 0.2, 2, and 2 ng/ml (ppb), respectively. In the cdELISA, the anti-NACOS6 Abwas found to recognize NACOS3-NACOS6, but not Nacetyl-D-glucosamine (GlcNAc), NACOS2, and COSs; theanti-COS6 Ab recognized COS2-COS6 and COSM, but notglucosamine (GlcN) and NACOSs. The recognition pattern ofthe anti-COSM Ab was almost the same as that of the anti-COS6 Ab, except that the former recognized COS2 andCOS3 slightly better than the latter.

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