In order to prepare an ELISA plate which can be used for site-specific conjugation of oligonucleotide probe, partially hydrolysed nylon was used to coat ELISA plate, oligonucleotide probes were conjugated to ELISA plate by covalent conjugation of 5′ phosphate group of the probe with the amino groups of the hydrolysed nylon under the catalysis of water soluble carbodiimide. The method was used to detect PCR product of Mycoplasma pneumoniae, and reproducible result was acquired. The reported method can be used in molecular detection of clinical pathogens.