Journal of Advanced Biomedical Sciences

Objective: Klebsiella Pneumoniae is one of the most important infection bacteria of the Enterobacteriaceae family. It is also the second factor of hospital bacteremia due to the gram negative bacteria after Escherichia coli. In this research, this bacteria with the private 16SrDNA gene, was identified through PCR-ELISA technique.

Materials and Methods: In this method the dNTP labeled with Digoxigenin was used for amplifying the specific gene. Streptavidin was loaded in ELISA plate, and then the specific Biotinylated probe was used to connect the Polymerase Chain Reaction (PCR) product. Biotinylated probe was connected to the streptavidin and the amplified gene was attached to the probe. Finally, the digoxigenin antibodies were used to identify the PCR product. The reaction was measured with an ELISA reader.

Result: 16SrDNA of Klebsiella pneumoniae was amplified using the gene specific primers resulted in a fragment of the bp 260. The results of PCR-ELISA showed that this technique does not cross-react with the bacteria in their families. Additionally,  the sensitivity of 6.0 ng was evaluated.

Conclusion: In this research, PCR-ELISA technique was known as an accurate and rapid test for the detection of infection agents by using the specific gene. PCR-ELISA can act as an alternative method instead of time-consuming, less sensitive, and expensive techniques such as culture bacteria in blood agar plates, Macconkey agar, Realtime PCR and differential biochemical tests which are currently used in the research labs.