Identification of pathogens living in biofilms of chronic infections has been difficult with PCR, serological, biochemical and culture techniques. The study aims at the detection of bacterial pathogens in biofilms of biological secretions using SPR analysis Biacore. The antibodies were developed by isolating mononuclear lymphocytes from the blood of the patients who sustained systemic infection. The isolated lymphocytes had antibody secreting B cells (plasma cells) which were identified using flow cytometry analysis. The antibodies produced (n=4) were used to immobilize CM5 chip of Biacore to detect the bacteria in ulcer secretions with wound secretions of healthy volunteers as controls. The results from Surface Plasmon Resonance (SPR) analysis and culture technique were compared and statistically there was no significant difference obtained. The results from present study suggest that SPR analysis could be used as an alternative system for detection of bacteria in poly-microbial samples and detect the organisms that might not be discovered by culture or PCR method.