Based on the prepared group-specific antibodies against polychlorinated biphenyls (PCBs), a direct competitive real-time fluorescent quantitative immuno-polymerase chain reaction (rt-IPCR) assay was developed. The purpose of the assay was the determination of multi-residue PCBs in indoor air samples. In the assay, male New Zealand white rabbits were immunized with an immune antigen mixture composed of PCB12-O-BSA, PCB37-O-BSA, and PCB77-O-BSA. The specific polyclonal antibodies (pAbs) to multi-residue PCBs were obtained and used to develop a direct competitive rt-IPCR assay. The specificities of the pAbs were examined by the indirect competitive enzyme-linked immunosorbent assay (id-ELISA). The assays were found to be highly specific for PCB congeners as well as Aroclors 1248 and 1242. The effect of optimal reagent concentrations on reducing background fluorescence was also investigated. Using the optimized assay, a standard curve for Aroclor 1248 was prepared. The linear range for the determination of PCBs was 10 to 10⁶ fg mL⁻¹ with a correlation coefficient of 0.98 and a detection limit of 10.25 fg mL⁻¹. The entire procedure was then evaluated using spiked air samples. The rt-IPCR results for the air samples were confirmed by gas chromatography/mass spectrometry and ELISA. Recovery was lower or higher with agitation but would still be acceptable for use in an on-site field test to provide rapid, semiquantitative, and reliable test results for detection of PCBs in air samples.