Nowadays, chemiluminescent immunoassay (CLIA) has gained increasing attention in different fields including clinical diagnosis and environmental monitoring because of its high sensitivity and specificity simple equipment and wide linear range. In this study, the colloidal CuO nanoparticles and CuO/MWCNT nanocomposite as the efficient nanocatalysts were introduced and compared for amplification of the luminol CLIA system. In the present work, we applied the antibody-nanomaterials conjugate without horseradish peroxidase (HRP) enzyme as the CLIA reaction catalyst. The CuO nanoparticles-antibody con- jugates and CuO/MWCNT nanocomposite-antibody conjugates were prepared using the covalent binding The results show that CuO nanoparticles and CuO/MWCNT nanocomposite enhance the luminol chemi- luminescence intensity, significantly. Under the optimal conditions, the calibration plot for the standard sample of the hepatitis B surface antigen and chemiluminescence intensity was approximately linear within the range of 3.5 × 10−9–2.5 × 10−6 and 2.2 × 10−9–5.0 × 10−6 g ml −1. The limit of detection for the assay using CuO nanoparticles and CuO/MWCNT nanocomposite were 1.8 and 0.85 ng ml−1, respectively which these results were comparable to that achieved of the enzyme-linked immunosorbent assay. The present CLIA method based on the new antibody-nanomaterials conjugates and chemiluminescence system has introduced for first time and offered great promise for simple and sensitive analysis of biologica samples.