The purified NusA-IRAS (10-120aa) protein (50 μg in a volume of 80 μL) was mixed with an equal volume of Freund's complete adjuvant. The antigen-adjuvant mixture was injected into 6 female BALB/c mice and was followed by three booster injections at 2-week interval in incomplete Freund's adjuvant. The mouse with the highest antibody titer tested by ELISA was boosted intraperitoneally with 100 μg NusA-IRAS (10-120aa) protein without adjuvant 3 days before the cell fusion. Feeder layer cells were prepared 1 day prior to fusion. Splenocytes from mice with the highest ELISA antibody titers were fused with murine myeloma cells SP2/0 following standard procedures [25] . Culture supernatants were collected after fusion and initially screened by ELISA with purified NusA-IRAS (10-120aa) fusion proteins as antigens. Positive hybridoma clones were selected with the limiting dilution method, and stable hybridoma clones were obtained after 3 cloning cycles. Isotypes of antibodies were identified with a mouse subisotype panel (Bio-Rad). The pristine-primed BALB/c mice were injected intraperitoneally with 1 × 10 6 hybridoma cells per mouse in order to acquire abundant mAbs. The ascitic fluid was collected and mAbs were purified with a protein A/G affinity column (Amersham Pharmacia Biotech).