Design of enzyme-linked immunosorbent assay method for detection of anti-streptolysin-O antibodies on base of recombinant streptolysin-O

  Introduction: Group A, β hemolytic streptococci are among the major causative agents of otorhinolaryngology infections. Inadequate treatment of disease may lead to serious disorders such as acute rheumatic fever and glomerulonephritis. Anti-streptolysin-O (ASO) is commonly used as a marker in the diagnosis of infection. Purification of native streptolysin-O has several difficulties and its industrial production process is time consuming with very low yield and the risk of biological contamination. In this study, we used a recombinant streptolysin-O protein as an antigen to detect ASO antibodies in enzymes-linked immunosorbent assay (ELISA).   Materials and Methods: We amplified streptolysin-O gene by polymerase chain reaction (PCR) method and subcloned in prokaryotic expression vector PET28a. E.coli. BL21-DE3-plySs strain was transformed with PET28a-streptolysin-O and gene expression was induced by IPTG. ELISA microplates were coated with different concentration of streptolysin-O protein. Level of ASO antibodies were detected by ELISA method. The results obtained from ELISA method were compared with inhibition of hemolysis assay as a standard method.   Results: The results showed that there is a positive significant correlation between the ELISA and inhibition of hemolysis method(r=0.97, p=0.0001). The sensitivity and specificity of ELISA for detection of ASO anti bodies were 100% and 83%, respectively.   Conclusion: ELISA developed with recombinant streptolysine O showed a good sensitivity for detection of ASO antibodies. It is suggested that this method could be suitable for immunoassays.