Enzyme-linked immunosorbent assay for group A Streptococcal anti-DNase B in human sera, using recombinant proteins - Comparison to the DNA methyl green micromethod

• Published in: Journal of immunological methods Vol. 451; pp. 111 - 117
• Main Authors: Das, Sarita, Dileepan, T., Johnson, D.R., Kaplan, E.L., Patrick Cleary, P.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.12.2017
• Subjects: antibodies; Antibodies, Bacterial - blood; antigens; Antigens, Bacterial - genetics; Antigens, Bacterial - immunology; Biomarkers - blood; Coloring Agents - chemistry; deoxyribonucleases; Deoxyribonucleases - genetics; Deoxyribonucleases - immunology; DNA; DNA methyl green micromethod; dnaseB; ELISA; enzyme-linked immunosorbent assay; Enzyme-Linked Immunosorbent Assay - methods; genes; Group A Streptococcus; His-tagged DNase B; Humans; Methyl Green - chemistry; nationalities and ethnic groups; neutralization; Polymerase Chain Reaction; Predictive Value of Tests; recombinant proteins; Recombinant Proteins - immunology; Reproducibility of Results; Streptococcal Infections - blood; Streptococcal Infections - diagnosis; Streptococcal Infections - genetics; Streptococcal Infections - immunology; Streptococcus pyogenes; Streptococcus pyogenes - enzymology; Streptococcus pyogenes - genetics; Streptococcus pyogenes - immunology; streptolysin; Time Factors; His-tagged DNase B; SCPA; dnaseB; Group A Streptococcus; Streptococcus pyogenes; DNA methyl green micromethod; ASO; PCR; DMGM; ADB; ELISA
• ISSN: 0022-1759; 1872-7905; 1872-7905
• DOI: 10.1016/j.jim.2017.09.006

Among the four known Streptococcal nucleases comprising of DNase A, B, C and D; DNase B is the most common, and determination of the levels of antibody to DNase B (ADB) is often used to confirm a clinical diagnosis of Streptococcus pyogenes/group A Streptococcal (GAS) infection. The commonly used assays for antibodies that neutralize DNase B or streptolysin O activity use partially purified antigens that often fail to detect antibody changes subsequent to culture documented infections. Therefore, an enzyme-linked immunosorbent assay (ELISA) was developed employing his-tagged recombinant DNase B as plate antigen for comparison to the commonly used DNA methyl green micromethod (DMGM). DNAs from various Streptococcal species were screened for presence of dnaseB gene by PCR. Measurements of ADB in sera collected from subjects belonging to different ages, and ethnic groups were used to compare the two methods. dnaseB was not detected by PCR in DNA samples isolated from different strains of group B (GBS), C (GCS) and G (GGS) Streptococci. The ADB based ELISA proved to be highly sensitive and more responsive to changes in antibody concentration than DMGM. Use of recombinant DNase B eliminates the variability associated with the enzyme, partially purified from Streptococcal culture supernatants from various commercial sources and may provide a more reliable source of antigen to a wider group of laboratories concerned with GAS diagnosis. •dnaseB was detected by PCR in DNA samples of group A Streptococci.•An ELISA is developed to aid clinical diagnosis of the patients suffering from group A Streptococcal (GAS) infection.•It is based on a Histidine-tagged recombinant DNase B derived from Streptococcus pyogenes.•This assay has been found to be more sensitive to determine the levels of antibody to DNase B (ADB) than the ADB titer method.