The ELISA were performed on
polystyrene microtiter plates (Nunc, MaxiSorp) coated with LPS (2a
antigen) at the final concentration of 10 microg/ml. The
antigen was extracted from Klebsiella rhinoscleromatis Rh32 by the
trichloroacetic acid and separated by
ethanol (Boivin method). The
antibodies against the LPS were detected by ELISA in serum samples collected from 65 patients suspected in clinical investigation for
rhinoscleroma in Poland from 1970 to 2009. Additionally, the specificity of the
antigen was tested using serum sample of immunized rabbit and 30 sera of patients from control group, with high level of
antibodies to different bacterial pathogens. All serum samples were diluted 1:100. The concentrations of
IgA,
IgG and
IgM antibodies were expressed as optical density (OD) measured at the wavelength of 450 nm. The cut-off limit of serum
antibodies was set at mean antibody OD determined in the sera of 30 blood donors exceeded by three standard deviations. The presence of
IgA and
IgG antibodies were detected by ELISA in 33 (50,8%) and
IgM in 28 (43,1%) of patients. Most of the serum samples (75%) with high level of specific
antibodies were obtained from patients before 1980. On the other hand
antibodies to K. rhinoscleromatis were detected only in 2 (6,7%) patients from the control group and none of blood donors. In conclusion, our home-made ELISA, based on purified LPS of K. rhinoscleromatis showed high specificity and sensitivity in the diagnosis of
antibodies to K. rhinoscleromatis in comparison to the
complement fixation test. The presence of high level of specific
IgA,
IgG and
IgM antibodies in the sera obtained in different stages of disease may showed that during the
rhinoscleroma is permanent stimulation of antibody production.