Title : Rickettsial disease serodiagnosis – Current trends
Abstract:
Background: Clinical Rickettsial diseases are found worldwide, yet these agents are not amenable to in vitro propagation outside of proper BSL2 facilities. Diagnosis has utilized both PCR and antibody detection, with serology products currently available for automated (ELISA), manual (IFA and MIF), and rapid test formats for both IgG-specific and IgM-specific antibodies.
Methods: Our current ELISA assays are made using specific and highly purified antigens with high specificity and reproducibility. For Spotted Fever Group (SFG) we utilize the SFG lipopolysaccharide antigen for IgG-specific assays and a recombinant β-peptide antigen for IgM-specific assays. Although IFA and MIF assays require a fluorescence microscope to visualize Rickettsia, the IFA IgG titers are specifically required by CDC for our national epidemiology. Although the specific methodology is not described, the acute phase diagnostic use of IFA and MIF assays for IgG requires a serum pre-treatment step utilizing goat anti-human IgM antiserum to diminish competition for antigen binding by the IgM peak response. The same serum pretreatment is required for IgM-specific testing by IFA and MIF, except that the serum diluent used contains goat anti-human IgG antiserum.
Results: Both IFA and MIF assays are enhanced by pre-treating the acute phase sera with diluted goat anti-human IgG for IgM-specific assays and goat anti-human-IgM for IgG-specific assays. The difference observed is a several-fold enhancement that depends upon the timing of the acute phase serum, generally >/= 8-fold from <1:64 to at least 1:256. The initial appearance of IgG titers can be seen earlier in the time course of the acute response and the IgM is seen for a longer time, as well.
Conclusion: When IFA is the only acceptable assay, the Serum Pre-Treatment protocol enhances both IgG and IgM titers, quite often turning otherwise negative results into positive titers. Otherwise, the appropriate ELISA assays are more accurate and sensitive.
