Reference materials were produced to standardize the immunoglobulin class specificity and potency of immunofluorescent anti-IgM conjugates used for diagnostic tests for congenital syphilis. In attempting to mimic essential immunologic characteristics of syphilitic and nonsyphilitic infant sera, we evaluated these sera in comparison with processed adult sera. We were quite surprised to discover that some syphilitic babies do not produce significant quantities to IgM antibody to T. pallidum in response to their infection, as would be expected; instead, they make relatively large amounts of IgM anti-IgG. We found this to be true also for newborns and infants infected with cytomegalovirus, rubella, and toxoplasmosis. To our knowledge, this observation has not been previously reported. However, it could have been predicted from the knowledge that older infants and young children normally produce IgM antibodies to maternal IgG allotypes (Gm factors). We are disturbed that these findings suggest that currently recommended indirect immunofluorescence IgM tests for perinatal infection may not be disease specific. Our observations may be important for a better understanding of basic immunologic mechanisms of fetal-maternal to tolerance and fetal response to life-threatening infection.

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The fluorescent treponemal antibody-absorption double-staining step-by-step procedure and proposed reference reagents for the test are described. The test and reagents were evaluated in two separate laboratories on 265 fresh sera, and test results were compared with the reference fluorescent treponemal antibody-absorption test results performed in a third laboratory. The data indicate that the tests are comparable in the areas where the test is recommended for use. Problems with inadequate light filtration occurred, but these could be resolved. This test is recommended for use with microscopes equipped with incident illumination.

Immunofluorescent staining of Treponema pallidum was studied to clarify the effect of three factors on the results of the fluorescent treponemal antibody-absorption test: (i) heat inactivation of sera at 56 degrees C for 30 min before testing, (ii) use of multicircle slides, and (iii) tungsten illumination to visualize and assess unstained treponemes on reactive as well as nonreactive smears. It was found that serum inactivation before testing was not necessary for detection of immunoglobin G antibody, but an immunoglobulin M prozone was detected in unheated serum. On multicircle slides, it was demonstrated that a false-positive reaction could be obtained in 30 s at 37 and 25 degrees C if a smear where a nonreactive serum had been placed was crossed by a strongly reactive serum from another circle. Tungsten illumination proved necessary for correct assessment of unstained treponemes on all fluorescent treponemal antibody-aborption test smears, reactive or nonreactive. The possible role of these factors in incorrect fluorescent treponemal antibody-absorption test results is discussed.