C-220. Validation of a Serum Histoplasma Antigen Assay

J. L. Cloud1, S. K. Bauman2, J. M. Pelfrey2, K. Ludwig1, E. R. Ashwood1,3;
1ARUP Inst. for Clin. and Experimental Pathology, Salt Lake City, UT, 2Immuno-Mycologics, Inc., Norman, OK, 3Univ. of Utah, Salt Lake City, UT.

A validated polyclonal sandwich enzyme immunoassay (EIA) for the detection and quantitation of histoplasma antigens in human urine has successfully been in use at ARUP Laboratories. The performance evaluation of the assay resulted in good precision, both within-run and between-run. The assay is linear when testing dilutions of the highest calibrator, but non-linear when testing dilutions of a positive human serum sample. The assay, as performed on urine samples, is invalid with untreated human sera. Serum pre-treatment with pronase has been shown to work well in reducing interference from proteins and blocking epitopes by immune complexes. Pronase-treating urine samples, however, results in lower level of detection and cannot be used. To validate pronase-treated serum samples in the Histoplasma Antigen EIA, we tested 71 samples that were previously tested by MiraVista Diagnostics (MVD) to determine percent agreement in positive and negative samples. Thirty-seven of the samples were compared with the MVD second generation assay while 34 samples were compared with the current MVD third generation assay. The second generation assay comparison resulted in an R2 of 0.5957 while the improved third generation comparison resulted in an R2 of 0.7514. After excluding samples testing in the equivocal range, 24 of 28 (86%) MVD positive samples were in agreement and 28 of 28 (100%) MVD negative samples were in agreement. We conclude the serum histoplasma antigen test performs well and is valid in our hands. Our studies further show that antigenic differences between urine and serum sample matrices are present.