Immunoreactivity of cryptococcal antigen is not stable under prolonged incubations in human serum

Abstract

The stability of cryptococcal antigen from Cryptococcus neoformans serotype A and D strains at different temperatures in serum and other solvents was studied. Samples stored at -20 or 4 degrees C had equivalent stabilities as measured by the Premier Cryptococcal Antigen kit and the Cryptococcal Antigen Latex Agglutination System (CALAS) kit. However, using the Premier Cryptococcal Antigen kit, there was a 91% loss of reactivity in samples incubated in human serum for 4 weeks at 37 degrees C. A loss of reactivity of more than 99% was observed after incubation at 45 degrees C for 4 weeks. The capsular antigen was not detected by the Premier Cryptococcal Antigen kit after 16 weeks at elevated temperatures. Antigen titers were also reduced in a latex agglutination assay (CALAS) after 4 weeks at 37 and 45 degrees C. The loss of antigen reactivity was a function of pH and temperature.

FIG. 1.

The immunoreactivity of GXM in different solvents decreased at elevated temperatures. GXM from strain H99 was measured with the Premier Cryptococcal Antigen kit after 4 weeks (A) and 16 weeks (B) of incubation at −20, 4, 37, and 45°C. The samples in the different solvents (four bars for each solvent) were incubated at −20, 4, 37, and 45°C (shown left to right for each solvent). The absorbance of the −20°C samples was set at 100, and the remaining samples were normalized against this value.

FIG. 2.

The loss of O acetylation on GXM incubated in human serum is detected by EIA. GXM from strain H99 that was incubated for 4 weeks in human serum was measured in EIAs using MAbs that can distinguish the acetylation state of the polysaccharide. (A) EIA using MAb 2D10 as the capture MAb and MAb 18B7 as the detecting MAb. (B) EIA using MAb 21D2 as the capture MAb and MAb 18B7 as the detecting MAb.