History of (103)-b-D-glucans testing

Lehmann and Reiss were the first to report the measurement of circulatingAspergillus fumigatusanti-gen in rabbits and humans with IA in 1978 [16].

Theirstrategy involved the development of an antiserum against serum from rabbits experimentally infected with A. fumigatus. They detected a single antigenic moiety that circulated in the blood of infected rabbits and humans with proven IA [16]. They later determined that galactomannan was the molecule being detected by the rabbit antiserum [17].

During these experiments, Betaglucans was isolated and characterized fromA. fumigatus cell walls, but found to be non-antigenic [17]. In 1968, Levin and Bang developed an assay for bacterial endotoxin using the amebocytes ofLimulus polyphemus[the American horseshoe crab] [18]. During pyrogenicity testing of carboxy-methylated Betaglucans that was being studied as an anti-tumor agent, Kakinuma and colleagues noted that Betaglucans consistently turned theLimu-lus test positive , despite confirmation of non-pyrogeni-city in inoculated animals [19].

Morita and colleagues, by studying Tachypleus tridentatus [Japanese horseshoe crab] amebocyte lysate fractions, demonstrated that Betaglucans triggered theLimulustest coagulation cascade via a separate proenzyme, which was termed Factor G (Fig. 1) [20].

Obayashi and colleagues developed a chromo-genic test based on the recombination of the different amebocyte lysate fractions and proposed that use of recombined fractions containing Factor G, but not Factor C, which recognizes endotoxin, could be used for non-invasive testing for the diagnosis of invasive fungal diseases (IFD) [21].

Following proof of principle studies [22] and after confirmation that Betaglucans was indeed the substrate that bound to Factor G to trigger the reaction cascade of their chromogenic test [23], Obayashi and colleagues published the first multicenter validation study in 1995 [11].

A similar Betaglucans test was developed using amebocyte lysate fractions of the American horseshoe crab and approved by the FDA in 2004 as an aid in the diagnosis of fungemia and deep-seated mycoses [24] following a prospective validation study [12]