Prokaryotic expression of glucans binding protein and application for detecting invasive fungal infection

Abstract

Objective To explore the potential application of glucans binding protein (GBP) in the detection of soluble glucans from serum sampled from patients with invasive fungal infection (IFI). Methods The prokaryotic plasmid pET30a-GBP252-668 coding for 252 to 668 amino acid in GBP as an active domain of GBP of Limulus polyphemus was constructed. The recombinant GBP252-668 was expressed and purified.Then a double GBP252-668 sandwich method was developed to detect (1,3)-β-D-glucans in the serum of patients with IFI. Meanwhile, the results were compared with G-test. Results Recombinant GBP252-668 (molecular weight is about 46 ku) was obtained and the double GBP252-668 sandwich method was established and optimized via chessboard method, the concentrations of GBP252-668 were 8 and 16 g/L for coating and detection respectively. The serum sampled from 48 patients with IFI and from 48 healthy donors were examined.and results were analysed. The sensitivity and specificity of double GBP252-668 sandwich method were 91.67% and 89.58% respectively and the sensitivity was non-inferior to G-test, and specificity of double GBP252-668 sandwich method was even higher than that of G-test (79.17%). Conclusions The GBP252-668 sandwich method is a potential detection method for IFI diagnosis. The method is non-inferior to G-test and its specificity is even higher than G-test. It is expected to be a new commercially available kit for IFI detection.

Keywords

• invasive fungal infection(ifi)|glucans binding protein|sandwich method|clinical application