Highly sensitive immunoassay based on E. coli with autodisplayed Z-domain.
The Z-domain of protein A has been known to bind specifically to the F(c) region of antibodies (IgGs). In this work, the Z-domain of protein A was expressed on the outer membrane of Escherichia coli by using "Autodisplay" technology as a fusion protein of autotransport domain. The E. coli with autodisplayed Z-domain was applied to the sandwich-type immunoassay as a solid-support of detection-antibodies against a target analyte. For the feasibility demonstration of the E. coli based immunoassay, C-reactive protein (CRP) assay was carried out by using E. coli with autodisplayed Z-domain. The limit of detection (LOD) and binding capacity of the E. coli based immunoassay were estimated to be far more sensitive than the conventional ELISA. Such a far higher sensitivity of E. coli based immunoassay than conventional ELISA was explained by the orientation control of immobilized antibodies and the mobility of E. coli in assay matrix. From the test results of 45 rheumatoid arthritis (RA) patients' serum and 15 healthy samples, a cut-off value was established to have optimal sensitivity and selectivity values for RA. The CRP test result of each individual sample was compared with ELISA which is the reference method for RA diagnosis. From this work, the E. coli with Z-domain was proved to be feasible for the medical diagnosis based on sandwich-type immunoassay.
Jose
Joachim
J
Institute of Pharmaceutical Chemistry, Heinrich Heine University, Duesseldorf, Germany.
Park
Min
M
Pyun
Jae-Chul
JC
D001172:Arthritis, Rheumatoid / Q000175:diagnosis / Q000378:metabolism; D002097:C-Reactive Protein / Q000032:analysis*; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D057230:Limit of Detection; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013205:Staphylococcal Protein A / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; D002097:C-Reactive Protein