Detection of H. pylori antibody profile in serum by protein array.
OBJECTIVE
To detect multiple H. pylori antibodies in serum samples of individuals who carry H. pylori by protein array.


METHODS
Recombinant H. pylori antigens, urease B subunit (UreB), vacuolating toxin A (VacA) and cytotoxin associated gene A protein (CagA), were prepared and immobilized in matrixes on nitrocellulose membrane by robotics to bind the specific immunoglobulin G (IgG) antibodies in serum. Staphylococcus protein A (SPA) labeled by colloid gold was used to integrate the immuno-complex and gave red color signal. The scanner based on charge-coupled device (CCD) could collect the image signal and convert it into digital signal.


RESULTS
When human IgG was printed on the membrane in increasing concentrations and incubated with immunogold, a linear dose response curve was obtained and the detection limit for IgG was about 0.025 ng. The cutoff values, which were defined as the mean grey level plus 3 times of standard deviation, were 27.183, 28.546 and 27.402, for anti-UreB IgG, anti-CagA IgG and anti-VacA IgG, respectively, as 400 human serum samples with negative H. pylori antibodies were detected by the protein array. When 180 serum samples from patients in hospital were employed for detection of IgG against UreB, CagA and VacA, the sensitivity of the protein array was 93.4%, 95.4%, 96.0%, and the specificity was 94.8%, 94.4% and 97.5%, respectively, as compared with the results obtained by ELISA. The assay also showed high reproducibility, uniformity and stability, and the results were available within 30 min.


CONCLUSIONS
The protein array is a very practical method for rapid detection of multiple antibodies in serum samples. It is especially useful for large scale epidemiological investigation of the infection of H. pylori.
Han
Feng-Chan
FC
Institute of Genetic Diagnosis, The Fourth Military Medical University, 17 West Changle Road, Xi'an 710032, Shaanxi Province, China. fengchan@fmmu.edu.cn
Li
Xu-Jun
XJ
Jiang
Hong
H
Qin
Li-Peng
LP
Li
Ding
D
Guo
Yan-Hai
YH
Liu
Zhi-Guang
ZG
Zhang
Li
L
Yan
Xiao-Jun
XJ
D000907:Antibodies, Bacterial / Q000097:blood*; D000942:Antigens, Bacterial; D016480:Helicobacter pylori / Q000276:immunology*; D006801:Humans; D040081:Protein Array Analysis / Q000191:economics / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D044967:Serum / Q000737:chemistry; D013272:Stomach Diseases / Q000175:diagnosis
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial