Characteristics of 26 kDa antigen of H. Pylori by Monoclonal Antibody.

Iran J Allergy Asthma Immunol 2015 Feb;14(1):113-9

Department of Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.

Alkylhydroperoxide reductase (AhpC, the 26 kDa antigen) is one of the abundant antioxidant enzymes in Helicobacter pylori and seems to have a good potential for use in development of immunoassays to detect H. pylori infection in clinical specimens. This study aimed to investigate some properties of this antigen by the produced monoclonal antibodies. Five established hybridoma cell lines secreting monoclonal antibodies (MAbs) against 26 kDa antigen of H. pylori were cultivated and MAbs were purified by affinity chromatography. Subsequently, MAbs were conjugated with biotin, and different combinations of capture and tracer antibodies used in sandwich ELISA. Immunoblotting of bacterial extracts were performed to estimate aggregation status of the antigen. Release of antigen from the cultivated bacteria on solid media was examined by sandwich ELISA, and also, existence of interference in fecal extract was investigated by immunoblotting and sandwich ELISA. Our findings showed that the MAbs against 26 kDa antigen of H. pylori could recognize three bands of nearly 25 kDa, 50 kDa, and 75 kDa in immunoblotting. This study also indicated presence of more antigens in the culture medium around the bacteria than the bacterial extract itself. The results of sandwich ELISA and immunoblotting on fecal extracts suggest the presence of interfering agents that prevent detection of antigen by antibody in ELISA but not in immunoblotting. In this study the oligomerization of the 26 kDa antigen, presence of interfering agents in stool matrix, and release of antigen to outside of bacteria, were demonstrated.

Download full-text PDF

Source
February 2015
6 Reads
1 Citation
1.013 Impact Factor

Publication Analysis

Top Keywords

kda antigen
20
sandwich elisa
16
antigen pylori
12
elisa immunoblotting
12
antigen
10
interfering agents
8
mabs kda
8
kda kda
8
presence interfering
8
immunoblotting study
8
monoclonal antibodies
8
release antigen
8
kda
7
pylori
5
immunoblotting
5
elisa
5
antigen cultivated
4
existence interference
4
interference fecal
4
antigen release
4

Similar Publications