Proteomics-based identification of DEAD-box protein 48 as a novel autoantigen, a prospective serum marker for pancreatic cancer
Section snippets
Materials and methods
Sera and cells. Sera were obtained at the time of diagnosis from 60 patients with pancreatic cancer, after informed consent was given. Sera from 60 healthy individuals and from 140 patients with other diseases (including 30 with colorectal cancer, 30 with gastric cancer, 30 with hepatocellular cancer, 30 with lung cancer, and 20 with chronic pancreatitis) were used as controls. Hep-2 cells were grown in RPMI1640 (Gibco) supplemented with 10% fetal bovine serum.
Sample preparation. Cells were
Reactivity of sera from pancreatic cancer patients with DDX48
Hep-2 cell proteins were separated by SDS–PAGE and then transferred onto nitrocellulose membranes. Sera from 55 patients with pancreatic cancer and from 52 healthy donors were screened individually for the presence of autoantibodies. The results revealed that 87.27% pancreatic patient sera contained autoantibodies recognizing Hep-2 proteins in the 20–170 kDa range, and most of them showed more than one positive bands, while only 8 out of 52 (15.38%) normal individuals observed positive
Discussion
Pancreatic cancer presents many challenges to clinicians and researchers searching for more effective ways to combat its often-devastating effects. Among the central challenges of this disease is the identification of markers for improved diagnosis.
In this study, we first employed 1-D immunoblot assay to screen the repertoire of autoantibodies in pancreatic cancer patients, and then implemented a proteomic approach which involves 2-D gel electrophoresis, immunoblot analysis, and mass
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