Enzyme Immunoassay for the Determination of H. pylori IgG Level in Human Serum
Introduction
Helicobacter pylori (H. pylori) is a gram-negative bacterium that colonizes the gastric mucosa. The organism is found in 95-98% of patients with duodenal ulcers and 60-90% of patients with gastric ulcers (Peterson, 1991). Various diagnostic tests, including bacteriological, histological, and serological methods, estimate that 90% of symptomatic patients are infected, while about 50% of adults over 50 years old are colonized without clinical symptoms (Podolsky, 1989). Studies have demonstrated that eradication of the organism through antimicrobial therapy correlates with symptom resolution and disease cure (McGuigan, 1988).
Clinical Significance
H. pylori infection is associated with peptic ulcers, chronic gastritis, and an increased risk of gastric cancer. Accurate diagnosis is essential for guiding appropriate treatment. Serological tests, such as the H. pylori IgG ELISA, offer a non-invasive and effective method for detecting past and ongoing infections. IgG antibodies against H. pylori remain detectable for months or even years, making ELISA a useful screening tool for epidemiological studies and diagnostic purposes (Perez & Blaser, 1991).
Diagnostic Methods
Patients with gastrointestinal symptoms suggestive of H. pylori infection can be diagnosed using two primary methods:
- Invasive Techniques: These include biopsy followed by culture, histological examination, or direct urease activity detection.
- Non-Invasive Techniques: These include urea breath tests and serological methods such as ELISA.
Biopsy-based tests are subject to errors due to sampling issues and bacterial contamination. H. pylori infection triggers a humoral immune response, producing specific antibodies (IgG, IgM, and IgA). The H. pylori IgG ELISA test is a highly accurate and convenient method for determining bacterial colonization and is considered the technique of choice for IgG detection.
Test Principle
The H. pylori IgG ELISA Kit is based on an indirect ELISA technique. Microtiter wells are coated with a specific quantity of H. pylori antigens. A diluted serum sample is then incubated in the wells. If H. pylori-specific antibodies are present, they will bind to the antigens. After an incubation period at room temperature, unbound antibodies are removed through washing. The wells are then treated with an anti-human IgG horseradish peroxidase (HRP)-conjugated secondary antibody. After a second incubation and wash step, a chromogen-substrate solution is added, leading to the development of a blue color. The reaction is stopped with an acidic stop solution, changing the color to yellow, which is then measured spectrophotometrically at 450 nm. The intensity of the color is directly proportional to the concentration of anti-H. pylori IgG in the sample.
References
- McGuigan, J. E. (1988). Peptic ulcer and gastritis. Harrison’s Principles of Internal Medicine, 12th ed., 1229-1248.
- Peterson, W. L. (1991). H. pylori and peptic ulcer disease. New England Journal of Medicine, 324, 1024-1047.
- Podolsky, I. (1989). Prevalence of H. pylori in healthy subjects and patients with peptic disease. Gastroenterology, 96 (Suppl.), A394.
- Perez, C. I., & Blaser, M. O. (1991). Serodiagnosis of H. pylori: Comparison of enzyme-linked immunosorbent assays. Journal of Clinical Microbiology, 29, 1635-1639.