The Helitope®-RUO Kit is an indirect ELISA test for the detection of IgG antibodies to CagA peptide epitopes in human serum. The test takes less than 2 operating hours. Diluted human serum samples are added to wells pre-coated with: 1) Selected CagA-derived peptides originating from across multiple strains of H. pylori, and for which cross-reactivity to other antigens is minimal; or 2) Proprietary peptide-based control that lacks CagA-derived antigenic sequences, to assess sample-specific background binding to the assay surface. The samples are incubated for 30 minutes. After incubation and washing, human antibodies targeting the CagA peptide antigens remain bound to the plate surface. Secondary antibody conjugated to horseradish peroxidase (HRP) targeting human IgG is then added to each well and the plate is incubated for 45 minutes. The ELISA wells are washed once again before a tetramethylbenzidine (TMB) substrate is added. An acidic stopping solution is finally used to stop the reaction and the rate of enzymatic turnover of the substrate is determined by measuring absorbance at 450 nanometers. A calibrator (Mouse-human chimeric IgG antibody raised against CagA peptide) is included to validate the integrity of the test and to establish the assay-specific cut-off (protocol 1).

Protocol 1:
Protocol 1 is non quantitative and determines if a serum sample is CagA positive or negative using a cut off formula. Up to 22 samples may be evaluated in this protocol. In addition to the samples and the calibrator, negative control (sample diluent only) is included in each assay run. Each sample is run in duplicate on (1) antigen-coated wells, and on (2) Control wells.

Protocol 2:
Protocol 2 is a semi-quantitative protocol. A standard curve is created by diluting out the calibrator. The relative levels of CagA antibodies in the serum can be estimated using this standard curve and are expressed in arbitrary units (AU). Up to 16 samples may be evaluated using protocol 2. In addition to the samples and the standard curve, a negative control (sample diluent only) is included in each assay run. Each sample is run in duplicate on (1) antigen-coated wells, and (2) on Control wells.

Helicobacter pylori is a gastric bacterium associated with chronic infection and an increased risk of gastric disease, including gastric cancer. The cytotoxin-associated gene A (CagA) protein is a major virulence factor expressed by a subset of H. pylori strains and is a common target of host antibody responses. (Parsonnet et al., 1997; Hatakeyama, 2004). Only a subset of H. pylori strains within any population carries the cag pathogenicity island and expresses CagA. Consequently, not all H. pylori-infected individuals develop detectable anti-CagA antibody responses. Studies have indicated that IgG anti-CagA-antibodies can remain preferentially detectable in the blood even after the H. pylori infection has been treated or the bacteria has been cleared from the stomach, serving as a marker of past CagA+ H. pylori infection (Sörberg et al., 1997). Current serology tests for CagA+ H. pylori antibodies are limited in clinical usefulness due to antigenic heterogeneity and cross-reactivity, which reduces their specificity. The Helitope®-RUO Kit has been developed using linear CagA epitopes (peptides) with high antibody reactivity in CagA+ H. pylori-infected cases, and low levels in non-infected individuals. Compared to whole antigen, these peptides exhibit lower cross-reactivity in non-infected individuals or in individuals infected with H. pylori that do not produce CagA (“CagA-”), creating the basis for a more accurate and specific serology test.

anti-CagA antibodies (IgG) in Human serum

0.15 AU/mL -10AU/mL (semiquantitative)

96%

>95%

Human

ELISA

Colorimetric

Human Serum

For research use only.

Australia

6 months from purchase.

Store at 2-8°C

Shipped on ice packs