Western blotting is useful in the salivary diagnosis of Helicobacter pylori infection

Abstract

Background: The salivary diagnosis of Helicobacter pylori infection offers attractive possibilities for the epidemiological study of infection in children. Salivary enzyme linked immunosorbent assay (ELISA) is less reliable then serum ELISA, owing to variable transudation of immunoglobulin. In addition, children are more difficult to study because of lower specific serum antibody concentrations to H pylori. The performance of salivary western blotting in comparison with serum western blotting and serum ELISA was investigated in school children.

Subjects and methods: Paired serum and saliva specimens were obtained from 669 [corrected] school children aged 9-11 in 10 British towns. All saliva and serum specimens were first analysed by ELISA; subsequently, western blotting of both specimens was performed on 31 and 34 specimens, respectively, to establish the criteria for positivity for western blotting. The remaining 121 specimens were then tested blindly and saliva was compared with the serum.

Results: The sensitivity and specificity of salivary ELISA in the 669 [corrected] specimens was 32 of 50 (64%) and 530 of 619 (86%) [corrected], respectively, when compared with serum ELISA. The western blotting validation was performed on 28 subjects with positive serum and positive salivary ELISA, 28 saliva positives with negative serum, 16 saliva negatives with positive serum, and 50 doubly negative subjects. Compared with serum western blots, the sensitivity and specificity of salivary western blots was 38 of 47 (81%) and 68 of 75 (91%), respectively. Using serum ELISA as the gold standard, the sensitivity and specificity were 32 of 44 (73%) and 72 of 78 (92%), respectively, the specificity being significantly higher than salivary ELISA (p < 0.001).

Conclusion: Salivary western blotting for IgG is useful in the diagnosis of H pylori infection and is superior to ELISA. It also permits the identification of pathogenic strains.

Figure 1 Frequency distribution of optical densities in 665 subjects who had salivary enzyme linked immunoabsorbent assay (ELISA) performed and who had matching serum specimens. A cut off point of 0.41 optical density units was used.

Figure 2 Frequency distribution of serum enzyme linked immunoabsorbent assay (ELISA) values (n = 129).

Figure 3 Sensitivity and specificity (%) of saliva enzyme linked immunoabsorbent assay (ELISA) against serum ELISA by cut off point in 665 subjects with samples for both.

Figure 4 Samples used to establish criteria for a positive and negative western blot for serum and saliva. Serum western blots: lanes 1 and 2, serum control CagA VacA positive strains; lanes 3–9, serum enzyme linked immunoabsorbent assay (ELISA) positive samples; lanes 10–19, serum ELISA negative samples. Saliva western blots: lanes 20 and 21, serum ELISA positive samples; lanes 22 and 23, serum ELISA negative samples; lanes 24 and 25, serum ELISA positive samples. Band sizes are in kDa.

Figure 5 Further representative saliva samples used to establish positive and negative criteria. Lane 1, positive serum control; lanes 2–4, serum enzyme linked immunoabsorbent assay (ELISA) negative specimens; lanes 5–11, serum ELISA positive specimens; lanes 12–25, serum ELISA negative specimens. Band sizes are in kDa.