Annals of Oncology Advance Access published October 21, 2011

original article Annals of Oncology

doi:10.1093/annonc/mdr384

Helicobacter pylori infection assessed by ELISA and by

immunoblot and noncardia gastric cancer risk in
a prospective study: the Eurgast-EPIC project
C. A. Gonzalez1*, F. Megraud2, A. Buissonniere2, L. Lujan Barroso1, A. Agudo1, E. J. Duell1,
M. C. Boutron-Ruault3,4, F. Clavel-Chapelon3,4, D. Palli5, V. Krogh6, A. Mattiello7, R. Tumino8,
C. Sacerdote9, J. R. Quiros10, E. Sanchez-Cantalejo11, C. Navarro12, A. Barricarte13,
M. Dorronsoro14, K.-T. Khaw15, N. Wareham15, N. E. Allen16, K. K. Tsilidis16, H. Bas Bueno-de-
Mesquita17,18, S. M. Jeurnink17,18, M. E. Numans19, P. H. M. Peeters19, P. Lagiou20,
E. Valanou21, A. Trichopoulou21, R. Kaaks22, A. Lukanova-McGregor22, M. M. Bergman23,

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H. Boeing23, J. Manjer24, B. Lindkvist25, R. Stenling26, G. Hallmans27, L. M. Mortensen28,
K. Overvad28, A. Olsen29, A. Tjonneland29, K. Bakken30, V. Dumeaux30, E. Lund30, M. Jenab31,
I. Romieu31, D. Michaud32, T. Mouw32, F. Carneiro33, C. Fenge34& E. Riboli35
1
Unit of Nutrition, Environment and Cancer, Cancer Epidemiology Research Program, Bellvitge Biomedical Research Institute (IDIBELL), Catalan Institute of Oncology,
Barcelona, Spain; 2INSERM U853, Bordeaux; 3Centre for Research in Epidemiology and Population Health, Institut Gustave Roussy, Villejuif; 4Paris South University,
Villejuif, France; 5Molecular and Nutritional Epidemiology Unit, Cancer Research and Prevention Institute (ISPO), Florence; 6Department of Preventive & Predictive
Medicine, Nutritional Epidemiology Unit, Fondazione IRCCS Istituto Nazionale dei TumoriMilan; 7Department Of Clinical And Experimental Medicine, Federico Ii
University, Naples; 8Cancer Registry and Histopathology Unit, Civile M.P. Arezzo Hospital, Ragusa; 9Centre for Cancer Epidemiology and Prevention (CPO

original
Piemonte), Turin, Italy; 10Public Health and Participation Directorate, Health and Health Care Services Council, Asturias; 11Andalusian School of Public Health,

article
CIBER Epidemiologa y Salud Publica (CIBERESP), Granada; 12Department of Epidemiology, Murcia Health Council, CIBER Epidemiologa y Salud Publica
(CIBERESP) Murcia, Murcia; 13Navarre Public Health Institute, CIBER Epidemiologa y Salud Publica (CIBERESP), Pamplona; 14Public Health Division of Gipuzkoa
and Ciberesp, Basque Regional Health Department, San Sebastian, Spain; 15Department of Public Health and Primary Care, University of Cambridge, Cambridge;
16
Cancer Epidemiology Unit, Nuffield Department of Clinical Medicine, University of Oxford, Oxford, UK; 17National Institute for Public Health and the Environment
(RIVM), Bilthoven; 18Department of Gastroenterology and Hepatology, University Medical Centre Utrecht (UMCU), Utrecht; 19Julius Center for Health Sciences and
Primary Care, University Medical Center Utrecht, Utrecht, The Netherlands; 20WHO Collaborating Center for Food and Nutrition Policies, Department of Hygiene,
Epidemiology and Medical Statistics, University of Athens Medical School, Athens; 21Hellenic Health Foundation, Athens, Greece; 22Department of Cancer
Epidemiology, German Cancer Research Center, Heidelberg; 23Department of Epidemiology, German Institute of Human Nutrition Potsdam-Rehbrucke, Potsdam,
Germany; 24Department of Surgery, Skane University Hospital Malmo, Lund University, Malmo; 25Department of Internal Medicine, Division of Gastroenterology and
Hepatology, Sahlgrenska University Hospital, Gothenburg; 26Department of Medical Biosciences, Pathology, Umea University, Umea, Sweden; 27Department of
Public Health and Clinical Medicine, Nutritional Research, Umea University, Umea, Sweden; 28Department of Epidemiology, School of Public Health, Aarhus
University, Aarhus; 29Danish Cancer Society, Institute of Cancer Epidemiology, Diet Cancer and Health, Copenhagen, Denmark; 30Department of Community
Medicine, University of Troms, Tromso, Norway; 31International Agency for Research on Cancer (IARC-WHO), Lyon, France; 32Department of Epidemiology and
Biostatistics, School of Public Health, Imperial College London, London, UK; 33Institute of Molecular Pathology and Immunology of the University of Porto
(IPATIMUP) and Medical Faculty/HS Joao, Porto, Portugal; 34Department of Clinical Pathology, Odense University Hospital, Odense, Denmark; 35School of Public
Health, St Marys Campus, Imperial College London, London, UK

Received 29 April 2011; revised 6 July 2011; accepted 6 July 2011

Background: In epidemiological studies, Helicobacter pylori infection is usually detected by enzyme-linked

immunosorbent assay (ELISA). However, infection can spontaneously clear from the mucosa during the progression of
atrophy and could lead to substantial under-detection of infection and underestimation of its effect on gastric cancer
(GC) risk. Antibodies detected by western blot are known to persist longer after the loss of the infection.
Methods: In a nested casecontrol study from the Eurogast-EPIC cohort, including 88 noncardia GC cases and 338
controls, we assessed the association between noncardia GC and H. pylori infection comparing antibodies detected
by western blot (HELICOBLOT2.1) to those detected by ELISA (Pyloriset EIA-GIII
).
Results: By immunoblot, 82 cases (93.2%) were H. pylori positive, 10 of these cases (11.4%) were negative by ELISA
and only 6 cases (6.8%) were negative by both ELISA and immunoblot. Multivariable odds ratio (OR) for noncardia GC

*Correspondence to: Dr C. A. Gonzalez, Unit of Nutrition Environment and Cancer,

Department of Epidemiology, Catalan Institute of Oncology (ICO), Gran Via s/n
Hospitalet, Barcelona 34-08908, Spain. Tel: +34-932607401; Fax: +34-932607787;
E-mail: cagonzalez@iconcologia.net

The Author 2011. Published by Oxford University Press on behalf of the European Society for Medical Oncology.
All rights reserved. For permissions, please email: journals.permissions@oup.com
original article
comparing immunoglobulin G positive versus negative by ELISA was 6.8 [95% confidence interval (CI) 3.015.1], and
by immunoblot, the OR was 21.4 (95% CI 7.164.4).
Conclusions: Using a western blot assay, nearly all noncardia GC were classified as H. pylori positive and the OR
was more than threefold higher than the OR assessed by ELISA, supporting the hypothesis that H. pylori infection is
a necessary condition for noncardia GC.
Key words: Helicobacter pylori, noncardia gastric cancer, prospective study, western blot
introduction
Helicobacter pylori infection is a well-established cause of
sporadic noncardia gastric cancer (GC) [1]. In epidemiological
studies, H. pylori infection is usually detected by enzyme-linked
immunosorbent assay (ELISA). A combined analysis of 12
casecontrol studies nested within cohorts [2] found an overall
odds ratio (OR) of 3.0 [95% confidence interval (CI) 2.33.8]
for the risk of noncardia cancer, measuring anti-H. pylori
immunoglobulin G (IgG) antibodies by ELISA. In a meta-
analysis [3] of 10 casecontrol studies (using ELISA in 8 studies
and western blot in 2) in which the infection by H. pylori was
assessed by CagA seropositivity, the OR for noncardia GC was
2.71 (95% CI 1.74.2).
Severe chronic atrophic gastritis (SCAG) and extended
intestinal metaplasia are believed to create conditions in which
H. pylori is unable to survive, and, as a consequence, there is
a gradual reduction of H. pylori colonization of the gastric
mucosa during the progression of preneoplastic lesions.
Therefore, there is a potential to underestimate the association,
mainly in casecontrol studies, because of under-detection of
H. pylori infection in blood collected after the diagnosis of GC.
It has been shown that anti-CagA antibodies measured by
western blot analysis (immunoblot) persist longer than specific
IgG antibodies detected by conventional ELISA [48], and
therefore, it may be possible to detect previous H. pylori infection
in some ELISA-negative subjects using immunoblot. These studies
have shown that the association between H. pylori infection and
GC is much stronger than considered previously and led to the
hypothesis that H. pylori may be a necessary cause of GC [9].
In the European Prospective Investigation into Cancer and
Nutrition (Eurgast-EPIC) study, we have previously reported
[10] that using only antibodies against H. pylori lysate, there
was non-statistically significant association with noncardia GC.
The aim of this study is to assess the magnitude of the risk
associated with H. pylori infection in a new set of noncardia GC
from the Eurogast-EPIC study, comparing antibodies detected
by western blot analysis to those detected by ELISA.
materials and methods
subjects
The study subjects belong to the EPIC cohort [11]. Briefly, the EPIC cohort
includes about half million individuals, mostly aged 4065 years, recruited
between 1992 and 1998 in 23 centres from 10 European countries:
Denmark, France, Greece, Germany, Italy, The Netherlands, Norway, Spain,
Sweden and UK. At enrolment, blood samples were collected from most
participants. Follow-up is based upon population cancer registries in most
countries, except in France, Germany and Greece, where it is mainly
achieved by active contact with study subjects and review of health
insurance and pathology reports.
2 | Gonzalez et al.
Annals of Oncology
A nested casecontrol study within the EPIC cohort (EurGast) was
conducted to analyse the relationship between GC risk and baseline H.
pylori seropositivity status as well as other biomarkers. Each incident
noncardia GC case with an available blood sample was matched by sex, age
group (62.5 years), centre and date of blood collection (645 days) to four
control participants who were randomly selected from the cohort at risk at
the time of diagnosis of the index case. Cases were subjects newly diagnosed
of GC defined by code C16 of the International Classification of Diseases,
10th Revision (ICD-10). Noncardia GC were defined by the code C16.1 to
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C16.9. The previous analysis [10] included 233 GC cases (diagnosed mostly
before 2000) and 910 controls, using only ELISA for detection of H. pylori
antibodies. Cases for this new analysis, using ELISA and immunoblot, were
identified during an update of cancer incidence within the cohort that
included new cases diagnosed in the same EPICs participating countries
mostly between 2000 and 2004. An independent panel of pathologists
reviewed the original slides and pathology reports provided by each EPIC
centre for most of the cases, in order to confirm and validate the diagnosis,
tumour site and morphology [12].
determination of the H. pylori status by ELISA
Helicobacter pylori infection was determined by ELISA using the Pyloriset

EIA-GIII kit (Orion Diagnostica, Espoo, Finland). Briefly, a 1/200 dilution of
serum in buffer was introduced in H. pylori-coated microtiter wells. After 30
min incubation, the wells were washed and a peroxidase conjugated anti-
human IgG from rabbit was incubated for a further 30 min. After washing, the
tetramethylbenzidine substrate was added for 10 min and the optical density
(OD) measured at 450 nM. The results were expressed as unit per milliliter
according to a calibrator curve. A value 20 U/ml was considered as positive.
determination of the H. pylori status and the CagA status
by immunoblot
Immunoblot using the HELICOBLOT 2.1 kit (Genelab Diagnostics,
Singapore) was carried out. An H. pylori lysate enriched with recombinant
antigens is electrophoretically prepared and transferred on to the nitrocellular
strips, which are commercially available. Individual strips were incubated
with a 1/100 dilution of serum in blotting buffer on a tray. After 60 min of
incubation, the strip was washed and a goat anti-human IgG conjugated with
alkaline phosphatase was added for a further 60 min. After washing, the 5-
Bromo-4 chloro-3 indolylphosphate and nitrobluetetrazolium substrate was
added for 15 min. The criteria for H. pylori positivity were the presence of
one of the following bands 89 KD, 37 KD, 35 KD or both the 30 KD and 19.5
KD bands, and for CagA positivity, the presence of a 116 KD band.
determination of pepsinogen I levels
Pepsinogen I level was determined by ELISA using the kits from Biohit
(Helsinki, Finland). Human pepsinogen-captured antibodies (monoclonal) are
absorbed in microtiter wells. A dilution of 1/5 of the serum was added. After
60 min incubation, a peroxidase conjugated anti-human pepsinogen was added
for a further 60 min. After washing, the tetramethylbenzidine substrate was
added for 30 min and the OD was measured at 450 nM. A calibration curve was
used to calculate the pepsinogen concentration. All determinations were carried
out at room temperature. A pepsinogen 1 level <22 lg/l indicated SCAG.
Annals of Oncology
statistical analysis
Data are presented as the number and proportion of controls and cases for
each H. pylori assay used. Conditional logistic regression was used to
estimate the OR for noncardia GC risk, adjusting for potential confounders:
education (none, primary, technical/professional, secondary or university),
cigarette smoking (never, former and current) and average daily dietary
intakes (fruit, vegetables and red and processed meat).
results
During a mean follow-up of 10.65 years (range 0.317.6 years),
in the update of cancer incidence from the EPIC cohort, 195
new histologically confirmed adenocarcinoma cases (51 from
the cardia, 88 from the noncardia, 4 mixed and 52 for which
the site was not identified) were diagnosed. The analysis
presented here is based on these 88 noncardia GC cases (of
which 31 were intestinal type, 32 diffuse and 25 mixed or
undefined) and 338 matched controls. In 16 (18.2%) of these
cases (Table 1), SCAG was present according to serological
original article
was higher than those positive by ELISA (11.1%). Similar
results were observed by immunoblot. We did not observe
any differences regarding the proportion of negative cases
between intestinal and diffuse histological subtypes (data not
shown).
Table 2 shows the ORs of noncardia GC associated with H.
pylori seropositivity, according to the assay, adjusting for
potential confounders. The OR was 6.8 (95% CI 3.015.1)
comparing IgG positive versus negative by ELISA and 21.4
(95% CI 7.164.3) comparing IgG positive versus negative by
immunoblot.
discussion
Our prospective study found that the OR for noncardia GC and
H. pylori seroprevalence is almost fourfold higher by
immunoblot than by ELISA. It supports the evidence that
measuring only IgG by ELISA to classify H. pylori infection

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status may lead to a substantial underestimation of the effect of
levels of pepsinogen I. The median serum concentration of H. pylori infection in noncardia GC risk [4, 8].
pepsinogen I for all noncardia cases was 64.3. The mean years Similar results have been observed in three previous studies
of follow-up of noncardia cases with SCAG was relatively that compared results from ELISA with immunoblot. In
similar to those without SCAG (9.0 and 10.7 years, a population-based casecontrol study from Sweden [4]
respectively), but cases with SCAG were significantly older (including 206 noncardia GC and 238 controls), the adjusted
compared with those without SCAG (mean age at blood
extraction 61.3 and 55.3 years, respectively, P 0.001).
Table 2. ORa and 95% CI for the association of exposure to Helicobacter
The seroprevalence of H. pylori antibodies in noncardia
pylori infection detected through IgG ELISA or immunoblot and
cases and controls by assay is shown also in Table 1. In 16
noncardia gastric cancer risk in the EurGast-EPIC study
cases (18.2%), IgG antibodies detected by ELISA were
negative (eight without SCAG and eight with SCAG). Only six
cases (6.8%) (three with SCAG and three without SCAG) H. pylori infection Cases/controls OR (95%
detected through (88/338) CI)
were negative for both assays, while 82 (93.2%) of noncardia
GC cases were positive for previous H. pylori infection by H. pylori IgG negative 16/150 1.0 (reference)
immunoblot. Among controls, 188 (55.7%) were positive by (ELISA)
ELISA, while 199 (58.9%) were positive by immunoblot. We H. pylori IgG positive 72/188 6.81 (3.0115.08)
observed 9 controls that were positive by ELISA but negative (ELISA)
by immunoblot and 20 controls negative by ELISA but H. pylori IgG negative 6/139 1.0 (reference)
(Immunoblot)
positive by immunoblot. Concordance by immunoblot assay
H. pyrlori IgG positive 82/199 21.42 (7.1364.35)
between H. pylori antibodies and CagA antibodies for all cases
(Immunoblot)
and controls was almost complete, and we observed only one
case and four controls reported as negative for CagA but CI, confidence interval; ELISA, enzyme-linked immunosorbent assay; IgG,
positive for H. pylori by immunoblot. Out of the 52 cases for immunoglobulin G; OR, odds ratio.
which the anatomical site was undefined, 6 cases (11.5%) (all a
Conditional logistic regression (matched by age, sex, centre and date of
without SCAG) were negative for the two assays. The blood extraction) adjusted by smoking status, school level, red and
proportion of cases with SCAG but negative by ELISA (50%) processed meat intake and fruit and vegetables consumption.

Table 1. Seroprevalence of Helicobacter pylori IgG antibodies (ELISA) and IgG antibodies (Immunoblot) in noncardia gastric adenocarcinoma cases and
matched controls, in the EurGast-EPIC cohort study

Test results Controls Cases

ELISA Immunoblot N (%) PI median PI < 22 N (%) PI median PI < 22
IgG H. pylori (25th75th percentile) lg/ml n (%) (25th75th percentile) lg/ml n (%)
2 2 130 (38.5) 58.9 (46.376.4) 3 (2.3) 6 (6.8) 12.2 (8.658.4) 3 (50.0)
2 + 20 (5.9) 53.4 (46.293.1) 1 (5.0) 10 (11.4) 28.9 (9.172.8) 5 (50.0)
+ 2 9 (2.7) 62.2 (56.377.4) 1 (11.1) 0 0
+ + 179 (53.0) 69.6 (53.287.6) 6 (3.4) 72 (81.8) 68.9 (41.593.6) 8 (11.1)
Total 338 (100) 65.1 (49.780.5) 11 (3.3) 88 (100) 64.3 (37.790.7) 16 (18.2)

ELISA, enzyme-linked immunosorbent assay; IgG, immunoglobulin G; PI, pepsinogen I.

doi:10.1093/annonc/mdr384 | 3
original article
OR for seropositivity of H. pylori infection was 2.2 as measured
by ELISA and 21.0 as measured by immunoblot. In a hospital-
based casecontrol study from Germany [9] (including 68
noncardia GC and 360 controls), the OR increased from 3.7 by
ELISA to 18.3 by immunoblot. Finally, in a nested casecontrol
study from Australia [8] (including 34 noncardia GC and 134
controls), the OR increased from 2.3 by ELISA to 10.6 by
immunoblot.
We found that the prevalence of seronegativity in noncardia
GC decreased from 18.2% by ELISA to 6.8% by immunoblot,
confirming the underestimation of the prevalence of H. pylori
infection using ELISA. In a meta-analysis [3] of casecontrol
studies, which included 10 studies (western blot was used only
in two of these studies), and >1700 noncardia GC cases, which
assessed the relationship between GC and H. pylori by CagA
seropositivity, 37.7% of the noncardia GC cases were negative
for CagA antibodies. On the contrary, using immunoblot in the
population-based casecontrol study from Sweden [4], only 4%
of noncardia GC cases were negative, while in the nested case
control study from Australia [8], it was 6%, which are both
relatively similar to the proportion observed in our study. It
was recently estimated that the H. pylori attributable fraction of
noncardia GC cases is 74% in developed countries and 78% in
developing countries [13]. Our results, as well as those from
other studies indicate that this fraction may be higher, and that
H. pylori infection could be a necessary condition of noncardia
GC [9]. A necessary cause is not necessarily a sufficient cause
(meaning other factors are required). It is well known that
despite the high prevalence of H. pylori infection, only a small
proportion of infected subjects develop GC. This underlines the
relevance of the role of other cofactors, such as diet, smoking
and genetic susceptibility, acting as component of a sufficient
cause, in the same or different causal mechanisms, depending
of the relative prevalence of these factors in a population [14].
Our results suggest that the underestimation of the
prevalence of H. pylori infection by ELISA is greater in cases
with SCAG, which is expected since during progression of
gastric atrophy, the bacterium cleans from the gastric mucosa.
As was expected in our study, noncardia cases with SCAG were
older than cases without SCAG. In our study, SCAG was
defined as pepsinogen I levels <22 lg/l; this cut-off has
previously demonstrated to have relatively high sensitivity
(89.5%) and specificity (91.5%) for the screening of SCAG in
the general population [15].
We observed 5 cases without SCAG out of 10 noncardia GC
that were H. pylori negative by ELISA but positive by
immunoblot. In a study designed to assess the current mucosal
condition of 64 GC cases through histology and culture [16], 4
out of 12 cases that were H. pylori negative by ELISA and CagA
positive by immunoblot had no evidence of gastric atrophy. In
addition, no evidence of atrophy was found in 8 out of 10 cases
that were negative by both serological tests. In our study, we
found that three noncardia GC cases were negative for H. pylori
infection by both assays and had no evidence of SCAG. We also
checked the available pathological information for the six
noncardia GC cases that were H. pylori negative, to verify that
there were no errors in the anatomical subsite.
The best method to diagnose H. pylori infection in different
research situations is still not clarified [17]. We used
4 | Gonzalez et al.
Annals of Oncology
a commercial immunoblot test (HELICOBLOT 2.1) that has
been shown to have a sensitivity of 100% for current and of
92% for previous H. pylori infection [17, 18], meaning that
some false-negative results can be expected in subjects with
previous infection. We observed that 93.2% of noncardia GC
were positive by immunoblot, which is consistent with the
sensitivity of the method.
Our study has several clinical implications. The immunoblot
might be useful for modelling the relationship between the
decline in H. pylori infection prevalence and declines in gastric
cancer incidence [19]. Our study should not be a basis for
justification of massive eradication therapy, because there is no
evidence yet of the benefit of this approach in the general
population [20]. Furthermore, this could lead to the activation
of antibiotic-resistant strains of other pathogens [1]. However,
it is a good reason to continue efforts for developing vaccines
against H. pylori.
Our study design has several strengths. Most of the cases
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were validated by a panel of expert pathologists. The
prospective nature of EPIC allowed H. pylori serology to be
accurately determined in healthy subjects before the onset of
the disease, which is an important strength in comparison to
casecontrol studies. In addition, as far as we know, this is the
first prospective study in European populations showing results
of both assays in noncardia GC cases. A limitation is the
relatively small sample size of noncardia GC cases. The length
of follow-up is also relatively short and does not allow to
estimate changes in the magnitude of risk according to the time
between blood collection and cancer diagnosis.
In conclusion, we found that the proportion of H. pylori-
negative cases by ELISA is reduced by threefold when results of
western blot are taken into account and accordingly, the OR is
more than threefold higher than that assessed by ELISA. Our
results support the hypothesis that H. pylori infection may be
a necessary cause of sporadic noncardia gastric cancer. Given
that the sensitivity of immunoblot is <100%, an improvement
in the accuracy of tests for the detection of past H. pylori
infection is still needed to clarify if this small fraction of
negative cases is indeed false negative.
acknowledgements
The authors want to acknowledge the pathology panel
members: Fatima Carneiro, Hendrik Blaker, Claus Laszlo Igali,
Gabriella Nesi and Roger Stenling for their contribution to the
collection and review of paraffin tumour blocks, slides and
pathology reports.
funding
This work was supported by the Health Research Fund (FIS) of
the Spanish Ministry of Health (Exp P10710130), La Caixa (BM
06-130) and Red Tematica de Investigacion Cooperativa en
Cancer (R06/0020) (Spain). The coordination of EPIC is
financially supported by the European Commission
(DGSANCO) and the International Agency for Research on
Cancer. The national cohorts are supported by the Health
Research Fund (FIS) of the Spanish Ministry of Health,
Regional Governments of Andaluca, Asturias, Catalunya,
Annals of Oncology
Basque Country, Murcia and Navarra (Spain); Danish Cancer
Society (Denmark); Liguecontre le Cancer, 3M, Mutuelle
Generale de lEducation Nationale, Institut National de la Sante
et de la Recherche Medicale (France); Deutsche Krebshilfe,
Deutsches Krebsforschungszentrum and Federal Ministry of
Education and Research (Germany); Ministry of Health and
Social Solidarity, Stavros Niarchos Foundation and Hellenic
Health Foundation (Greece); Italian Association for Research
on Cancer (AIRC) and National Research Council (Italy);
Dutch Ministry of Public Health, Welfare and Sports (VWS),
Netherlands Cancer Registry (NKR), LK Research Funds,
Dutch Prevention Funds, Dutch ZON (Zorg Onderzoek
Nederland), World Cancer Research Fund (WCRF), and
Statistics Netherlands (The Netherlands); Norwegian Cancer
Society (Norway); Swedish Cancer Society, Swedish Scientific
Council and Regional Government of Skane and Vasterbotten
(Sweden); Cancer Research UK, Medical Research Council,
Stroke Association, British Heart Foundation, Department of
Health, Food Standards Agency, and Wellcome Trust (UK).
Some authors are members of Environmental Cancer Risk,
Nutrition and Individual Susceptibility, a network of excellence
funded by the European Commission (6th EU Framework
Programme for Research and Development).
disclosure
The authors declare no conflicts of interest.
references
1. Correa P, Houghton JM. Carcinogenesis of Helicobacter pylori. Gastroenterology
2007; 133: 659672.
2. Helicobacter and Cancer Collaborative Group. Gastric cancer and Helicobacter
pylori: a combined analysis of 12 case control studies nested within prospective
cohorts. Gut 2001; 49: 347353.
3. Huang JQ, Zheng GF, Sumanac K et al. Meta-analysis of the relationship
between cagA seropositivity and gastric cancer. Gastroenterology 2003; 125(6):
16361644.
4. Ekstrom AM, Held M, Hansson LE et al. Helicobacter pylori in gastric cancer
established by CagA immunoblot as a marker of past infection. Gastroenterology
2001; 121(4): 784791.
original article
5. Sorberg M, Engstrand L, Strom M et al. The diagnostic value of enzyme
immunoassay and immunoblot in monitoring eradication of Helicobacter pylori.
Scand J Infect Dis 1997; 29(2): 147151.
6. Mini R, Annibale B, Lahner E et al. Western blotting of total lysate of Helicobacter
pylori in cases of atrophic body gastritis. Clin Chem 2006; 52(2): 220226.
7. Enroth H, Kraaz W, Rohan T et al. Does the method of Helicobacter pylori
detection influence the association with gastric cancer risk? Scand J
Gastroenterol 2002; 37(8): 884890.
8. Mitchell H, English DR, Elliott F et al. Immunoblotting using multiple antigens is
essential to demonstrate the true risk of Helicobacter pylori infection for gastric
cancer. Aliment Pharmacol Ther 2008; 28(7): 903910.
9. Brenner H, Arndt V, Stegmaier C et al. Is Helicobacter pylori infection a necessary
condition for noncardia gastric cancer? Am J Epidemiol 2004; 159(3): 252258.
10. Palli D, Masala G, Del Giudice G et al. CagA+ Helicobacter pylori infection and gastric
cancer risk in the EPIC-EURGAST study. Int J Cancer 2007; 120(4): 859867.
11. Riboli E, Hunt KJ, Slimani N et al. European Prospective Investigation into Cancer
and Nutrition (EPIC): study populations and data collection. Public Health Nutr
2002; 5(6B): 11131124.
12. Carneiro F, Moutinho C, Pera G et al. Pathology findings and validation of gastric
Downloaded from http://annonc.oxfordjournals.org/ by guest on September 20, 2015
and esophageal cancer cases in a European cohort (EPIC/EUR-GAST). Scand J
Gastroenterol 2007; 42(5): 618627.
13. Parkin DM. The global health burden of infection-associated cancers in the year
2002. Int J Cancer 2006; 118(12): 30303044.
14. Buehler JW. Surveillance. In Rothman KJ, Greenland S (eds), Modern
Epidemiology, 2nd edition. Philadelphia, PA: Lippincott Williams & Wilkins 1998.
15. Kekki M, Samloff IM, Varis K, Ihamaki T. Serum pepsinogen I and serum gastrin
in the screening of severe atrophic corpus gastritis. Scand J Gastroenterol Suppl.
1991; 186: 109116.
16. Ye W, Held M, Enroth H et al. Histology and culture results among subjects with
antibodies to CagA but no evidence of Helicobacter pylori infection with IgG
ELISA. Scand J Gastroenterol 2005; 40(3): 312318.
17. Monteriro L, Bergey B, Gras N, Megraud F. Evaluation of the performance of the
Helico Blot 2.1 as a tool to investigate the virulence properties of Helicobacter
pylori. Clin Microbiol Infect 2002; 8(10): 67969l.
18. Siman JH, Engstrand L, Berglund G et al. Evaluation of western blot CagA
seropositivity in Helicobacter pylori-seropositive and -seronegative subjects. Clin
Diagn Lab Immunol 2005; 12(2): 304309.
19. Yeh JM, Goldie SJ, Kuntz KM, Ezzati M. Effects of Helicobacter pylori infection
and smoking on gastric cancer incidence in China: a population-level analysis of
trends and projections. Cancer Causes Control 2009; 20(10): 20212029.
20. Herrera V, Parsonnet J. Helicobacter pylori and gastric adenocarcinoma. Clin
Microbiol Infect 2009; 15(11): 971976.
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