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Year : 2015  |  Volume : 21  |  Issue : 6  |  Page : 360-366
Rapid fecal calprotectin testing to assess for endoscopic disease activity in inflammatory bowel disease: A diagnostic cohort study

1 Department of Medicine, London Health Sciences Centre, University of Western Ontario, London, Ontario, Canada
2 Department of Medicine, London Health Sciences Centre, University of Western Ontario, London, Ontario; Department of Medicine, King Abdulaziz University, Jeddah, Kingdom of Saudi Arabia
3 Department of Pediatrics, London Health Sciences Centre, University of Western Ontario, London, Ontario, Canada
4 Department of Medicine, London Health Sciences Centre, University of Western Ontario, London, Ontario; Department of Pediatrics, London Health Sciences Centre, University of Western Ontario, London, Ontario, Canada
5 Department of Microbiology, London Health Sciences Centre, University of Western Ontario, London, Ontario, Canada

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Date of Submission12-May-2015
Date of Acceptance02-Sep-2015
Date of Web Publication4-Dec-2015


Background and Aim: With increasing numbers of patients diagnosed with inflammatory bowel disease (IBD), it is important to identify noninvasive methods of detecting disease activity. The aim of this study is to examine the diagnostic accuracy of fecal rapid calprotectin (FC) testing in the detection of endoscopically active IBD. Patients and Methods: All consecutive patients presenting to outpatient clinics with lower gastrointestinal symptoms were prospectively recruited. Patients provided FC samples. Sensitivity (Sn), specificity (Sp), positive predictive value (PPV), and negative predictive value (NPV) for FC were calculated. Receiver–operator characteristics (ROC) curve was used to identify the ideal FC cutoff that predicts endoscopic disease activity. Correlation between FC and endoscopic disease activity, disease location, and C-reactive protein (CRP) levels were measured. Results: One hundred and twenty-six patients, of whom 52% were females, were included in the final analysis with a mean age of 44.4 ± 16.7 years. Comparing FC to endoscopic findings, the following results were calculated: A cutoff point of 100 μg/g showed Sn = 83%, Sp = 67%, PPV = 65%, and NPV = 85%; and 200 μg/g showed Sn = 66%, Sp = 82%, PPV = 73%, and NPV = 77%. Based on ROC curve, the best FC cutoff point to predict endoscopic disease activity was 140 μg/g. Using this reference, FC levels strongly correlated with colorectal, ileocolonic, and ileal disease and predicted endoscopic activity. Conclusions: FC is an accurate test when used as an initial screening tool for patients suspected of having active IBD. Given its noninvasive nature, it may prove to reduce the need for colonoscopy and be an added tool in the management of IBD.

Keywords: Crohn's, diagnostic accuracy, inflammatory bowel disease, ulcerative colitis

How to cite this article:
Kwapisz L, Mosli M, Chande N, Yan B, Beaton M, Micsko J, Mennill PW, Barnett W, Bax K, Ponich T, Howard J, Tirolese A, Lannigan R, Gregor J. Rapid fecal calprotectin testing to assess for endoscopic disease activity in inflammatory bowel disease: A diagnostic cohort study. Saudi J Gastroenterol 2015;21:360-6

How to cite this URL:
Kwapisz L, Mosli M, Chande N, Yan B, Beaton M, Micsko J, Mennill PW, Barnett W, Bax K, Ponich T, Howard J, Tirolese A, Lannigan R, Gregor J. Rapid fecal calprotectin testing to assess for endoscopic disease activity in inflammatory bowel disease: A diagnostic cohort study. Saudi J Gastroenterol [serial online] 2015 [cited 2023 Jan 28];21:360-6. Available from:

Patients presenting to gastroenterologists with abdominal pain and/or diarrhea are often suspected of having either inflammatory bowel disease (IBD) or irritable bowel syndrome (IBS). Classically, IBD is a chronic disease that has a relapsing and remitting pattern and its two main entities include Crohn's disease (CD) and ulcerative colitis (UC). Symptoms typically associated with IBD include diarrhea, rectal bleeding, abdominal pain, and weight loss. Over the past several decades there has been a dramatic rise in the incidence of IBD worldwide.[1],[2] Similarly, there has been a parallel increase in the rates of IBS.[3],[4] Compared with IBS, untreated IBD can be associated with poor outcome. Therefore, distinguishing IBD from IBS is necessary.

Traditionally, both noninvasive tests such as C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) as well as invasive procedures such as ileocolonoscopy and flexible sigmoidoscopy have been used to diagnose and evaluate IBD for disease activity. The drawback to serological markers, such as CRP and ESR are that they lack specificity for intestinal inflammation and can be elevated in systemic inflammatory states. Conversely, the disadvantages to colonoscopy are rooted in the nature of being invasive, uncomfortable for patients, and being associated with risk of adverse events. As such, there has been a growing need for simple, cost-effective, and noninvasive tests to detect intestinal inflammation and aid in differentiating IBD from IBS.

Fecal calprotectin (FC) is a biomarker that has demonstrated promise in its specificity for intestinal inflammation.[5] Calprotectin is a calcium- and zinc-binding protein that constitutes nearly 60% of neutrophil cytosolic protein.[6] Levels of calprotectin therefore indicate the presence of neutrophils and other immune cells released from damaged intestinal mucosa.[7] Calprotectin's major functions include both antibacterial and antifungal activity, inhibition of metalloproteinases, and induction of apoptosis.[8] Two large meta-analyeis have showed that FC is a very useful tool in discriminating between IBD and IBS.[5],[9] The pooled sensitivity and specificity of FC was found to be as high as 93% and 96%, respectively. However, in those studies FC levels were measured using an enzyme-linked immunosorbent assay (ELISA) method and endoscopy was not always used as reference standard. More recently, less cumbersome and more efficient rapid point of care tests have been developed, which can reliably provide results within 15 min.[10],[11] A recent study showed that rapid FC testing has a positive correlation both to clinical and endoscopic severity in IBD.[12] This advantage may provide clinicians with the means to apply on site "personalized medicine" care in an efficient way. The calprotectin immunoassay by Quantum Blue (Buhlmann Laboratories, Schönenbuch, Switzerland) is designed to provide valuable information about the status of mucosal inflammation; can aid in predicting relapses and can detect calprotectin within a wide range of 100–1800 µg/g. These criteria in theory can provide even higher sensitivity in cases where inflammation is expected such as in patients with an established diagnosis of IBD.

The aim of our study is to evaluate the diagnostic accuracy of rapid point of care FC testing and correlate its results with endoscopic disease activity, disease location, and CRP as an aid in evaluating and establishing the diagnosis of IBD.

   Materials and Methods Top

Study population

We performed a prospective cohort study involving 130 consecutive adult patients, who were recruited between January 2013 and June 2014, presenting to specialized IBD outpatient gastroenterology clinics at London Health Sciences Centre (LHSC; University Hospital and Victoria Hospital) and St. Joseph's Hospital with signs and symptoms suggestive of active IBD. Diagnosis of IBD was based on typical clinical, endoscopic, and radiologic criteria supported by histological evidence of the disease. Inclusion criteria included age between 18 and 85 years as well as symptoms suggestive of IBD or a previously established diagnosis of IBD with symptoms of activity. All patients were investigated with either colonoscopy or flexible sigmoidoscopy within 4 weeks of their presentation as part of their diagnostic workup. Exclusion criteria included inability to perform endoscopy within the allocated time frame, a positive stool test for Clostridium difficile toxin or inability to provide a stool sample for analysis.

Study methods

Stool samples were collected from each patient (1:1) and subsequently analyzed for FC levels. Results were then compared with reports of the patient's endoscopic evaluation. Endoscopic assessments were performed by 10 board certified gastroenterologists, who were blinded to the results of the index test, as were the study personnel who received the FC results blinded to the endoscopic reports. The reference standard of endoscopic disease activity was used to determine the overall diagnostic accuracy of the test. Endoscopic evidence of active inflammation was noted when there was a mucosal break, exudate, or ulcerations present in keeping with traditional scoring systems such as the Mayo score for UC and simple endoscopic scoring for CD (SES-CD).[13],[14] In the absence of any mucosal abnormalities in the colon and terminal ileum; the endoscopic assessment was reported as normal. Data on demographics, medications, clinical parameters, and laboratory investigations were simultaneously collected.


The primary objective of this study was to assess the diagnostic accuracy of high-range rapid FC immunoassay in the diagnosis and assessment of IBD when compared with endoscopic activity. Secondary objectives included identifying the optimal FC cutoff point and using it to correlate FC levels with endoscopic disease activity, disease location, and CRP levels.

Methods of fecal calprotectin determination

All patients provided a stool sample within four weeks of their scheduled endoscopic evaluation. Less than 1 g of native stool was required. Samples could be kept refrigerated at 2°C—8°C for up to 6 days. If the samples could not be analyzed by the sixth day, they were kept frozen at −20°C and subsequently thawed when ready for measurement. All samples were analyzed by a high-range FC quantitative point of care test (FC-QPOCT), which is manufactured by Bühlmann, Quantum Blue (Buhlmann Laboratories, Schönenbuch, Switzerland).[15]

The QPOCT is a quantitative lateral flow assay, which with the help of a reader system provides results ranging from 100 to 1800 µg/g. The procedure consisted of three steps. First, samples were extracted. Next, 1:150 Chase Buffer solution was used to dilute the stool samples, followed by vortex and centrifuge. Lastly, 80 µL of diluted stool sample were loaded onto a cassette of a Quantum Blue Reader and a result was provided within 15 min. Two microbiologists who were appropriately trained were assigned to perform the FC measurements, while being blinded for endoscopic results and patients' clinical status.

Statistical analysis and sample size calculation

Descriptive statistics were expressed as means (±standard deviations (SD)) for continuous variables and as proportions for categorical variables. One-way ANOVA test was used to compare means. Sample size calculation was based on the premise that FC is a highly sensitive test for the evaluation of patients with suspected IBD. Assuming ileocolonoscopy is the gold standard with a sensitivity of 100%, and assuming a type 1 error of 0.05 and 80% power we estimated that 122 patients will need to undergo both tests to determine a sensitivity of fecal calprotectin of 95%.

Sensitivity (Sn), specificity (Sp), positive predictive value (PPV), and negative predictive values (NPV) were calculated using 100 and 200 µg/g as cutoff points and endoscopic activity as reference standard.

Simple linear regression was used to measure correlation. Pearson's correlation coefficient (r) and Spearman's rank correlation coefficient (rho) were used to express correlation between FC and CRP levels and disease location, respectively. Receiver operator characteristic (ROC) curve analysis was used to identify the FC-QPOCT and CRP points that best predict endoscopic disease activity for IBD. Simple and multiple logistic regression analyses were then used to characterize the relationship between abnormal FC levels, based on the cutoff point identified by ROC curve analysis, and disease location (rectal, colonic, colorectal, ileal and ileocolonic) and endoscopic activity (active versus inactive). Associations were expressed as odds ratios (OR). StataCorp (TX, USA) statistical software, version 12.0 was used for statistical analysis and a two-sided 5% significance level was used for all statistical inferences. Precision of estimates was measured using 95% confidence intervals (95% CI).

Ethical considerations

The Western University Research Ethics Board for Health Sciences Research Involving Human Subjects (HSREB) provided approval (#102975) for the conduction of this clinical study. All recruited patients provided written informed consent for participation in the study.

   Results Top

Patients' characteristics

Baseline characteristics of the study population are summarized in [Table 1]. Of the 170 patients who were eligible and offered enrollment, 130 provided stool samples for FC analysis. During the course of the study, four patients were not able to undergo endoscopy (colonoscopy or flexible sigmoidoscopy) and the final analysis was based on 126 patients. The mean age was 44.4 ± 16.7 and 52% were females. Exactly 50% of enrolled patients had known IBD prior to their endoscopy, whereas the other 50% had symptoms suggestive of IBD but had not yet been diagnosed. Endoscopic changes consistent with IBD were found in 53 (41%) patients, with 11 (8%) patients being newly diagnosed. Lastly, patients enrolled in the study presented to gastroenterologists with a variety of symptoms related to IBD including sole diarrhea at 22%, rectal bleeding at 21%, abdominal pain at 18%, and a mixture of multiple symptoms present 39% of the time.
Table 1: Baseline demographics and clinical characteristics (N=126)

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Primary end points

In this particular study, a newer version of a point of care immunoassay for FC was used to determine the diagnostic accuracy of the test. According to the manufacturer, values below 100 μg/g are indicative of either a noninflammatory state or mild inflammation in the gut. The results in our study for FC levels below 100 μg/g generated Sn = 83%, Sp = 67%, PPV = 65%, and NPV = 85%. Conversely, using FC values greater than 200 μg/g as a cutoff point, which is intended by the manufacturer to indicate active organic disease with inflammation in the gastrointestinal tract, resulted in Sn = 66%, Sp = 82%, PPV = 73%, and NPV = 77% [Table 2].
Table 2: Diagnostic accuracy results for FC; cut-off points of <100, <200, and <140 µg/g, and for CRP using cut-off points s of <5, <7, and <10 mg/L

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Secondary end points

For our cohort of patients, 60% had levels of CRP measured. Compared with active inflammation observed through endoscopy, CRP values (using a cutoff of 5 mg/dL) in this study showed Sn = 44%, Sp = 72%, PPV = 63%, and NPV = 54% [Table 2].

Location of inflammation was also recorded and correlated to FC levels. Overall, 29 patients had active disease limited to the colon or rectum (55%), 16 patients had activity localized to the ileum (30%), and 8 patients had ileocolonic involvement (15%). Within each location, Sn was calculated and found to be 90% for the colon/rectum, 88% for ileocolonic disease, and 69% for the ileum. Also, consistently higher FC levels were observed for active disease that had at least partial colonic involvement (whether colon and rectum or ileocolonic). When only the terminal ileum was involved, a much lower sensitivity was observed as well as lower overall FC levels. There was a statistically significant difference between groups as determined by one-way ANOVA (F (4,48) = 5.04, P =0.0018) [Table 3].
Table 3: Disease location and its impact on FC sensitivity

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To better characterize the correlation between FC and CRP, simple linear regression was performed. FC, though weakly, nevertheless significantly positively correlated with CRP levels (r = 0.017, 95% CI: 0.006–0.03, P = 0.003) [Figure 1] and more strongly positively correlated with disease location (rho = 0.5191, 95% CI: 0.006–0.03, P ≤ 0.00001).
Figure 1: A scatter plot correlating fecal calprotectin with C-reactive protein levels

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ROC curve analysis identified 140 μg/g as the optimal FC cutoff point to predict IBD endoscopic disease activity (area under the curve (AUC) =0.81, 95% CI: 0.73–0.87) with Sn = 77%, Sp = 73%, positive likelihood ratio (LR+) = 2.8 and negative likelihood ratio (LR−) =0.3 [Figure 2]. Similarly, the optimal cutoff point for CRP was identified as 4 mg/dL (area under the curve (AUC) =0.584, 95% CI: 0.46–0.69) with Sn = 60%, Sp = 60%, positive likelihood ratio (LR+) = 1.5 and negative likelihood ratio (LR−) = 0.7.
Figure 2: Receiver operator characteristic curve analysis identified 140 μg/g and 4 mg/dL, as the optimal cutoff points for fecal calprotectin and C-reactive protein, respectively

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Using this cutoff point, FC results were dichotomized into normal and abnormal. Simple and multiple logistic regression analyses were performed. Abnormal FC was highly predictive of endoscopic disease activity on simple (OR = 9.7, 95% CI: 4.24–22.25; P ≤ 0.0001) and multiple (OR = 11.3, 95% CI: 2.68–47.68; P = 0.001) logistic regressions (adjusted for disease location). FC was also predictive of disease location (OR = 1.99, 95% CI: 1.43–2.77; P ≤ 0.0001). Similar analysis was performed to identify disease locations that predicted an abnormal FC test result. A significant positive association was observed with colorectal (OR = 27.14, 95% CI: 3.27–225.15; P = 0.002), ileocolonic (OR = 19, 95% CI: 2.21–163.79; P ≤ 0.007) and ileal disease (OR = 3.49, 95% CI: 1.15–10.56; P ≤ 0.027), and colonic involvement by itself perfectly predicted an abnormal FC level as no patient with active colonic disease had a normal FC result. There was a significantly negative association between FC and rectal disease (OR = 0.085, 95% CI: 0.035–0.205; P ≤ 0.0001) [Figure 3].
Figure 3: Box plots correlating fecal calprotectin levels with (a) disease location, and (b) endoscopic findings

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   Discussion Top

In this study, we examine the diagnostic accuracy with a rapid FC point of care assay (FC-QPOCT) and correlate its levels to disease locations, CRP levels, and endoscopic disease activity. We used a new POC test that has high-range detection capability but is also designed to provide results within 15 min, allowing a much more efficient and useful method compared with prior arduous ELISA methods. The results we report are consistent with it being a highly accurate method of FC measurement in relation to a well-established gold standard, which has not been used as comparator in many previously reported studies.[16],[17],[18] A recent study by Lobaton et al. reported, using a cutoff point of 160 μg/g, an Sn of 64.9% and an Sp of 83.9% with the similar FC assay and demonstrated a very strong correlation with an ELISA-based FC assays (Intra-class correlation coefficient (ICC = 0.904, 95% CI: 0.864-0.932; P = 0.001).[18] Since the literature supports that disease activity, based on clinical criteria, does not highly correlate with endoscopic disease activity in IBD patients, especially with CD [19] we elected not to use this as a clinical endpoint.

With a growing need to find new, reliable, and sensitive biomarkers in the evaluation of IBD, FC has proven to be quite valuable in discriminating between IBD and IBS.[20] The population of patients we included in this study is representative of the majority of patients with IBD, with a prominent bi-modal age group distribution. Two evenly matched groups of patients were enrolled into the study with either signs or symptoms suggestive of IBD or established diagnosis of IBD with symptom activity. This adds to the generizability of our results. It was found that for the lower cutoff value of 100 μg/g, Sn of 83% and NPV of 85% were observed. For the higher cutoff of 200 μg/g, a higher overall Sp of 82% and PPV of 73% were noted. These figures are in line with previously reported diagnostic accuracy studies for ELISA-based FC assays.[21],[22],[23] The high NPV and low LR − we report, can be useful clinical characteristics of FC to consider when it is used to triage IBD patients presenting with lower GI symptoms for colonoscopy, as negative results would be highly suggestive of inactive disease.

Further subanalysis demonstrated that areas of active disease played a prominent role in the diagnostic accuracy of FC. For colonic involvement, the overall sensitivity of FC was much better. With sole involvement of the ileum or the rectum, FC levels proved to be much less sensitive. Furthermore, there were six patients who had evidence of pancolitis and their average FC levels were the highest observed at an average of 1710. It can be surmised from our study, that the high-range immunoassay used for FC is potentially much more valuable and reliable for colonic IBD and cannot be relied on in patients with distal "rectal" UC.

Classically, CRP has been considered a poor marker of IBD activity with reported sensitivity estimates as low as 50%.[24] In our study, only 60% of patients received blood work, which included CRP levels prior to their endoscopy. We recognize the limitation around this analysis, but still found Sn of 44%, Sp of 72%, NPV of 54%, and PPV of 63% and that CRP poorly correlated with FC-QPOCT. One can see FC's superiority over CRP despite the limited data.

The ideal cutoff point for FC tests has been a subject of debate and results have varied widely.[25],[26],[27] Our results, which are based on a properly powered study sample, suggest that 140 μg/g most accurately predicts endoscopic disease activity and this correlated well with endoscopic disease activity and disease location. The limitation to this analysis is that the FC-QPOCT does not report results below 100 or above 1800 μg/g and therefore cutoff points outside that range could not be assessed.

Lastly, it is important to mention that we did not have strict exclusion criteria and acknowledge that there are a variety of conditions that can artificially increase FC levels as studies show that benign conditions such as menstrual bleeding and epistaxis as well as diseases such as bacterial enteritis/colitis, eosinophilic esophagitis, microscopic colitis, peptic ulcer disease, colorectal cancer, polyps, diverticulitis, and nonsteroidal anti-inflammatory drugs can all induce an increase in FC readings.[28],[29],[30],[31],[32],[33] Overall, 23 patients in our study were found to have polyps on endoscopy. Of those patients, 64% had elevated FC levels without endoscopic evidence of IBD. As a result, one can see that the presence of polyps can certainly affect the interpretation of FC levels. Furthermore, some important clinical confounders such as disease severity were not controlled for in our analyses and we acknowledge these limitations. Nevertheless, rapid point of care FC assays are highly accurate in detecting endoscopic disease activity for IBD, especially colonic disease, and can be used to triage patients for colonoscopy in an effort to preserve resources and improve patient care.

   Conclusions and Recommendations Top

Although FC has shown to be a very useful addition to the diagnosis of IBD, many questions still remain. An optimum cutoff has not yet been determined in literature and even though FC has shown tremendous promise in discerning between IBD and IBS, its exact future remains unclear. FC's destiny may lie as a biomarker that is used to trend response to therapy in disease states or be a simple tool that primary care doctors along with gastroenterologists can use to screen for IBD prior to endoscopy. In our study, it is important to note that FC levels were much more accurate when disease activity was found in the colon, this factor suggests that its use may be more beneficial in UC and Crohn's colitis versus noncolonic CD. Further studies are certainly on the horizon for FC and in time it may prove to be a very valuable tool for daily use in the investigation of numerous gastrointestinal diseases. Based on our results, we recommend using FC-QPOCT to screen patients suspected of having IBD using a cutoff point of 140 μg/g to discriminate between active and inactive disease.


We would like to acknowledge and thank all the nurses and hospital staff who assisted us in acquiring the study samples.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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Correspondence Address:
Dr. Mahmoud Mosli
Department of Medicine, King Abdulaziz University Hospital, King Abdulaziz University, Jeddah, Kingdom of Saudi Arabia

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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/1319-3767.170948

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