Use of the urinary trypsinogen-2 dipstick test in early diagnosis of pancreatitis after endoscopic retrograde cholangiopancreatography (ERCP)

doi:10.4236/ojgas.2013.36049

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Open Journal of Gastroenterology, 2013, 3, 289-294 OJGas

http://dx.doi.org/10.4236/ojgas.2013.36049 Published Online October 2013 (http://www.scirp.org/journal/ojgas/)

Use of the urinary trypsinogen-2 dipstick test in early

diagnosis of pancreatitis after endoscopic retrograde

cholangiopancreatography (ERCP)

Hasan El-Garem1, Enas Hamdy2, Sherif Hamdy1, Mohammad El-Sayed1*, Aish a Elsharkawy1,

Azmi Mohammed Saleh1

1Department of Endemic Medicine and Hepatology, Faculty of Medicine, Cairo University, Cairo, Egypt

2Department of Clinical Pathology, Faculty of Medicine, Cairo University, Cairo, Egypt

Email: *[email protected]

Received 20 August 2013; revised 25 September 2013; accepted 6 October 2013

which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

ABSTRACT

Background: Acute pancreatitis is one of the most se-

rious complications of ERCP. Early diagnosis of post

ERCP pancreatitis helps physicians to provide inten-

sive care and possible medical treatment as early as

possible. Trypsinogen-2 in urine is a good diagnostic

and prognostic marker of acute pancreatitis. Objec-

tives: To evaluate the diagnostic value of urinary

trypsinogen-2 dipstick test for early diagnosis of post

ERCP pancreatitis. Methods: A total of 37 patients

with obstructive jaundice were tested with the uri-

nary trypsinogen-2 dipstick test and serum levels of

amylase and lipase before ERCP and 6 hours after

ERCP. Results: Post ERCP pancreatitis was diag-

nosed in 6 (16%) of 37 patients. The sensitivity, speci-

ficity, positive predictive value and negative predic-

tive value of urinary trypsinogen-2 dipstick test at 6

hours after ERCP were 100%, 97%, 86%, 100% re-

spectively. At the cutoff level (130 U/L) for lipase, the

positive predictive value and negative predictive va-

lue all were (100%), however, the positive predictive

value and negative predictive value for amylase levels

at cutoff (122 U/L) were 60%, 100% respectively. Se-

rum lipase level was the best test for diagnosing post

ERCP pancreatitis followed by the urinary trypsino-

gen-2 dipstick test. Conclusions: The urinary trypsi-

nogen-2 dipstick test can be used as a rapid and easy

test for early diagnosis of post ERCP pancreatitis

with high sensitivity and specificity.

Keywords: ERCP; Pancreatitis; Urinary Trypsinogen-2

Dipstick Test

1. INTRODUCTION

Since its first reported application in 1968, endoscopic

retrograde cholangiopancreatography (ERCP) has be-

come a valuable procedure for examination and treat-

ment of pancreaticobiliary diseases [1]. One of the most

serious complications of (ERCP) is acute pancreatitis.

The reported incidence varies from 1.3% to 24.4% [2].

Measurement of serum amylase and lipase levels after

the procedure may have a possible role for early recog-

nition of post-ERCP pancreatitis [3]. Asymptomatic ele-

vation in serum amylase and lipase activities after ERCP

is common, occurring in approximately 25% to 75% of

all patients, owing to the lack of specificity of pancreatic

enzymes [4]. Reports indicate that serum and urinary

trypsinogen-2 concentrations increase in patients with

acute pancreatitis and that trypsinogen-2 levels can ser-

ve as a more sensitive diagnostic marker for pancreatitis

relative to amylase or lipase [1,5]. A rapid test strip has

been developed for the detection of trypsinogen-2 in

urine (The urinary trypsinogen-2 dipstick test—UT2DST-

actim pancreatitis) which is based on the immunochro-

matography principle and shows a good sensitivity and

specificity in diagnosing acute pancreatitis [6].

The aim of this study was to evaluate the diagnostic

value of urinary trypsinogen-2 dipstick test for early

diagnosis of post-ERCP pancreatitis.

2. PATIENTS AND METHODS

Thirty seven adult patients with obstructive jaundice

were referred to Kasr El-Aini Endoscopy Unit to perform

ERCP.

2.1. Inclusion Criteria

1) Adult patients above 18 years.

*Corresponding author.

OPEN ACCESS

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290

2) Patients having biliary obstruction as diagnosed by

elevated serum bilirubin and abdominal imaging show-

ing biliary obstruction.

2.2. Exclusion Criteria

1) Pancreatitis within 60 days of ERCP.

2) History of drug that may lead to acute pancreatitis.

3) Age less than 18 years.

4) Pregnant patients.

5) Known renal disease.

6) Known acute pancreatitis.

7) History of pancreatic/biliary surgery.

After an informed consent by the patients the selected

patients were subjected to:

Full clinical assessment (history taking and clinical

examination), laboratory investigations including (com-

plete blood count (CBC), Bilirubin (total and direct),

(ALT), (AST), alkaline phosphatase (ALP), Prothrom-

bin time and concentration (PT & PC), urea, creatinine,

serum amylase, serum lipase, urinary trypsinogen-2 dip-

stick test (UT2DST).

3. SPECIMEN COLLECTION

Two samples of 5 ml fasting venous blood were collec-

ted one before & the other 6 hours after ERCP for each

patient, the samples were left to clot and then centrifuged

at 3000 rpm for 5 minutes. The serum was then separated

& stored at −20˚C for measuring amylase and lipase.

Two Urine samples were taken one before & the other

6hours after ERCP for trypsinogen-2 determination by

UT2-DST.

3.1. Principle of UT2DST

The Actim© pancreatitis dipstick (MedixBiochemica, Kau-

niainen, Finland) strip, an immunochromatographic test,

was used for urine trypsinogen-2 determination (de-

tection limit 50 µg/L). The tip of the strip was im-

mersed into a urine-containing vial and was held for 20

seconds before being completely taken out of the vial.

The strip was then kept at room temperature for 5 mi-

nutes to observe whether urine reacted with blue latex

particles covered by monoclonal antitrypsinogen-2 anti-

bodies. Excess (>50 µg/L) urinary trypsinogen-2 caused

the occurrence of 2 blue stripes (one for trypsinogen-2 &

the other as control), while only one stripe (referred to as

the control stripe) was observed when urinary trypsino-

gen-2 concentration was within the normal range. The

appearance of the control stripe confirmed the accuracy

of the assay, while no blue stripes on the test strip sug-

gested an erroneous test, in which case the test should be

repeated.

Hyperamylasemia was defined as an increase of serum

amylase of greater than the upper limit of normal (82

U/L).

Hyperlipasemia was defined as an increase of serum

lipase of greater than the upper limit of normal (60 U/L).

Acute pancreatitis was diagnosed by abdominal pain

persistent for at least 24 hours, associated with serum

amylase and lipase equal to or greater than three times

the upper normal limit [4].

Abdominal ultrasonography was done for all patients

with special emphasis on the pancreas and biliary system

to establish the presence of biliary obstruction and to

diagnose its cause.

ERCP: The examination was done by using Olympus

duodenoscope video system Lucera CV 260SC or using

the Pentax lateral view endoscope ED-3440T and ED-

3485T.

The contrast material used was urograffin. The can-

nulation of the ductal system was done under X-Ray

screening to show the cause, the level and site of obstruc-

tion. Precut sphincterotomy was performed if cannula-

tion was difficult All patients were monitored at least for

six hours after the procedure to detect symptoms and

signs of complications (e.g. tachycardia, hypotension,

fever, vomiting and abdominal pain). Measurement of

serum amylase and lipase was done as mentioned by

sampling of blood at six hours post-ERCP. Urine sample

was taken 6 hours post ERCP for Trypsinogen-2 deter-

mination by UT2DST. Patients were then either hospi-

talized or followed up.

3.2. Statistical Analysis

Data were statistically described in terms of mean and

standard deviation (SD), frequencies (number of cases)

and percentages when appropriate. Comparison of quan-

titative variables between the study groups was done

using t-student test for parametric data or Mann Whit-

ney U test for non-parametric data in independent sam

ples. For comparing categorical data, Fischer’s exact test

was performed when appropriate. A probability value (P

value) less than 0.05 was considered statistically signi-

ficant. Kappa was used for the agreement. All statistical

calculations were done using SPSS (Statistical Package

for the Social Science; SPSS Inc., Chicago, IL, USA)

version 15 for Microsoft Windows.

4. RESULTS

The study included 37 patients referred to the gastroin-

testinal endoscopy unit, Kasr El Aini hospital to perform

ERCP. The age of the patients ranged between 22 and 77

years with a mean of 50 ± 1 years. Those patients were

23 males (62%) and 14 females (38%). From the 37 pa-

tients, 6 of them developed pancreatitis post ERCP

(16.2%) (3 males and 3 females) while 31 of them did

not (83.8%) as shown in Figure 1.

H. El-Garem et al. / Open Journal of Gastroenterology 3 (2013) 289-294 291

No significant differences were seen between the two

groups of patients (non pancreatitis group and pancreati-

tis group) as regard symptoms, signs, ultrasound and

ERCP findings. The laboratory results of the studied pa-

tients showed that there are no significant differences

between both groups of patients except for the lipase and

amylase serum levels after ERCP (post lipase and post

amylase) (P = 0.000) for both of them (Table 1).

The UT2DST before ERCP (pre_UT2DST) was nega-

tive in all studied patients.

non pancreatitis

pancreatitis

16.20%

83.80%

Figure 1. percentage of pancreatitis in the studied group.

Table 1. laboratory results of the 2 studied groups.

Non pancreatitis

N = 31

Pancreatitis

N = 6

Mean ± SD Mean ± SD

P value

Hb (gm/dl) 12.171 ± 2.29 11.717 ± 1.95 0.654

WBCx1000 6.2 ± 5 8.300 ± 4.5 0.32

Plateletsx1000 232.65 ± 132.92 326.80 ± 217.855 0.396

Bil.T (mg/dl) 8.080 ± 10.1 5.350 ± 20.6 0.92

Bil.D (mg/dl) 5.000 ± 6.5 3.500 ± 13.8 0.76

AST (U/L) 75.00 ± 56 62.00b ± 254 0.83

ALT (U/L) 56.00 ± 71 55.00 ± 293 1.0

GGT (U/L) 283.19 v ± 197.389 282.33 ± 135.848 0.992

ALK.Ph (U/L) 335.00 ± 260 325.0 ± 334 0.95

Urea (mg/dl) 31.52 ± 11.304 30.83 ± 15.171 0.899

Creatinine

(mg/dl) 0.892 ± 0.3032 1.017 ± 0.2137 0.345

Pre_lipase

(U/L) 27.9 ± 16.6 24.1 ± 5.3 0.591

Pre_amylase

(U/L) 67.1 ± 29.5 119.3 ± 65.3 0.109

Post_lipase

(U/L) 28.9 ± 14.7 260.5 ± 52.8 0.000

Post_amylsae

(U/L) 102.9 ± 165.8 1314.3 ± 379.8 0.000

Post ERCP UT2DST was negative in 30 patients of

the non pancreatitis group (96.8%) and positive in one of

them (3.2%) (Table 2).

The test was positive in all patients with Pancreatitis

(100%). The sensitivity of the post ERCP UT2DST was

100% the Specificity was 97% with PPV 86%, NPV

100% and the P value was <0.01.

Comparison between serum lipase and amylase levels

post ERCP in relation to UT2DST test shows that posi-

tive UT2DST test was significantly associated with

higher amylase and lipase serum levels after ERCP (post

amylase and post lipase) as shown in Table 3 (P < 0.01).

Post_UT2DST was positive when lipase >130 U/L

with sensitivity 86%, specificity 100%, AUC (Area un-

der curve) 0.914, P < 0.01. Post_UT2DST was positive

when amylase >122 U/L with sensitivity 100%, speci-

ficity 90%, AUC 0.981, P < 0.01 (Figur e 2).

Assessment of post serum lipase level at cutoff 130

U/L in relation to final diagnosis revealed that the sensi-

tivity, PPV, specificity and NPV all were 100% (Table

4).

Assessment of post serum amylase level at cutoff 122

U/L in relation to final diagnosis revealed that the Sensi-

tivity = 100%, PPV = 60%, specificity = 87% and NPV

= 100% (Table 5).

Agreement of post_UT2DST and post serum lipase

level at cutoff 130 U/L showed that the Kappa = 0.91 (P

< 0.01). While agreement of Post_UT2DST and post

serum amylase level at cutoff 122 U/L showed that the

Kappa 0.77 (P < 0.01).

5. DISCUSSION

This study was performed on 37 adult patients indicated

Table 2. Post ERCP UT2DST.

Non pancreatitis

N = 31

Pancreatitis

N = 6

Total

N = 37

NegCount30 0 30

Post

UT2DST PosCount1 6 7

P < 0.01.

Table 3. Comparison between serum lipase and amylase post

ERCP in relation to UT2DST.

Positive UT2DST

N = 7

Negative UT2DST

N = 30

Mean ± SD. Mean ± SD.

P value

Pre_lipase (U/L) 22.5 ± 6.4 28.4 ± 0.67 0.58

Pre_amylase (U/L)121.1 ± 59.8 64.9 ± 27.4 0.06

Post_lipase (U/L)226.3 ± 102.5 29.1 ± 14.9 <0.01

Post_amylsae (U/L)1144.1 ± 568.3 102.3 ± 168.6 <0.01

H. El-Garem et al. / Open Journal of Gastroenterology 3 (2013) 289-294

292

Source of the Curve

ost_Lipase

ost_am

lase

Reference Line

1.0

0.8

0.6

0.4

0.2

0.0

Sensitivity

0.0 0.2 0.4 0.6 0.8 1.0

1-Specificit

ROC Curve

Figure 2. ROC curve for serum amylase and lipase levels post

ERCP vs. results of post UT2DST test.

Table 4. Assessment of post serum lipase level at cutoff 130 in

relation to final diagnosis.

Non pancreatitis

n = 31

Pancreatitis

n = 6 Total

<130 U/L Count 31 0 31

Post_lipase

>130 U/L Count 0 6 6

Table 5. Assessment of post serum amylase level at cutoff 122

in relation to final diagnosis.

Non pancreatitis

(n = 31)

Pancreatitis

(n = 6) Tot al

<122 U/L Count 27 0 27

Post_amylase

>122 U/L Count 4 6 10

for ERCP, their ages ranged between 22 and 77 years

with a mean ± SD of 50 ± 1 years. Upon studying the

patients after ERCP, from the 37 patients 6 of them de-

veloped pancreatitis post ERCP (16.2%) while 31 of

them did not (83.8%).

The percentage of post ERCP pancreatitis in our study

agreed with other studies like Murray et al., (2003) [7]

which was 15.5%, Sankaralingam et al., (2007) [2]

which was16% and Kobayashi et al., 2011 [1] which was

12%.

Safwat et al., (2011) [8] performed a study in Egypt

and found that the overall incidence of post ERCP pan-

creatitis in their study was (14%), which agreed with the

percentage of post ERCP pancreatitis in our study.

The percentage of clinically detectable acute pancreas-

titis (AP) after ERCP ranges from 0 to 39% [9]. This

variability in the reported rates of post ERCP pancreatitis

is attributed to differences in criteria used for diagnosis,

differences in patient populations, endoscopic techniques

used, number of cases, aetiology and endoscopic experi-

ence [10].

The higher rates of pancreatitis in our study above the

rates reported in some studies may be attributed to more

strict parameters used for diagnosis in those studies. In

our study we used threefold elevation of amylase level as

a marker for diagnosis of acute pancreatitis. Other stud-

ies as that done by Testoni and Bagnolo, (2001) [11]

revealed that the frequency of post-ERCP/sphinctero-

tomy pancreatitis was between 1.3% and 7.6% and Free-

man et al., (2001) [12] found that the frequency of post-

ERCP pancreatitis that occurred was (6.7%), both of

them used higher levels of serum amylase (five folds

elevation) as a diagnostic marker for pancreatitis.

The Actim Pancreatitis MedixBiomedica urine test

strip measures concentrations of urinary trypsinogen-2

with a detection limit of the test of approximately 50

μg/L. It has been shown that the results from a urinary

trypsinogen-2 test strip agrees well with the rise in serum

trypsinogen-2 for a diagnosis of post-ERCP pancreatitis,

but the urine dip stick cannot be used to assess the sever-

ity of pancreatitis because it is not a quantitative meas-

urement [13].

In our study, 6 of the 37 patients developed post ERCP

pancreatitis (PEP), all of them had positive post ERCP

Urinary Trypsinogen-2-Dipstick test (UT2DST) with a

Sensitivity 100%, Specificity 97%, PPV 86%, NPV

100% and the P value was <0.01.

Kemppainen et al., (1997) [13] tested 106 patients un-

dergoing ERCP with a urinary trypsinogen-2 test strip 6

hours after ERCP, in whom 11 patients (10.4%) devel-

oped post-ERCP pancreatitis, they showed that the sensi-

tivity and specificity of urinary trypsinogen-2 strip test in

diagnosing post-ERCP pancreatitis were 81% and 97%,

respectively.

The high negative predictive value (98%) supported

the clinical value in excluding the development of post-

ERCP pancreatitis.

In another small-scale study, Sankaralingam et al.,

(2007) [2] tested 30 patients undergoing ERCP with uri-

nary trypsinogen-2 test strip at 1 and 4 hours after ERCP,

in whom 5 patients (17.2%) developed post-ERCP pan-

creatitis, they demonstrated that urinary trypsinogen-2

test had 100% sensitivity and negative predictive value at

1 and 4 hours after ERCP. The specificities at 1 and 4

hours after ERCP were 91% and 96%, respectively.

Tseng et al., (2011) [14] examined 150 patients under-

going ERCP with urinary trypsinogen-2 test strip before

and 3 hours after ERCP, in whom 13 (8.7%) patients

developed post-ERCP pancreatitis, urinary trypsinogen-2

strip test at 3 hours after ERCP had high sensitivity

(84.6%), specificity (97.1%), and negative predictive

value (98.5%) that are compatible also with the report of

Kemppainen et al., (1997) [13].

H. El-Garem et al. / Open Journal of Gastroenterology 3 (2013) 289-294 293

Another study reported that urinary trypsinogen-2 had

a 94% sensitivity for diagnosing acute pancreatitis [6],

Hedstrom et al., (1996) [15] also reported that the level

of trypsinogen-2 correlated with the severity of acute

pancreatitis.

Serum levels of amylase and lipase were examined at

6 hours after ERCP procedure in our study. We found

that the elevations of serum lipase have high negative

and positive predictive values which were both (100%),

the negative predictive value for the serum amylase was

(100%) but the positive predictive value was (60%)

which was lower than that of urinary trypsinogen-2 strip

test (86%), and there was a significant difference be-

tween levels of post ERCP amylase and lipase in both

groups.

Tseng et al., (2011) [14] examined the serum levels of

amylase and lipase at 3 hours after ERCP procedure and

found that the elevations of serum amylase or lipase (3 or

5 times normal) have high negative predictive values

(94.8% - 99.1%); however, their positive predictive va-

lues (36.4% - 42.9%) were markedly lower than that of

urinary trypsinogen-2 strip test (73.3%).

Our data showed a significantly higher mean level of

serum amylase 6 hours after ERCP in the pancreatitis

group (1314.3 ± 379.8 U/L) relative to the non pancreati-

tis group (102.9 ± 165.8 U/L) which agreed with the

Kobayashi et al., (2011) [1] study which showed that the

mean level of amylase 2 hours after ERCP in the pan-

creatitis group was (969.6 ± 220.4 U/L) relative to the

healthy group which was (120.4 ± 13.5 U/L).

However, in our study 4 of 31 cases of non-pancrea-

titis showed elevated serum levels of the amylase. In pre-

vious reports, post-ERCP levels of pancreatic enzymes in

cases with non pancreatitis peaked 6 hours after ERCP,

Kobayashi et al., 2011 [1] found that 14 of 60 cases of

non pancreatitis group showed greater than 3-fold eleva-

tion in amylase levels.

This may be due to the mechanics of the endoscopy

procedure which can cause enzymes to be reabsorbed

from the digestive tract via several mechanisms, salivary

gland stimulation associated with the insertion of a

mouthpiece or endoscope, insertion of a scope into the

gastrointestinal tract or introduction of air into the gas-

trointestinal tract [1].

Post-ERCP hyperamylasemia is reported by many au-

thors to be extremely common reaching up to 70% [16,

17].

The finding of post ERCP hyperamylasemia may be

attributed to other maneuvers used during ERCP as ma-

nipulation of the papilla during difficult cannulation,

pancreatic duct cannulation or injection, precut sphinc-

terotomy, balloon dilatation and extraction of large stones.

The mechanical trauma to the papilla or pancreatic

sphincter during instrumentation may cause transient

obstruction of outflow of pancreatic juice. Also, subject-

ing the pancreatic duct to a sudden increase in pressure

may be the cause of post ERCP hyperamylasemia. Pas-

sage of common bile duct stones is also known to cause

hyperamylasemia [18].

In conclusion, the urinary trypsinogen-2 dipstick test

can be used as an easy and rapid test for early diagnosis

of post-ERCP pancreatitis with high sensitivity and

specificity and can help clinicians to provide intensive

care and possible medical treatment as early as possible.

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