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Correlation of salivary and nasal lavage pepsin with MII‐pH testing

Article in The Laryngoscope · July 2019

DOI: 10.1002/lary.28182

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Correlation of Salivary and Nasal Lavage Pepsin With MII-pH

Testing

Miles J. Klimara, MD ; Nikki Johnston, PhD; Tina L. Samuels, MS ; Alexis M. Visotcky, MS;
David M. Poetker, MD; Todd A. Loehrl, MD; Joel H. Blumin, MD ; Jonathan M. Bock, MD

Objectives: Laryngopharyngeal reflux (LPR) is a common upper airway disease. Salivary pepsin is a proposed marker for
LPR; however, the optimal time for collection of specimens for pepsin detection and pepsin’s presence in the oral and nasal
secretions relative to concurrent multichannel intraluminal impedance-pH (MII-pH) monitoring are unknown.
Study Design: Prospective case-control study with an experimental design.
Methods: Patients undergoing MII-pH testing for evaluation of LPR and asymptomatic control subjects were selected.
Nasal lavage and saliva samples were collected in the clinic prior to MII-pH probe placement. Additional saliva samples were
obtained an hour after each meal and upon waking the following morning. Nasal lavage and salivary pepsin were measured by
ELISA.
Results: Twenty-six patients undergoing MII-pH testing and 13 reflux-free control patients were enrolled. Salivary pepsin
was detected in 11 of 26 patients with suspected LPR and 0 of 13 controls. Pepsin was most frequently detected in the speci-
men provided upon waking at an average concentration of 186.9 ng/mL. A significant correlation was observed between sali-
vary pepsin in waking samples to MII-pH measurements, including reflux bolus duration, and proximal and distal recumbent
reflux episodes (P < 0.05). A significant correlation was also observed between salivary pepsin upon waking or sinus lavage
and reflux symptom index (P < 0.05).
Conclusion: Pepsin in salivary and nasal lavage samples demonstrated an association with MII-pH-documented LPR. Pep-
sin detection was most frequent in morning samples, supporting use of morning salivary pepsin levels as a potential noninva-
sive technique for LPR diagnosis.
Key Words: Laryngopharyngeal reflux, pepsin, saliva, multichannel intraluminal impedance-pH monitoring.
Level of Evidence: 2
Laryngoscope, 00:1–6, 2019

INTRODUCTION the reflux symptom index (RSI) and reflux finding score
Laryngopharyngeal reflux (LPR) is defined as reflux (RFS) have demonstrated poor correlation with objective
of gastric contents into the larynx and pharynx and pre- measures of reflux events.3 Multichannel intraluminal
sents with a wide variety of laryngeal symptoms such as impedance-pH probe monitoring is widely regarded as the
globus sensation, dysphonia, and dysphagia. LPR has tra- most reliable alternative but remains relatively more inva-
ditionally been diagnosed via clinical history combined sive, costly, and time-consuming. Therefore, there is sub-
with laryngoscopic findings such as pseudosulcus vocalis, stantial interest in alternative methods for evaluation of
ventricular obliteration, posterior commissure hypertro- suspected LPR.
phy, and thick mucosal secretions.1,2 However, tools for Pepsin is a digestive enzyme that is implicated in the
historical and laryngoscopic identification of LPR such as mucosal inflammation underlying the pathophysiology of
LPR.4,5 Because pepsin is exclusively found in gastric
From the Department of Otolaryngology and Communication secretions, its presence in airway and salivary secretions
Sciences (D.M.P., T.A.L., J.H.B., J.M.B.); the Department of Microbiology and has been suggested as a less expensive and invasive, as
Immunology (M.J.K., N.J.); and the Institute for Health and Equity, well as a more sensitive and specific, marker for reflux-
Division of Biostatistics (T.L.S., A.M.V.), Medical College of Wisconsin,
Milwaukee, Wisconsin, U.S.A. related airway processes.5,6 Indeed, a sensitive immuno-
Additional supporting information may be found in the online assay for pepsin previously demonstrated up to 100%
version of this article. sensitivity and 89% specificity for reflux as identified by
Editor’s Note: This Manuscript was accepted for publication on
June 24, 2019. pH monitoring.7 Since the introduction of MII-pH monitor-
Presented at the 2019 Triological Sections Combined Sections Meet- ing, more recent work has identified the presence of pepsin
ing, Coronado, California, U.S.A., January 24–26, 2019. in the saliva of many patients with impedance-confirmed
This study was supported by the Medical College of Wisconsin
Department of Otolaryngology and Communication Sciences. J.M.B. is a proximal reflux, suggesting a potential role for pepsin in
paid consultant for Diversatek Healthcare. The authors have no other the diagnosis of LPR.8,9 Proximal reflux is also suggested
funding, financial relationships, or conflicts of interest to disclose.
Send correspondence to Jonathan M. Bock, MD, Department of Oto-
as a contributor to rhinologic disease such as chronic
laryngology and Communication Sciences, Medical College of Wisconsin, rhinosinusitis (CRS), a notion bolstered by literature iden-
8701 Watertown Plank Road, Milwaukee, WI 53226. E-mail: tifying a high prevalence of reflux in patients with poor
[email protected]
outcomes following endoscopic sinus surgery for CRS.10,11
DOI: 10.1002/lary.28182 Pepsin in nasal lavage has been studied as a potential

Laryngoscope 00: 2019 Klimara et al.: Salivary Pepsin and MII-pH in LPR
1
marker for extraesophageal reflux (EER) in CRS, and a impedance sensor array placed at or just above the upper esopha-
study by Ozmen et al. demonstrated pepsin in nasal geal sphincter under visualization with flexible laryngoscopy,
lavage of 82% of patients undergoing endoscopic sinus sur- according to our previously described protocol.14 Prior to placement
of the probe, the nose was prepared with topically administered
gery for CRS.12
atomized 4% lidocaine mixed 50:50 with 0.05% oxymetazoline.
However, there is no established protocol for the col-
Patients were instructed to carefully register upright and supine
lection of salivary specimens for detection of pepsin as a periods, mealtime, and symptoms (cough, dysphonia, globus sen-
marker of proximal reflux. In a study of gastroesophageal sation, or throat clearing) for symptom correlation. MII-pH data
reflux disease (GERD) patients, Hayat et al. demon- was uploaded to the manufacturer’s analysis software and inter-
strated that salivary pepsin was significantly more likely preted by the physician as described in our previously described
to be detected in specimens collected in the postprandial protocol.14 Tests were generally interpreted as positive for
period.13 More recently, however, Na et al. studied a pop- LPR with any significant pharyngeal acid episodes (defined as
ulation of patients with proximal reflux confirmed with pH < 4.0) or > 40 proximal impedance events based on prior nor-
MII-pH monitoring and found that salivary pepsin levels mative data.15
were significantly higher upon awakening than after
meals or when symptoms occurred.9 Therefore, the opti-
mal timing for collection of salivary pepsin for the pur- Nasal Lavage and Salivary Specimen Collection
pose of diagnosing EER remains uncertain. Further, in Prior to placement of MII-pH probe, nasal lavage samples
the aforementioned studies, healthy controls exhibited were obtained via Lukens trap after instilling 5 to 10 cc of saline
positive salivary pepsin.9,13 The utility of salivary pepsin to one nostril with a small syringe. Over the course of the
in establishing a diagnosis of LPR is therefore unclear. 24 hours testing period, subjects were instructed to expectorate
their saliva into a tube at 1 hour after lunch, 1 hour after dinner,
Moreover, there is little data regarding the prevalence of
when waking and standing in the morning, and 1 hour after eat-
reflux to the level of the nasopharynx and pepsin’s pres-
ing breakfast. Patients were instructed to refrigerate the sam-
ence in nasal lavage specimens in patients with LPR. ples and bring them to their visit for probe removal the following
Therefore, the major goal of the study herein was to day. Samples were stored at −80 C until analysis was completed.
examine the presence and level of salivary pepsin at vari-
ous time points, as well as the presence of pepsin in nasal
secretions, with respect to the presence of proximal reflux
Pepsin ELISA and Western Blot
as determined by traditional methods of LPR diagnosis. Pepsin ELISA was performed on all saliva and lavage spec-
We hypothesize that MII-pH reflux event data will corre- imens according to a previously described protocol.16 Specimens
late with salivary and nasal lavage pepsin detected. with an A450 greater than two-fold the absorbance of a sample
diluent blank were considered positive for pepsin for the pur-
poses of analysis. A representative subgroup of the collected
MATERIALS AND METHODS specimens was analyzed with sodium dodecyl sulfate - polyacryl-
amide gel electrophoresis according to a previously established
Subjects and Study Design protocol for qualitative corroboration of the ELISA data.16
This prospective case series was approved by the institu-
tional review board of the Medical College of Wisconsin
(Milwaukee, WI). Patients with suspected LPR, including post-
nasal drip and CRS, were offered dual pH-MII testing by one of Statistical Analysis
the two senior authors (J.H.B. and J.M.B.) after laryngology clinic Continuous measures were summarized using mean/
evaluation. All patients were offered testing as part of their ini- standard deviation and compared by Wilcoxon Mann-Whitney
tial clinical evaluation if they were suspected of having LPR, and test. Categorical measures were assessed by Fisher exact test.
patients were included if they agreed to testing. Healthy controls Spearman rank correlation coefficient was estimated to evalu-
>18 years of age with no diagnosis or symptoms of reflux or use ate the association between salivary pepsin; nasal lavage pepsin;
of anti-reflux medications were offered enrollment via print clinical data including RSI and RFS; and MII-pH parameters
advertisements at the Medical College of Wisconsin and including longest bolus duration, total distal and recumbent reflux
Froedtert Hospital, Milwaukee, WI and in scheduled visits to the episodes, and proximal and distal acid reflux episodes. Patients
Froedtert Hospital otolaryngology clinic. Patients undergoing with incomplete data were excluded from analysis with respect to
evaluation for suspected LPR were not compensated. Controls any missing variables. The level of significance was considered to
were provided with a stipend via a $400 gift card for their partic- be two-sided P value <0.05.
ipation in the study to compensate for travel time and impedance
testing that is not the standard of care and has the possibility of
slight discomfort. RSI questionnaires were completed for each RESULTS
subject prior to study. Flexible laryngoscopy was performed on
Twenty-six patients clinically suspected to have LPR
all patients as part of MII-pH placement, and RFS was obtained.
and 14 normal controls were enrolled, submitted salivary
All patients were assessed after abstaining from proton pump
inhibitor therapy for 1 week, H2 blockers for 48 hours, and and nasal lavage specimens, and underwent MII-pH mon-
CaCO3 the day of the study. itoring. Of the patients with suspected LPR, 14 (39%)
were on some form of acid suppressive therapy prior to
enrollment. One recruited control was excluded from
24-Hour MII-pH Monitoring analysis due to impedance events associated with a sali-
Subsequently, MII-pH studies were completed with the vary specimen positive for pepsin, suggesting a diagnosis
Diversatek ComforTEC LPR dual pH-MII testing apparatus of LPR. All completed studies had waveforms deemed
(Diversatek Healthcare, Milwaukee, WI), with the proximal pH and adequate for interpretation. Demographic data and

Laryngoscope 00: 2019 Klimara et al.: Salivary Pepsin and MII-pH in LPR
2
 TABLE I. TABLE III.
Demographics of Patients and Healthy Controls. Characteristics and ELISA Results of Patients With Suspected LPR.
Patients, n = 26 Controls, n = 13 Morning Salivary Nasal Lavage
RSI RFS Pepsin (ng/mL) Pepsin (ng/mL)
Gender, n (%)
MII-pH positive for LPR
Male 9 (35) 4 (31)
23 9 149 38730
Female 17 (65) 9 (69)
24 9 57 0
Age, mean 49 34
27 10 0 44460
A statistically significant difference was detected in mean age between 29 7 356 1322
the patient and control groups. However, age was not significant when
adding age as a predictor to linear regression models. 31 9 0 0
7 13 0 0
13 10 0 0
ELISA results are summarized in Tables I and II, respec-
tively. Of MII-pH studies in patients with suspected LPR, 18 8 0 0

19 of 26 (73%) were interpreted as positive for evidence of 23 7 0 0

significant LPR (Tables III and IV), whereas four of 19 5 – 37010
13 (31%) of studies were interpreted as positive in 21 5 – 61480
healthy controls. Pepsin was detected in at least one spec- 29 5 2528 0
imen submitted by 11 of 26 (42%) patients with clinically 31 0 0 0
suspected LPR, in eight of 19 (42%) patients with proxi- 32 7 0 0
mal reflux confirmed by MII-pH monitoring, and in three 10 8 0 0
of seven (43%) patients with negative MII-pH studies
13 8 0 0
(Tables II–III, and IV). Pepsin was detected in nasal
20 5 0 0
lavage of seven of 26 (26.9%) patients with clinically
26 5 465 0
suspected LPR, and in five of 19 (26.1%) patients with
LPR confirmed with positive impedance studies 30 8 0 0

(Tables II–III, and IV). Pepsin was detected in the nasal MII-pH negative for LPR
lavage of two of seven patients with negative MII-pH 34 7 69 1224
studies, both of whom additionally provided salivary spec- 26 3 824 14994
imens positive for pepsin (Tables III and IV). No pepsin 31 3 0 0
was detected in saliva or nasal lavage of healthy controls. 14 5 0 0
19 3 38 0
13 10 0 0
TABLE II. – – 0 0
Comparison of Patients and Healthy Controls.
RSI, RFS, and pepsin ELISA results of all patients with clinically
Patients, n = 26 Controls, n = 13 suspected LPR. Missing values represented with –.
LPR = laryngopharyngeal reflux; RFS = reflux finding score; RSI = reflux
Pepsin positive, any specimen, 11 (42%) 0 symptom index.
n (%)**
Pepsin positive, morning, n (%)* 8 (33%) 0
Pepsin positive, after breakfast, 1 (4%) 0 Pepsin was most frequently detected in specimens
n (%) collected the morning after waking. The mean salivary
Pepsin positive, after lunch, n (%) 1 (4%) 0 pepsin level in patients with suspected LPR in specimens
Pepsin positive, after dinner, n (%) 3 (12%) 0
Pepsin positive, probe placement, 0 0
n (%)
TABLE IV.
Pepsin positive, nasal lavage, 7 (27%) 0 Comparison of Pepsin ELISA Results in Suspected LPR Patients
n (%) With Positive and Negative MII-pH Studies.
Pepsin, morning salivary, ng/mL 186.9 (−39.4 to 413.2) 0
(95% CI)* MII-pH Positive MII-pH Negative

Pepsin, nasal lavage, ng/mL 7662 (747 to 14577) 0

Pepsin, any specimen positive, n (%) 8 of 19 (42%) 3 of 7 (43%)
(95% CI)
Pepsin, morning saliva positive, n (%) 5 of 17 (29%) 3 of 7 (43%)
RFS (95% CI) 6.8 (5.6 to 8.0) 4.9 (2.9 to 7.0)
Missing 2 0
RSI (95% CI)*** 22.5 (19.3 to 25.7) 2.4 (0.7 to 4.2)
Pepsin, nasal lavage positive, n (%) 5 of 19 (26%) 2 of 7 (29%)
Results of pepsin ELISA, RFS, and RSI in patients and healthy con- Morning saliva, ng/mL 209.1 133
trols. Fisher exact test compared categorical data and Wilcoxon
Mann-Whitney tests compared continuous data. Nasal lavage, ng/mL 9631.7 2316.9
*P ≤ 0.05.
**P ≤ 0.01 Comparison of pepsin ELISA results in patients with clinically
***P ≤ 0.001. suspected LPR, categorized by final MII-pH study interpretation.
CI = confidence interval; RFS = reflux finding score; RSI = reflux symp- LPR = laryngopharyngeal reflux; MII-pH = multichannel intraluminal
tom index. impedance-pH.

Laryngoscope 00: 2019 Klimara et al.: Salivary Pepsin and MII-pH in LPR
3
Fig. 1. Mean pepsin in nasal lavage at probe placement and in salivary lavage specimens of patients upon waking, postprandially, and at
probe placement. Error bar represents one standard deviation. LPR = laryngopharyngeal reflux.

collected the morning after waking was 186.9 ng/mL; and There is no agreed-upon cutoff for salivary pepsin as
mean pepsin in specimens collected in the clinic following a diagnostic marker in LPR, likely due to considerable
probe placement, after breakfast, lunch, and dinner were variability in the levels of salivary pepsin reported in var-
0 ng/mL, 0.3 ng/mL, 0.1 ng/mL, and 12.3 ng/mL, respec- ious studies of LPR and GERD.17 The levels of pepsin
tively (Fig. 1). Significant correlation was observed between detected in the study herein are roughly in line with some
salivary pepsin upon waking and the longest bolus dura- previous investigations of salivary pepsin levels in GERD
tion (R = 0.357, P = 0.0301), proximal recumbent acid epi- and LPR.13,18 However, this is markedly higher than is
sodes (R = 0.479, P = 0.0027), proximal recumbent episodes seen in other studies of LPR, including a well-controlled
of any pH (R = 0.391, P = 0.0166), distal recumbent acid study by Na et al. that similarly measured pepsin in saliva
episodes (R = 0.402, P = 0.0137), distal recumbent episodes at several time points throughout the day, which showed a
of any pH (R = 0.334, P = 0.043), and RSI (R = 0.446, mean salivary pepsin level of 17.2 ng/mL.9,19 Less than
P = 0.0073) (Table V). Mean nasal lavage pepsin in patients half of the patients with proximal reflux detected by MII-
with suspected LPR was 7662 ng/mL, and the levels of pep- pH testing in the study herein exhibited pepsin in saliva,
sin in nasal lavage significantly correlated with RSI and the presence of pepsin did not predict positive MII-pH
(R = 0.362, P = 0.0275) (Tables II and V). No significant cor- testing. In contrast, Hayat et al. has previously utilized
relation was observed between the RFS and salivary pep-
sin upon waking (R = −0.094, P = 0.5987) or nasal lavage
pepsin (R = 0.053, P = 0.7573) (Table V).
TABLE V.
Correlation of Salivary and Nasal Lavage Pepsin With Clinical Data.

DISCUSSION Salivary Pepsin, Nasal Lavage

Morning (R) Pepsin (R)
Previous literature has demonstrated an association
of salivary pepsin and proximal reflux events identified by Longest bolus duration 0.357* 0.201
impedance monitoring.8 However, the precise relationship Proximal recumbent episodes, total 0.391* 0.166
between salivary pepsin and MII-pH data is unclear, and Proximal recumbent episodes, acid 0.479** 0.089
a recent systematic review has pointed to the substantial Distal recumbent episodes, total 0.334* 0.171
variability in salivary pepsin levels throughout the day as
Distal recumbent episodes, acid 0.402* 0.159
a major challenge for its use for diagnosis of LPR.17 The
DeMeester score 0.165 −0.026
primary aim of this study was therefore to assess the rela-
RSI 0.446** 0.362*
tionship between MII-pH parameters and pepsin in a pro-
spective fashion. A secondary goal of this study was to RFS −0.094 0.053
determine the optimal time for saliva collection for the Spearman correlation of salivary and nasal lavage pepsin with selected
measurement of pepsin as a marker for LPR, by correla- clinical and MII-pH data.
tion of pepsin with MII-pH testing, as well as to investi- *P ≤ 0.05.
**P ≤ 0.01.
gate the relationship between nasal lavage pepsin and MII-pH = multichannel intraluminal impedance-pH; R = Spearman cor-
evidence of reflux obtained with MII-pH testing. relation coefficient, RSI = reflux symptom index; RFS = reflux finding score.

Laryngoscope 00: 2019 Klimara et al.: Salivary Pepsin and MII-pH in LPR
4
the commercially available Peptest (RD Biomed, Hull, comparisons to this technique cannot be drawn. As
East Yorkshire, United Kingdom) to detect pepsin in the described above, there is also no gold standard for the inter-
saliva of 80% of patients with proximal reflux identified by pretation of impedance data and the diagnosis of LPR.2
MII-pH testing and only 36% of patients without identifi- Basic parameters of MII-pH data are prone to false positive
able proximal reflux.8 Such discrepancies may be due to and false negative events due to artifacts caused by, for
heterogeneity in the methods used for collection of speci- instance, swallows mimicking reflux events.22,23 Indeed, in
mens, techniques used to measure pepsin levels, and the study at hand, four of 13 (31%) of healthy controls had
MII-pH testing. However, because the levels of pepsin in MII-pH studies consistent with LPR. Although pepsin’s
specimens provided upon waking did correlate significantly absence in the saliva and nasal lavage of all 13 controls sup-
with a number of impedance parameters in the study ports its potential as a tool for identifying true cases of LPR,
herein, pepsin in saliva does appear to provide corrobora- further study is clearly necessary to determine the signifi-
tive evidence of the presence of proximal reflux. False nega- cance and appropriate interpretation of discordant results
tives with respect to MII-pH testing may occur due to the between pepsin and MII-pH testing. Of particular interest,
inherently intermittent nature of laryngopharyngeal reflux only eight of 23 (35%) of subjects with positive MII-pH tests
and hence variability in pepsin in saliva. Given the absence had detectable pepsin in saliva, although it should be noted
of pepsin specimens obtained from healthy controls, it that this includes several healthy controls with positive
might be possible to mitigate this pitfall by repeated sam- MII-pH testing, all of whom had negative tests for pepsin.
pling or testing of pooled salivary specimens. The detection The study herein should therefore only be considered sup-
of pepsin most frequently in samples collected the morning portive of further investigation of pepsin assays for the non-
upon waking corroborates the work of Na et al., contrary to invasive diagnosis of LPR and should not be misinterpreted
previous reports that salivary pepsin is most likely to be as demonstrating the utility of pepsin assays as a compo-
detected postprandially.9,13 Future studies aiming to fur- nent of routine clinical care. Finally, the study population is
ther delineate the role of salivary pepsin as a diagnostic small, and the subject groups differed significantly in terms
marker should consider repeated or pooled morning saliva of age, which may limit generalizability of these findings.
collection, and comparative studies of common methods of However, when adding age as a predictor to linear regres-
pepsin measurement might serve to clarify the underlying sion models, age was not significant.
cause for the differences seen between studies of salivary Morning salivary pepsin on waking correlates with
levels of pepsin in reflux-related disease. both reflux symptoms and with objective evidence of reflux
The levels of salivary pepsin on waking correlated on MII-pH monitoring, although not all patients with a
with both MII-pH data and RSI scores but were not sig- positive MII-pH study had pepsin in the waking saliva
nificantly correlated with RFS. This corroborates prior sample. Future studies may be able to utilize repeated col-
data, which suggests the RFS has a poor correlation with lection of morning saliva specimens to reduce the fre-
objective evidence of reflux events and has only modest quency of false negatives and hence increase the
specificity for the diagnosis of LPR.14,20 In conjunction diagnostic utility of salivary pepsin, potentially allowing
with a randomized, blinded study of the RFS in which its use as a screening tool prior to MII-pH testing. Nasal
Branski et al. demonstrated the measure’s poor interrater lavage pepsin may be a useful adjunctive tool in suspected
reliability, this data raises concern regarding the use of LPR and CRS, but because normative data for nasal
the RFS as a diagnostic tool.21 lavage pepsin concentration in patients with and without
Although pepsin was only detected in nasal lavage LPR is not available, its utility cannot be determined.
specimens of a minority of patients with LPR in this study, Future studies of pepsin in waking salivary samples and
all of these patients exhibited either pepsin in their saliva nasal lavage with a larger patient population may reveal
or evidence of proximal reflux on MII-pH monitoring. This their value in the diagnostic evaluation of suspected LPR.
is consistent with prior data correlating the presence of
reflux with CRS.10,12 Because pepsin in nasal lavage is evi-
dence of reflux to the level of the nasopharynx, it is inter- CONCLUSION
esting that patients with positive nasal lavage specimens Pepsin is present in saliva and nasal lavage of
did not uniformly exhibit both positive salivary pepsin and patients with LPR, and salivary pepsin on waking was
MII-pH testing. This may be due to inherent differences in significantly higher in a group of patients with LPR
the rates and timing with which pepsin can be detected in symptoms and evidence of proximal reflux on MII-pH
nasal lavage and saliva. Alternatively, obtaining nasal testing than in healthy controls. Salivary pepsin on wak-
lavage may release pepsin, which is present in the nose or ing correlated with both reflux symptoms and with objec-
sinuses as a result of prior episodes of reflux, and patients tive evidence of reflux on MII-pH monitoring, and pepsin
might thereafter simply not exhibit reflux during the was not detected in the saliva or nasal lavage of asymp-
period of MII-pH monitoring. Lastly, it must be considered tomatic controls without MII-pH evidence of LPR. These
that current techniques for measuring and interpreting findings suggest that salivary pepsin upon waking may
impedance data might be insufficient for the detection of be a useful diagnostic tool in the evaluation of LPR.
some cases of low-volume reflux.
This study has several clear limitations. First, mea-
surements of pepsin in saliva during episodes of patient- Acknowledgment
identified symptoms is one method of specimen collection The authors would like to thank Sergey Tarima, PhD, of
described in the literature that was not performed, and thus the Medical College of Wisconsin (MCW) Division of

Laryngoscope 00: 2019 Klimara et al.: Salivary Pepsin and MII-pH in LPR
5
Biostatistics for assistance with the data analysis and symptoms with endoscopic examination findings and potential prognostic
variables. Laryngoscope 1997;107:504–510.
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14. Cumpston EC, Blumin JH, Bock JM. Dual pH with multichannel
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