Objective—To develop and validate a radioimmunoassay
(RIA) for measuring canine pancreatic
lipase immunoreactivity (cPLI) in serum obtained
Sample Population—Serum samples from 47
Procedures—Canine pancreatic lipase (cPL) was purified
from pancreatic specimens of dogs. Antibodies
against cPL were raised in rabbits and purified by use
of affinity chromatography. A tracer was produced by
iodination of cPL with 125I. An RIA was established and
validated by determination of sensitivity, working
range, dilutional parallelism, spiking recovery, and
intra- and interassay variability. A reference range for
cPLI in serum was established by use of the central
95th percentile for samples obtained from 47 healthy
Results—Sensitivity and upper limit of the working
range were 0.88 and 863 µg/L, respectively.
Observed-to-expected ratios for serial dilutions
ranged from 84.9 to 116.5% for 4 samples. Observedto-
expected ratios for spiking recovery ranged from
82.8 to 128.6% for 4 samples. Coefficients of variation
for intra-assay variability for 4 serum samples
were 18.3, 4.2, 3.5, and 8.9%, whereas interassay
coefficients of variation were 29.2, 6.2, 3.9, and
4.4%, respectively. The reference range was 4.4 to
Conclusions and Clinical Relevance—We conclude
that the RIA described is sensitive, linear, accurate,
precise, and reproducible, with limited accuracy in the
high end of the working range and limited precision
and reproducibility in the low end of the working
range. Additional studies are needed to evaluate
whether this degree of accuracy, precision, and reproducibility
will negatively impact clinical use of this
assay. (Am J Vet Res 2003;64:1237–1241)
Objective—To describe the kinetics of urinary recovery
(UR) of 5 sugars used for gastrointestinal permeability
and mucosal function testing following orogastric
administration of lactose, rhamnose, xylose,
methylglucose, and sucrose.
Animals—7 healthy male Beagles.
Procedure—A sugar solution containing lactulose,
rhamnose, xylose, methylglucose, and sucrose was
administered by orogastric intubation to healthy dogs.
Urine samples were collected immediately before
sugar solution administration (baseline) and at 2-hour
intervals thereafter. The UR of the 5 sugars was determined
from urine concentrations measured by high
pressure liquid chromatography and pulsed amperometric
detection. Percent urinary recovery (%UR) of the
total UR up to 12 hours after sugar solution administration
was calculated for each sugar at 2-hour intervals.
Results—Mean %UR exceeded 85% for all 5 sugars
at 6 hours after orogastric administration of the sugar
solution and exceeded 90% after 8 hours.
Conclusion and Clinical Relevance—In healthy
dogs, a urine collection period of 6 hours is sufficient
for gastrointestinal permeability and mucosal function
testing following orogastric administration of lactulose,
rhamnose, xylose, methylglucose, and sucrose.
(Am J Vet Res 2002;63:845–848)
Objective—To validate an automated chemiluminescent
immunoassay for measuring serum cobalamin
concentration in cats, to establish and validate gas
chromatography-mass spectrometry techniques for
use in quantification of methylmalonic acid, homocysteine,
cysteine, cystathionine, and methionine in sera
from cats, and to investigate serum concentrations of
methylmalonic acid, methionine, homocysteine, cystathionine,
and cysteine as indicators of biochemical
abnormalities accompanying severe cobalamin (vitamin
B12) deficiency in cats.
Sample Population—Serum samples of 40 cats with
severe cobalamin deficiency (serum cobalamin concentration
< 100 ng/L) and 24 control cats with serum
cobalamin concentration within the reference range.
Procedure—Serum concentrations of cobalamin
were measured, using a commercial automated
chemiluminescent immunoassay. Serum concentrations
of methylmalonic acid, methionine, homocysteine,
cystathionine, and cysteine were measured,
using gas chromatography-mass spectrometry,
selected ion monitoring, stable-isotope dilution
Results—Cats with cobalamin deficiency had significant
increases in mean serum concentrations of
methylmalonic acid (9,607 nmol/L), compared with
healthy cats (448 nmol/L). Affected cats also had substantial
disturbances in amino acid metabolism, compared
with healthy cats, with significantly increased
serum concentrations of methionine (133.8 vs 101.1
µmol/L) and significantly decreased serum concentrations
of cystathionine (449.6 vs 573.2 nmol/L) and
cysteine (142.3 vs 163.9 µmol/L). There was not a significant
difference in serum concentrations of homocysteine
between the 2 groups.
Conclusions and Clinical Relevance—Cats with
gastrointestinal tract disease may have abnormalities
in amino acid metabolism consistent with cobalamin
deficiency. Parenteral administration of cobalamin
may be necessary to correct these biochemical
abnormalities. (Am J Vet Res 2001;62:1852–1858)
Objective—To determine serum lipase activities and pancreatic lipase immunoreactivity (PLI) concentrations in dogs with exocrine pancreatic insufficiency (EPI).
Animals—74 healthy dogs and 25 dogs with EPI.
Procedures—A diagnosis of EPI was made on the basis of clinical signs, low serum trypsin like immunoreactivity (TLI) concentration, and response to treatment with enzyme replacement. Median values for fasting serum lipase activity and serum PLI concentrations were compared between the 2 groups with a Mann-Whitney U test.
Results—Median fasting serum lipase activity was not significantly different between dogs with EPI (366.0 U/L) and healthy dogs (294.5 U/L), and only 1 dog with EPI had a serum lipase activity less than the lower limit of the reference range. Median serum PLI concentration was significantly lower in dogs with EPI (0.1 μg/L) than in healthy dogs (16.3 μg/L). All dogs with EPI had serum PLI concentrations less than the lower limit of the reference range.
Conclusion and Clinical Relevance—Serum lipase activity is not limited to the exocrine pancreas in origin, whereas serum PLI is derived only from the exocrine pancreas. Unlike in serum TLI concentrations, there was a small degree of overlap in serum PLI concentrations between healthy dogs and dogs with EPI. Serum TLI concentration remains the test of choice for diagnosis of EPI.
Objective—To determine cellular immunolocalization
of canine gastric lipase (cGL) and canine pancreatic
lipase (cPL) in various tissues obtained from clinically
Sample Population—Samples of 38 tissues collected
from 2 climically healthy dogs.
Procedures—The cGL and cPL were purified from gastric
and pancreatic tissue, respectively, obtained from
dogs. Antisera against both proteins were developed,
using rabbits, and polyclonal antibodies were purified by
use of affinity chromatography. Various tissues were collected
from 2 healthy dogs. Primary antibodies were
used to evaluate histologic specificity. Replicate sections
from the collected tissues were immunolabeled for cGL
and cPL and examined by use of light microscopy.
Results—Mucous neck cells and mucous pit cells
of gastric glands had positive labeling for cGL,
whereas other tissues did not immunoreact with
cGL. Pancreatic acinar cells had positive labeling for
cPL, whereas other tissues did not immunoreact
Conclusions and Clinical Relevance—We concluded
that cGL and cPL are exclusively expressed in gastric
glands and pancreatic acinar cells, respectively. Also,
evidence for cross-immunoreactivity with other lipases
or related proteins expressed by other tissues was not
found for either protein. Analysis of these data suggests
that gastric lipase is a specific marker for gastric
glands and that pancreatic lipase is a specific marker
for pancreatic acinar cells. These markers may have
clinical use in the diagnosis of gastric and exocrine
pancreatic disorders, respectively. (Am J Vet Res
Objective—To develop and validate an ELISA for
quantitative analysis of feline trypsin-like immunoreactivity
Sample Population—Purified feline cationic trypsin
(fCT) and rabbit anti-fCT antiserum; blood samples
from 63 healthy cats.
Procedures—A sandwich capture ELISA was developed,
using anti-fCT antiserum purified by affinity
chromatography that underwent biotinylation.
Purified fCT was used for standards. The assay was
validated by determination of sensitivity, working
range, linearity, accuracy, precision, and reproducibility.
A reference range was established by assaying
serum samples from the 63 healthy cats.
Results—Sensitivity was 1.23 µg/L; working range
was 2 to 567 µg/L. Ratios of observed versus
expected results for 4 samples tested at various
dilutions ranged from 90.0 to 120.7%. Ratios of
observed versus expected results for 5 samples
spiked with various concentrations of fCT ranged
from 82.0 to 101.8%. Intra- and inter-assay coefficients
of variability ranged from 9.9 to 11.1% and
from 10.2 to 21.7%, respectively. The reference
range for serum fTLI measured with this ELISA was
12 to 82 µg/L.
Conclusions and Clinical Relevance—Results suggest
that an ELISA can be used to measure serum
fTLI in cats. The ELISA was sufficiently sensitive, linear,
accurate, precise, and reproducible for clinical
use. (Am J Vet Res 2000;61:620–623)
Objective—To investigate postprandial changes in
serum concentrations of unconjugated bile acids in
Animals—7 healthy Beagles.
Procedure—Blood samples were obtained from dogs
at regular intervals up to 8 hours after consumption of
a meal. Serum concentrations of 5 unconjugated bile
acids were determined at each time point, using gas
chromatography-mass spectrometry with selected
Results—Total serum unconjugated bile acid concentration
was significantly increased, relative to baseline
values, at 360, 420, and 480 minutes after feeding.
Unconjugated cholic acid was significantly increased
at 360, 420, and 480 minutes. The proportion of total
unconjugated bile acids represented by cholic acid
was significantly increased at 240 to 480 minutes.
Deoxycholic acid was significantly increased at 360
and 420 minutes. Chenodeoxycholic acid was significantly
increased at 360 to 480 minutes. Lithocholic
acid was significantly increased at 180 minutes,
whereas no significant changes in ursodeoxycholic
acid were detected at any time point.
Conclusion and Clinical Relevance—Healthy
Beagles had significant increases in serum concentrations
and changes in the profile of unconjugated bile
acids after a meal. These increases persisted > 8 hours,
indicating that prolonged withholding of food is necessary
when to avoid the risk of a false-positive diagnosis
when assessing serum unconjugated bile acid concentrations
in dogs. (Am J Vet Res 2002;63:789–793
Objective—To describe the kinetics of demethylation
of 13C-aminopyrine in healthy dogs for use in determining
the most appropriate time for collection of
blood samples for a 13C-aminopyrine demethylation
blood test for evaluation of hepatic function.
Animals—9 healthy dogs.
Procedures—A 2-mL baseline blood sample was collected
into an evacuated heparinized tube, and 13Caminopyrine
was administered to each dog (2 mg/kg,
IV). Additional 2-mL blood samples were collected 15,
30, 45, 60, 75, 90, 105, 120, 135, 150, 180, 240, 300,
and 360 minutes after 13C-aminopyrine administration.
The CO2 was extracted from blood samples by addition
of a strong acid, and the percentage dose of
13CO2 (PCD) in the extracted gas was determined by
fractional mass spectrometry.
Results—No dogs had gross evidence of adverse
effects, and all had an increase in PCD after IV administration
of 13C-aminopyrine. The PCD had the least
variability among 5 variables used to evaluate hepatic
demethylating capacity. Peak PCD was detected at 30
minutes in 1 dog, 45 minutes in 5 dogs, 60 minutes
in 2 dogs, and 75 minutes in 1 dog. The mean PCD for
the 9 dogs peaked at 45 minutes after 13C-aminopyrine
Conclusions and Clinical Relevance—PCD appears
to be the preferable variable for evaluation of hepatic
demethylating capacity. Intravenous administration of
13C-aminopyrine leads to a consistent increase in PCD.
Mean PCD peaked 45 minutes after administration,
suggesting that blood sample collection 45 minutes
after 13C-aminopyrine administration may be appropriate
for use in estimating hepatic demethylating
capacity. (Am J Vet Res 2004;65:159–162)
Objective—To develop and validate a gas chromatography–mass spectrometry (GC-MS) method for determination of Nτ-methylhistamine (NMH) concentration in canine urine and fecal extracts and to assess urinary NMH concentrations in dogs with mast cell neoplasia and fecal NMH concentrations in dogs with protein-losing enteropathy.
Sample Population—Urine specimens were collected from 6 healthy dogs and 7 dogs with mast cell neoplasia. Fecal extracts were obtained from fecal specimens of 28 dogs with various severities of protein-losing enteropathy, as indicated by fecal concentration of α1-proteinase inhibitor.
Procedures—NMH was extracted directly from urine, and fecal specimens were first extracted into 5 volumes of PBSS containing 1% newborn calf serum. Nτ-methylhistamine in specimens was quantified via stable isotope dilution GC-MS. The assay was validated via determination of percentage recovery of known amounts of NMH and interassay coefficients of variation. Urinary excretion of NMH was evaluated by means of NMH-to-creatinine concentration ratios.
Results—Recovery of NMH in urine and fecal extracts averaged 104.6% and 104.5%, respectively. Interassay coefficients of variation ranged from 5.4% to 11.7% in urine and 12.6% to 18.1% in fecal extracts. Urinary NMH excretion was significantly increased in dogs with mast cell neoplasia, compared with that in healthy dogs. No correlation was detected between severity of protein-losing enteropathy and fecal NMH concentration.
Conclusions and Clinical Relevance—This method provided a sensitive, reproducible means of measuring NMH in canine urine and fecal extracts. High urinary NMH-to-creatinine concentration ratios in dogs with mast cell neoplasia are consistent with increased histamine release in this disease.