Review

Alkaline phosphatase:
beyond the liver
Nicole J. Fernandez, Beverly A. Kidney

Abstract: The alkaline phosphatases comprise a heterogeneous group of enzymes that are widely distributed in mammalian cells.
They often are associated with cell membranes, but their exact physiologic function is unknown. Despite this, alkaline
phosphatase activity is a very useful serum biochemical indicator of liver disease, particularly cholestatic disease. However,
increases in the activity of alkaline phosphatase in serum and other body fluids may reflect physiologic or pathologic changes
beyond those of hepatic origin. For example, nonhepatic increases in serum alkaline phosphatase activity are found in young
animals, in pregnant and lactating females, and in association with high fat diets. Bone disease, endocrine disease, neoplasia, and
other disorders can result in increased alkaline phosphatase activity. In addition, alkaline phosphatase activity may be increased
due to induction by certain drugs such as glucocorticoids and anticonvulsants. In this article, we will review the physiologic and
pathologic factors influencing the activity of alkaline phosphatase in serum and other body fluids, with an emphasis on disorders
beyond liver disease. (Vet Clin Pathol. 2007;36:223233)
2007 American Society for Veterinary Clinical Pathology

Key Words: Alkaline phosphatase, endocrine disease, enzyme induction, isoenzyme, isoform, review
u

Introduction
I.
II.
III.
III.
IV.

V.

VI.

VII.
VIII.
IX.
X.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Function and Distribution . . . . . . . . . . . . . . . . . . . .
Isoenzymes and Isoforms . . . . . . . . . . . . . . . . . . . .
Measurement of ALP in Biological Samples . . . . . .
Physiologic Effects on ALP . . . . . . . . . . . . . . . . . . .
A. Agerelated changes . . . . . . . . . . . . . . . . . . . . .
B. Pregnancy and lactation . . . . . . . . . . . . . . . . . .
C. Diet . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Induction by Glucocorticoids and
Anticonvulsants . . . . . . . . . . . . . . . . . . . . . . . . . .
A. Glucocorticoids and the steroid isoform . . . . . .
B. Anticonvulsants . . . . . . . . . . . . . . . . . . . . . . . . .
Disease Effects on Serum ALP . . . . . . . . . . . . . . . .
A. Hepatobiliary disease . . . . . . . . . . . . . . . . . . . .
B. Bone production and disease . . . . . . . . . . . . . . .
C. Endocrine disease . . . . . . . . . . . . . . . . . . . . . . .
D. Genetic and breed-related diseases . . . . . . . . . .
E. Neoplasia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
ALP in Other Fluids as a Marker of Disease . . . . .
Therapeutic Uses of ALP . . . . . . . . . . . . . . . . . . . .
Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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Alkaline phosphatase (ALP; EC 3.1.3.1) encompasses a group

of heterogeneous enzymes that catalyze the hydrolysis of
monophosphate esters at alkaline pH.1 Serum ALP activity is
used primarily as an indicator of hepatic disease; however,
there are numerous nonhepatic factors that result in an
increase in serum ALP activity. The ALP isoform profile in
serum is also influenced by many factors, including patient
age, drug therapy, endocrine disease, gestational stage, and
neoplasia. In this article, we will review the physiologic and
pathologic factors influencing serum ALP activity, with an
emphasis on disorders beyond liver disease. Use of ALP
measurement in other body fluids and tissues will also be
discussed. Unless otherwise noted, ALP activity refers to
serum activity of ALP.

Function and Distribution

ALP is active in a wide variety of organisms, from bacteria to
mammals, and thus has been described as having catalytic
promiscuity, a feature that may have contributed to its
evolutionary spread and ubiquitous nature.2 Although ALP is
one of the most studied enzymes, its physiologic function
has yet to be fully elucidated. In mammals, ALP is present
in serum in soluble form, but it also is found on many cellular membranes throughout the body.3 On the canalicular

From the Department of Veterinary Clinical and Diagnostic Sciences, Faculty of Veterinary Medicine, University of Calgary, and Animal Diagnostic Laboratory, CARE Centre, Calgary
Alberta Canada (Fernandez); and the Department of Veterinary Pathology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada
(Kidney). Corresponding author: Nicole J. Fernandez (fernandn@ucalgary.ca). 2007 American Society for Veterinary Clinical Pathology

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membranes of hepatocytes and the luminal surface of biliary

epithelial cells, ALP may be involved in choline transport
across cell membranes.4 In the brush border of epithelial cells
in the intestinal mucosa, it may be involved in the absorption
or hydrolysis of dietary phosphates.3 ALP is also found on the
cell membranes of osteoblasts and is necessary for bone
mineralization, but again, its exact function is unclear.3 In
semen, ALP is believed to be involved in sperm glycolytic
reactions and fructose formation.5 ALP is present on neuronal
membranes and at synaptic contacts in the primate cerebral
cortex, and a possible role for this neuronal isoform of ALP in
regulating neurotransmission has been proposed.6 ALP is also
found in granulocytes (neutrophils, eosinophils, and basophils) of many species, including humans, horses, and cattle,
but not in mature neutrophils of dogs and cats.7

Several methods are available for determining ALP

isoforms in domestic animals, including electrophoretic
mobility, stability to denaturation by heat or chemicals,
response to the presence of selected inhibitors, affinity for
specific lectins, and immunochemical characteristics.13 Results
of levamisole inhibition, heat inactivation, and affinity
electrophoresis have been shown to correlate strongly when
used to determine liver ALP and corticosteroid-induced ALP
isoforms in canine serum.14 Heat inactivation can be used to
differentiate between liver and bone ALP isoforms, but can be
imprecise because of difficulties in standardizing the many
variables of the assay, and may not identify other sources of
ALP.13,15 Modified electrophoretic techniques can be used to
reliably differentiate liver and bone ALP isoforms.13,15 Immunologic assays are the best method for measuring intestinal
and placental ALP isoforms.13 The reader is referred to other
sources for a more thorough discussion of methodology.1315

Isoenzymes and Isoforms

In humans, 4 gene loci encode 4 different ALP isoenzymes: 1)
a tissue-nonspecific isoenzyme, which is expressed in virtually
all tissues including liver, bone, and kidney; 2) an intestinal
isoenzyme, which is expressed in the brush border of the
intestinal mucosa; 3) a placental isoenzyme, which is expressed in trophoblasts during gestation and in small amounts
in the lung and cervix; and 4) a germ cell or placental-like
isoenzyme, which is expressed in small amounts in the testes
and thymus.3 These 4 genes appear to have evolved from
a single ancestral gene via multiple gene duplications, with
tissue-nonspecific and intestinal genes arising earliest, followed much later by the development of placental and germ
cell genes from the intestinal gene.3
In domestic mammals, only 2 gene loci encode ALP and
therefore 2 isoenzymes are produced, 1) a tissue-nonspecific
isoenzyme and 2) an intestinal isoenzyme.8 ALP from liver,
bone, kidney, and placenta is coded by the tissue-nonspecific
locus and is then modified, resulting in different isoforms of
ALP. Isoforms are produced by the post-translational modification of an isoenzyme (gene product); thus domestic animal
species have many isoforms of ALP but only 2 isoenzymes.
Seminal ALP, secreted by epithelial cells of the epididymis and
testes, is an isoform of tissue-nonspecific ALP that is unique to
the dog.9 The intestinal locus codes for intestinal ALP and, in
the dog, for a unique corticosteroid-induced isoform.1,10

Measurement of ALP in Biological Samples

ALP activity can be measured in any tissue, but for diagnostic
purposes it is typically measured in serum. In the healthy
adult dog, serum ALP activity is principally of hepatic origin,
with smaller contributions from bone and corticosteroidinduced isoforms.11 There is no measurable intestinal, renal, or
placental ALP in dog serum. The half-life of these isoforms is 3
to 6 minutes, in contrast to other ALP isoforms, which have
a half-life of approximately 70 hours in the dog. This difference
is caused by the presence of sialic acid residues on the liver,
bone, and corticosteroid-induced forms.1 In cats, the serum
half-life of ALP is shorter, at approximately 6 hours.12

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Physiologic Effects on ALP

Agerelated changes
Increased serum ALP activity has been consistently documented in neonates of many species. This phenomenon has
been further investigated in dogs and cats.16,17 In 13-day-old
Beagle and Beagle-type puppies, serum ALP activity was 30
times higher than adult values.16 This increase occurred within
24 hours of colostrum ingestion and did not occur in puppies
fed bitch milk-replacer. By day 10 after birth, serum ALP
activity was less than that measured before ingestion of
colostrum. During this time period, the bitches all had normal
serum ALP activity. Colostrum and milk collected from
bitches had 10 times the activity of bitch serum ALP until
day 10 after birth. The origin of the increased serum ALP
activity in the puppies was unclear; it may have been from the
colostrum or been released from alimentary viscera. An ALP
isoform profile was not determined in this study. In 12-dayold kittens, similar increases in serum ALP activity are seen
after ingestion of colostrum,17 and serum ALP can be used as
a marker of transfer of passive immunity in 12-day-old
kittens.18 By day 4, there is no significant difference in serum
ALP activity between colostrum-fed and colostrum-deprived
kittens.17 Colostrum intake does not result in increased serum
ALP activity in foals or calves.8,19
In a study of dogs 5 weeks to 2 years of age, serum
activity of the bone isoform of ALP (bone ALP) was highest
(approximately 4 times higher than adult values) in the
youngest dogs and then decreased dramatically in the first 3
months of life, reaching adult values by approximately 15
months of age.20 Various breeds were included in the study
and not evaluated separately in relation to ALP activity, so the
relationship between bone growth/size at maturity and
duration of increased bone ALP activity was not investigated.
In another study, total serum ALP activity was highest during
the first 16 weeks of life and no significant difference was
found between Beagles and Labrador Retrievers during this
time.21 In that study, serum ALP activity continued to decrease
until 1 year of age in both breeds; the median was slightly

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higher in Beagles compared to Labrador Retrievers after 16

weeks.
Total serum ALP activity was significantly higher in
kittens from birth to 8 weeks of age than in adult cats.17
Although ALP isoforms were not determined, the increase
was likely due at least in part to increased serum activity of
bone ALP, as serum from kittens ,15 weeks old has been
reported to contain only the bone isoform,22 and bone ALP
activity in cats less than 2 years old is significantly higher than
that of cats older than 2 years.23
Serum activity of bone ALP in newborn foals is 100 times
that of adults24 and accounts for 8092% of total serum ALP
activity8 as compared to 20% in horses .5 years old.25 Bone
ALP activity decreases in foals throughout the 23 weeks after
birth while serum activity of liver ALP remains constant.
In calves from birth to 83 days of age, total serum ALP
activity is increased compared to adult values.26 ALP activity
is highest at birth (up to 5 times adult values) and decreases
within 10 days to approximately 3 times adult values. ALP
isoforms were not determined, although in another study, the
presence of kidney ALP in neonatal calf serum was
demonstrated.27
The serum ALP isoform profile continues to change
throughout life. In a study of healthy dogs with no systemic
disease or history of steroid therapy, changes in the proportion
of bone, corticosteroid-induced, and liver ALP activity were
found with increasing age.28 In young dogs from 6 days to 1
year of age, bone ALP was the predominant form at 96% of
total serum ALP activity. In middle-aged dogs from 1 to 7
years old, the percentage of bone ALP decreased to 38%, and
in dogs .7 years of age it decreased further, to 26%. Bone ALP
thus constitutes the largest component of total serum ALP
activity in young dogs, but continues to make up a significant
portion in older dogs as well. Corticosteroid-induced ALP
(steroid ALP) was not significantly different between youngand middle-aged dogs (12 and 11% of serum ALP activity,
respectively), but rose to 27% in old dogs. This age-related
increase may be caused by increased stress and hypercortisolism. Interestingly, the authors noted that within the
young dog group, steroid ALP activity tended to be higher in
the youngest of these and then decreased in dogs approaching
1 year of age. No cause was determined, but they speculated
that stress might have resulted in the increase. Liver ALP
comprised ,10% of total serum ALP activity in young dogs.
In both middle-aged and old dogs, liver ALP accounted for
over 50% of the total ALP activity in serum. Thus liver ALP
constitutes the largest proportion of total serum ALP activity
in middle-aged and old dogs.
In cats, liver ALP is the primary contributor to total serum
ALP activity in healthy adults, while cats ,1 year of age have
a greater proportion of bone ALP.29

returns to within reference intervals within 20 days of

parturition.30
Increases in placental ALP activity have been detected in
the serum of pregnant queens and possibly in pregnant cows,
but not in pregnant bitches or mares.31,32 In queens, placental
ALP does not usually contribute to total ALP activity because
of its short half-life in serum (,2 minutes). However, in late
pregnancy, placental ALP activity has been detected in
serum.22 It is unclear whether the increased placental ALP
activity contributes to increased total serum ALP activity, or
whether total serum ALP activity remains within reference
intervals. In pregnant cows, an increase in total serum ALP
activity is observed in mid and late pregnancy.32 This increase
is likely caused by increased placental ALP activity, however
ALP isoforms have not been determined.
In the pregnant and postparturient bitch, a gradual
increase in activity of total serum ALP is seen from conception
to weaning, however, ALP activity remains within reference
intervals.33 Throughout gestation, the mean total serum ALP
activity increases from approximately 20 U/L to 30 U/L, an
increase that is too small to be clinically significant. In
pregnant mares, there is no significant change in total serum
ALP activity throughout gestation.30 In fact, a slight but
insignificant decrease in serum ALP activity was found in the
last months of pregnancy, which may be related to plasma
volume expansion. These differences between species could
possibly be explained by differences in placentation or
insufficient tissue concentrations in animals to raise serum
concentrations.30
Serum ALP activity is affected by stage of lactation as well
as by pregnancy. In lactating dairy cows, serum ALP activity
is increased because of increases in bone and liver ALP
activity.34 Serum ALP activity peaks in early lactation and then
gradually declines. ALP of mammary gland origin is present
in milk, but its contribution to serum ALP activity is unclear,
as it cannot be distinguished from the bone isoform using
established methods of isoform determination. Serum ALP
activity also shows seasonal variation in cows, independent of
reproductive status.32

Diet
In humans, increased intestinal ALP activity after eating may
lead to 5-fold increases in serum ALP activity, especially in
people consuming a high fat diet.35 Normally, ,10% of serum
ALP activity is from the intestine, but after eating fat-rich
meals, as much as 92% of ALP activity may be attributed to the
intestinal isoenzyme. In domestic animals, however, intestinal
ALP does not contribute to serum ALP activity. Horses on
a high fat diet do not have increased serum ALP activity
compared to horses on a high starch diet.36

Pregnancy and lactation

Induction by Glucocorticoids and Anticonvulsants

In pregnant human females, the activity of the placental

isoenzyme of ALP (placental ALP) is increased, resulting in
increased serum ALP activity. In the last trimester, this
increase may be up to 4 times normal. Serum ALP activity

Total serum ALP activity may be increased because of

a phenomenon known as enzyme induction.37 Certain drugs
such as glucocorticoids and anticonvulsants are thought to
increase the serum activity of ALP and other hepatic enzymes

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without causing hepatic dysfunction. Serum activity of both

liver and bone ALP may be induced. Dogs also have a unique
isoform of intestinal ALP called corticosteroid-induced ALP
(steroid ALP), although its serum activity increases in dogs
given anticonvulsants as well as steroids.38

Glucocorticoids and the steroid isoform

Increases in ALP activity can be detected in the liver, kidneys,
and intestinal tract as well as in the serum of dogs treated with
prednisone, but the greatest increases in tissue activity (of all
isoforms, including intestinal ALP) are seen in the liver.39 The
steroid ALP isoform is synthesized in the canine liver, but not
in the intestine or kidneys.40 Glucocorticoids are thought to
influence tissue ALP activity both at the level of gene
transcription and post-transcriptionally, depending on the
organ and ALP isoform involved.39
Total serum ALP activity is often increased in dogs with
increased cortisol levels because of hyperadrenocorticism or
iatrogenic hypercortisolism. In experimentally induced hypercortisolism, dogs given 4 mg/kg/d of prednisone IM for 10
days demonstrated a significant increase in total serum ALP
activity by day 3.41 This increase was due primarily to
increased serum activity of liver ALP. Liver ALP activity in
serum continued to increase throughout the 10 days of the
study, while hepatic bile acids remained within reference
intervals. This suggests that the increase in liver ALP activity
was not caused by cholestasis or induction by bile acids.
Steroid ALP activity in serum was not significantly increased
until day 7, and comprised only a small percentage of total
serum ALP activity (approximately 6%) by the final day of the
study, day 10. By day 10, bone ALP activity in serum was
significantly increased, and comprised a larger proportion of
total ALP activity than steroid ALP (approximately 10%). The
delay in the increase of steroid ALP activity in serum as
compared to liver ALP activity may reflect delayed expression
of the steroid ALP gene in liver tissue in response to
glucocorticoid treatment, although the mechanism responsible
for the delay is not clear.40
The percentage of total serum ALP activity attributable to
the steroid isoform is expected to increase over time, since
steroid ALP activity in dogs with naturally occurring hyperadrenocorticism is reported to account for 70100% of total
serum ALP activity.42-44 In dogs with a history of corticosteroid
therapy (duration and dosage uncertain) 60100% of total
serum ALP activity is reportedly caused by steroid ALP
activity.43,44 However, in a study of dogs given 4.4 mg/kg/d of
prednisolone for 30 days, only half of the dogs demonstrated
increased steroid ALP activity (comprising approximately
23% of the concurrent increase in total ALP activity) on day
30.28 Steroid ALP activity in the other half of the dogs was not
significantly different from control dogs or dogs receiving 1.1
mg/kg/d of prednisolone. This likely reflects individual
variation in response to corticosteroids, which has been
reported by others, for example Solter et al,45 although the
number of dogs in each group was low (6 dogs receiving 4.4
mg/kg prednisolone, 2 receiving 1.1 mg/kg).

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Serum ALP activity may also be increased in ill or stressed

dogs with increased levels of endogenous corticosteroids.
Although the relationship between stress and serum ALP
activity has not been elucidated, it is well established that
critical illness or exposure to stressful stimuli result in
increased basal cortisol levels and sometimes exaggerated
responses to ACTH stimulation testing in the dog.46,47 In
addition, the results of a study of 336 dogs with histologic
evidence of vacuolar hepatopathy suggest an association
between vacuolar hepatopathy, increased serum ALP activity,
and physiologic stress caused by acute or chronic illness.48
Forty-five percent of the dogs in that study had increased
serum ALP activity and no known history of corticosteroid
exposure or hyperadrenocorticism, although it was not clear if
any of these dogs had concurrent hepatobiliary disease. Data
on the proportion of total ALP activity caused by the steroid
isoform was not available.
Increases in total serum ALP activity persist after cessation
of corticosteroid therapy, but persistence depends on dose,
duration, and type of steroid. Ginel et al49 studied the effects
of several clinicallyrelevant corticosteroiddosing protocols.
Prednisone, at a dose of 1 mg/kg/d given orally for 3 weeks,
gave rise to increases that sometimes persisted for 3 weeks after
cessation of treatment, although serum ALP activity returned
to baseline levels in most dogs after 1 week. Dogs receiving
dexamethasone orally at a dose of 0.25 mg/kg/d for 1 week
had ALP activities within reference intervals by 3 weeks after
cessation of treatment. Increases caused by a single dose of
methylprednisone acetate at 1.1 mg/kg SC persisted as long as
5 weeks in some dogs, although significant increases persisted
for only 3 weeks. The authors suggested that a 3-week interval
for short-acting glucocorticoids and a 5-week interval for longacting glucocorticoids may be needed for serum ALP activity to
return to pretreatment levels, and that higher therapeutic doses
used for treatment of immune-mediated diseases may produce
increases of longer duration. In a series of dogs with iatrogenic
hypercortisolism caused by a variety of glucocorticoid treatment protocols, serum ALP activity was within reference
intervals in most dogs (24/28, 86%) at 6 weeks after withdrawal
of steroids,50 however, in a series of dogs receiving 4.4 mg/kg
prednisone IM for 14 days, serum ALP activity remained
increased at 8 times baseline values 6 weeks after cessation of
prednisone administration.51
Topical, ophthalmic, and otic preparations also induce
serum ALP activity and can result in clinical signs caused by
increased cortisol.50 In a study of 28 dogs diagnosed with
iatrogenic hyperadrenocorticism, 7 (25%) had received glucocorticoids only through topical, ophthalmic, or otic routes,
although it was not clear whether serum ALP activity was
increased in all of these dogs.50 Dogs given otic medications
containing corticosteroids demonstrated suppression of the
hypothalamic-pituitary-adrenal axis and increased serum
ALP activity that persisted for 2 weeks after cessation of
treatment.52,53 Dogs given ocular corticosteroids in another
study did not have increased serum ALP activity except
following hepatic biopsy. Biopsies showed changes consistent
with steroid hepatopathy.54
Clinicians often monitor liver enzymes in dogs on longterm steroid therapy, however, because of enzyme induction it

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can be difficult to differentiate increases caused by induction

from those caused by hepatic disease. In addition, corticosteroid therapy can result in steroid hepatopathy, a reversible
condition in which there is swelling of hepatocytes caused by
glycogen deposition.55 In general, impaired hepatic function is
uncommon in steroid hepatopathy, dogs rarely show signs of
hepatic disease or failure, and serum total bilirubin concentration will be normal, with serum bile acid concentrations
normal to mildly increased.56 Determination of ALP isoforms
may be of some use for noninvasive diagnosis of steroid
hepatopathy, although there is considerable overlap in isoform
profiles in dogs with hepatic disease and those with exposure
to corticosteroids.31 In dogs given 4.4 mg/kg/d prednisone for
14 days to induce steroid hepatopathy, serum ALP activity
was significantly increased by day 2 and continued to increase
for 6 days after corticosteroid administration was discontinued.51 This increase was due almost entirely to increased
steroid ALP activity, suggesting a correlation between the
steroid isoform and the development of steroid hepatopathy.
In another study, dogs receiving 4.4 mg/kg/d of prednisolone
for 30 days demonstrated only minor vacuolation of hepatocytes when increased serum ALP activity was due primarily
to increased liver ALP activity,28 but in dogs with similar
increases in serum ALP activity caused by both steroid and
liver ALP activity, severe hepatic vacuolation was seen
histologically. However, in both of these studies only 6 dogs
were used, and given the high dosages of glucocorticoids
used, the clinical relevance of the information is unclear.

Anticonvulsants
Serum activity of ALP may also be increased because of
enzyme induction caused by various anticonvulsants such as
phenobarbital, diphenylhydantoin and primidone.38,57 In dogs
receiving phenobarbital for treatment of epilepsy, total serum
ALP activity increased significantly within 3 weeks of
initiation of treatment.38 This increase was initially caused by
increased liver ALP activity. Steroid ALP activity was
increased at 6 months after initiation, and was the predominant isoform in 57.9% of dogs. The liver isoform
predominated in 36.8% of dogs, and 1 dog had approximately
equal amounts of each isoform. Bone ALP activity was also
increased at 6 months, but accounted for only a small
proportion of the total serum ALP activity.
It remains unclear whether these increases are caused by
enzyme induction, hepatotoxicity, or a combination of both.
Phenobarbital administration consistently results in increases
in serum ALP and ALT activity, but not in increased AST
activity and total bilirubin and bile acid concentrations, which
suggests a lack of hepatic disease.58 Gaskill et al59 reported that
enzyme induction is often inferred because dogs treated with
phenobarbital are clinically healthy despite increased ALP
activity. In order to separate enzyme induction from subclinical hepatotoxicity, they compared ALP activity in liver
homogenates to the activity of liver cytochrome P4502B, an
enzyme known to be induced by phenobarbital, in 5 dogs
receiving phenobarbital. ALP activity in liver homogenates
from treated dogs did not increase despite dramatic increases

Vol. 36 / No. 3 / 2007

in P4502B activity. Histopathologic examination of liver biopsies from 12 dogs demonstrated that phenobarbital-treated
dogs had similar but more severe changes than control dogs.
These results do not support enzyme induction as the
cause of increased serum ALP activity in dogs treated with
phenobarbital; however, other studies have found no histologic evidence of liver damage in dogs with increased serum
ALP activity following phenobarbital therapy. Muller et al58
described a diffuse increase in the size of hepatocytes and the
development of a ground glass appearance of the cytoplasm
after 10 weeks of phenobarbital therapy. The ground glass
appearance was attributed to increased smooth endoplasmic
reticulum secondary to microsomal enzyme induction, although electron microscopy would be needed for confirmation. Results of liver biopsies taken after 27 weeks of treatment
were similar and did not show evidence of progressive
changes caused by either enzyme induction or hepatic injury.
Dogs in this study also lacked radiographic and ultrasonographic evidence of liver damage.
Increases in serum ALP activity caused by phenobarbital
therapy persist for 35 weeks after cessation of therapy.60
Similarly, increases caused by diphenylhydantoin and primidone persist for 5 weeks.57

Disease Effects on Serum ALP

Hepatobiliary disease
Serum activity of ALP is proportional to tissue concentrations
and also depends on the half-life of the enzyme and its access
to the vasculature.31 Following insult to the liver, serum ALP
activity is increased because of de novo synthesis and elution
from hepatocyte and biliary epithelial membranes. In dogs
with naturally occurring hepatobiliary disease, the largest
increases in serum ALP activity (10-fold increase) are reported
in individuals with cholestasis.61 Chronic hepatitis, corticosteroid hepatopathy, and hepatic necrosis also result in large (6fold) increases in ALP activity. It should be noted that dogs
with experimentally induced hepatopathy following administration of exogenous glucocorticoids may have even larger
(up to 64-fold) increases in ALP activity.51 Hepatic neoplasia,
nodular regeneration, and hypoxia or hypotension may also
result in increased ALP activity.31 In addition, increased serum
ALP activity may be found in animals with hepatic lipidosis,
pancreatitis, or pancreatic neoplasia, likely secondary to
cholestasis.37 In humans, large (.10-fold) increases in ALP
activity are associated most commonly with intrahepatic
malignancies, bone metastases, and bacteremia.62
In 1 study, the sensitivity of serum ALP activity for
hepatobiliary disease in the dog was 7596%, however
specificity was only 51%.61 The low specificity of ALP activity
for hepatobiliary disease stems from the fact that serum ALP
activity can be increased by many nonhepatic causes. In cats
and horses, the diagnostic sensitivity of ALP activity in
detecting cholestasis is poor, and icterus often precedes an
increase in ALP activity.37 In cats, this is likely because the
magnitude of the increase in ALP is not as great because of the
short half-life of feline serum ALP and lower ALP activity per

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gram of liver tissue compared to the dog.63 In cattle, ALP

activity has moderate diagnostic sensitivity, however cholestatic disorders are uncommon.37 ALP activity is also of
limited usefulness in cattle because of its broad reference
interval; intervals are frequently determined without regard
for stage of lactation, which affects serum ALP activity in
dairy cows.34

Bone production and disease

The bone isoform of ALP is typically increased in young dogs
because of rapid bone growth, but increases may also be
observed in mature dogs with hyperparathyroidism, renal
disease, osteomalacia, osteomyelitis, and especially osteosarcoma.24,31 Diagnostically, suspicion of osteosarcoma (vs. other
types of sarcoma) is increased when ALP activity is detected
using cytochemical stains in fine needle aspirates of bony
lesions.64 Dogs with osteosarcoma often have increased serum
ALP activity as well, and measurement of serum ALP activity
in these dogs may have prognostic value. Dogs with osteosarcoma that have serum total and bone ALP activities
within reference intervals survive longer than dogs with
increased activities,65 and increased pretreatment serum total
and bone ALP activities are correlated with a shorter diseasefree interval after treatment.66,67
Dogs in which bone ALP activity in serum does not
decrease after amputation or limb-sparing surgery have
shorter survival and disease-free intervals than dogs in which
bone ALP activity decreases postsurgically.66 This study did
not include dogs with clinically detectable metastases at the
time of diagnosis, however, the influence of subclinical
metastasis on bone ALP activity in serum may be important.
In human patients that have undergone treatment for
osteosarcoma, an increase in total serum ALP activity may
be seen shortly before discovery of metastatic or recurring
lesions.68 Histologic grade, an important prognostic indicator
for canine osteosarcoma, appears to be independent of plasma
ALP activity,67 and although dogs with increased serum ALP
activity do not have higher grade or more undifferentiated
tumors,66 it has been suggested that dogs with high grade
tumors and increased serum ALP activity should be carefully
evaluated for metastatic disease.67 Serum ALP activity may
also be useful in formulating treatment plans following
surgery for osteosarcoma. Dogs with increased ALP activity
do not benefit from additional chemotherapy as compared to
dogs with normal ALP activity.67
ALP activity typically increases during fracture healing,
and measurement of serum ALP activity can be useful in
monitoring its progression.69 In dogs with uncomplicated
union of closed long bone diaphyseal fractures and in those
with delayed union, total serum ALP activity was increased at
time of presentation and continued to increase until 10 days
after surgery. This increase is presumably because of an
increase in activity of the bone isoform of ALP caused by
increased osteoblastic activity, but the authors measured only
total serum ALP activity. Subsequently, serum ALP activity
slowly declined, returning to within reference intervals when
bone union was complete. However, in dogs with poor

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fracture healing and nonunion, serum ALP activity was never

increased above reference intervals and did not show a similar
peak at 10 days postsurgery.
In horses and dogs, measurement of bone ALP activity in
serum is one of several methods used to assess bone in
a noninvasive way.25,70 In exercised horses, increased bone
ALP activity in serum reflects increased bone metabolic
activity and may aid in the early detection of bone disease
and in monitoring the effectiveness of treatment.25 In dogs,
serum bone ALP can be used as a marker of bone formation,
and the same reference intervals can be used for different
breeds, regardless of adult body size.70
Bone ALP activity can also be measured in synovial fluid,
and correlates with the degree of damage to articular cartilage
in horses with osteoarthritis.71 This finding suggests there is
a link between cartilage damage and bone turnover in
osteoarthritis and highlights the role of subchondral bone in
this disease. Interestingly, serum activity of bone ALP is not
useful as a marker of bone metabolic activity in dairy cows, as
ALP of osteoblastic origin is difficult to differentiate from ALP
originating from the mammary gland.72
In cats, bone ALP activity in serum has been investigated
as a potential marker of feline osteoclastic resorptive lesions,
a disease of undetermined etiology that commonly affects the
tooth root and alveolar bone of older cats, resulting in tooth
loss.23 Although this disease involves primarily bone resorption, the role of osteoblasts and bone formation is
unknown. No significant difference in bone ALP activity
was detected between affected cats and normal controls.
Incidentally, markers of bone resorption (deoxypyridinoline
and the carboxy-terminal telopeptide region of type I collagen)
also showed no significant difference between affected and
normal cats.

Endocrine disease
Serum ALP activity may be increased in canine endocrine
disorders such as hyperadrenocorticism, diabetes mellitus,
hypothyroidism, and hyperparathyroidism. This is perhaps
most intuitive in hyperadrenocorticism, a disease in which
there is increased endogenous corticosteroid. Increased total
serum ALP activity is the most common abnormality found in
routine blood work from dogs with hyperadrenocorticism73
and has been reported in 89.8% of dogs with spontaneously
occurring hyperadrenocorticism74 and 54% of dogs with
iatrogenic hypercortisolism.50 These increases are due at least
in part to increased steroid ALP activity, as previously
discussed.
Measurement of steroid ALP activity in serum has been
suggested as a diagnostic test for hyperadrenocorticism;
however, numerous researchers45,73,75,76 have concluded it is
not specific enough to be used as a diagnostic test, although it
may be useful as a screening test. Lack of steroid ALP activity
is likely a more useful test result, because it virtually rules out
hypercortisolism as a cause of increased serum ALP activity.76
Measurement of steroid ALP activity has not been found to be
useful for differentiating hyperadrenocorticism from iatrogenic hypercortisolism, liver disease, or diabetes mellitus.77

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Fernandez, Kidney

Although increased steroid ALP activity is reported to be 95%

sensitive for exposure to glucocorticoids, its specificity for
hyperadrenocorticism is only 18%.
Increased serum ALP activity is also a common finding
in canine diabetes mellitus, with 90% of diabetic dogs
reported to have increased total ALP activity.78 The reason
for the increase in ALP activity has not been conclusively
demonstrated, however it may be caused by hepatic lipidosis,
which frequently accompanies diabetes mellitus. Between 70
and 100% of dogs diagnosed with diabetes mellitus have
evidence of hepatic lipidosis at necropsy.78,79 Hyperadrenocorticism and pancreatitis are common concurrent disorders
in dogs with diabetes mellitus, and could also result in
increased serum ALP activity.78 Between 40 and 80% of the
total ALP activity in diabetic dogs is reportedly caused by the
steroid isoform,43 and in 1 study, steroid ALP was the only or
the predominant ALP isoform in diabetic dogs with increased
plasma ALP activity.44 The reason for this relatively high
proportion of steroid ALP activity is unclear, however 30% of
dogs with diabetes mellitus are reported to be lymphopenic,
likely reflecting stress, which could be associated with
increased endogenous corticosteroids and induction of
steroid ALP.31,78 Serum ALP activity is also significantly
increased in dogs with experimentally induced exocrine
pancreatic insufficiency.80 This is likely secondary to hepatic
lipidosis, which was found in all dogs in the study and was
considered multifactorial.
Hypothyroidism in dogs may be associated with increased serum ALP activity. In one study, 30% of hypothyroid
dogs had increased ALP activity, although most of these
increases were ,2 times the upper limit of normal, and some
of the dogs had concurrent disease (not specified but including
hepatic disease, hyperadrenocorticism, malignant neoplasia,
and cervical vertebral instability) or previous exposure to
glucocorticoids, which could explain the increase.81 The
mechanism for the increase is unknown, although in humans
with hypothyroidism, hepatic dysfunction has been implicated. In the dog, there is disagreement about whether the
increase is related to corticosteroid excess and enzyme
induction.73,81 Steroid ALP was reported to account for 20
80% of total serum ALP activity in hypothyroid dogs in 1
study,43 however the sample size was very small (n52). It was
unclear whether these dogs might have had other underlying
conditions that could have caused an increase in steroid ALP
activity.
Mild increases in serum ALP activity have been reported
in 50% (3/6) of dogs with primary parathyroid gland
hyperplasia.82 Although the specific isoforms present in these
dogs were not determined, increases were possibly caused by
alterations in bone metabolism and increased activity of bone
ALP, as has been previously reported.4 Increased serum ALP
activity is not a consistent finding in parathyroid hyperplasia,
however, and in cases of primary hyperparathyroidism and
renal and nutritional secondary hyperparathyroidism, ALP
activity may be within normal limits.4
In feline hyperthyroidism, serum ALP activity is increased in 7582% of cats.83 Although total serum ALP activity
may or may not be increased secondary to hyperthyroidism,
a correlation between total T4 concentration and liver ALP

Vol. 36 / No. 3 / 2007

activity has been reported.84 No correlation was found

between total T4 concentration and bone ALP activity,
however 88% of the cats had some bone ALP activity,
regardless of whether serum ALP activity was increased
above reference intervals. A study of hyperthyroid cats with
increased serum ALP activity found increased serum activity
of bone ALP in 100% of cats and increased liver ALP activity in
58% of cats.85 In the majority of these cats, .80% of total ALP
activity was attributable to bone ALP. Increases in serum
activity of bone ALP are likely caused by increased production
and release during active bone formation. T3 and T4 stimulate
both osteoclastic bone resorption and osteoblastic activity,
however, bone ALP is found primarily within osteoblasts.85
The cats in this study had no clinical signs such as osteoporosis
or pathologic fractures, which have been associated with
altered bone metabolism in hyperthyroid humans. Increased
liver ALP activity in hyperthyroidism has been attributed to
many factors, including direct toxic effects of thyroid
hormones on the liver and enzyme induction; however, liver
dysfunction is rare in both cats and humans with hyperthyroidism.85 Histologic changes in hepatic biopsies from
hyperthyroid cats are generally mild and nonspecific. In
another study, an additional ALP isoform from an unidentified tissue source was found in the serum of hyperthyroid cats
with increased ALP activity.86 The unidentified ALP isoform
was also increased in mature cats with increased serum ALP
activity but normal serum T4 concentrations.

Genetic and breed-related diseases

There are several reports of increased serum ALP activity
within specific related groups of dogs and of humans. Benign
familial hyperphosphatasemia has been recognized in related
litters of Siberian husky puppies.87 Eleven- and 16-week-old
puppies had markedly increased total serum ALP activity
(mean .5 times that of unaffected siblings and other unaffected Siberian huskies of the same age) without evidence of
disease. In 5 pups for which the isoform profile was
determined, the increase was attributable to an increase in
the activity of the bone isoform. Benign inherited hyperphosphatasemia has also been reported in several human
families.88 The increase in serum ALP activity results from
increased activity of bone and liver ALP in some families and
intestinal ALP in others. Recently, increased serum ALP
activity of undetermined cause has been reported in Scottish
terriers, however, the mechanism for the increase is unknown,
and isoform data is not available.89,90 Some but not all of these
dogs were known to be related.
In humans, serum ALP activity varies with ABO blood
group.91 Different blood groups have more or less intestinal
ALP in serum depending on the degree of erythrocyte
binding. Erythrocytes of blood group A bind avidly to
intestinal ALP, thus very little serum ALP activity is of
intestinal origin in people of blood group A. Erythrocytes of
blood groups B and O bind intestinal ALP to a much lesser
degree, thus individuals with these blood groups have more
intestinal ALP in their serum. The effect of this binding on total

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Alkaline Phosphatase Review

ALP activity has not been determined, although normally,

intestinal ALP accounts for ,10% of serum ALP activity.35
Decreased serum ALP activity has diagnostic value in
humans, namely for aiding the diagnosis of hypophosphatasia. Hypophosphatasia is a rare, autosomal recessive
inherited condition characterized by extremely low activity
of tissue-nonspecific ALP caused by a mutation in the gene
coding for this isoenzyme.92 This deficiency results in marked
demineralization of bone and can be fatal in neonates and
infants. The diagnosis is confirmed by demonstrating increased levels of inorganic pyrophosphate in the urine.
Disease conditions resulting from low ALP activity have not
been conclusively demonstrated in animals, however, the
discovery of a new type of urolith in 4 Persian cats and 1 Collie
dog in Sweden has prompted some speculation.93 The uroliths
are composed of potassium, magnesium, and pyrophosphate,
and the authors suggest they may develop in animals with
low activity of hydrolyzing ALP, similar to humans with
hypophosphatasia. Low ALP activity might result in supersaturation of urine with pyrophosphate and a predisposition
to develop crystalluria and urolithiasis. However, a genetic
basis for the disease was not supported as the 4 affected
Persian cats were not closely related, and a nutritional cause
for the uroliths could not be ruled out. The name perserite
has been introduced for these crystals, reflecting their
occurrence in Persian cats.

Neoplasia
ALP has received considerable interest regarding its use in
cancer diagnosis. In humans, several ALP isoforms are
associated with malignancy and may be useful in the
diagnosis of certain tumors.1 One is a variant of placental
ALP called Regan whose serum activity increases in malignancies of the lung, gastrointestinal tract, ovary, and uterus.
There is also a variant of germ cell or placental-like ALP called
Nagao, which is found in seminomas and other tumors and
a third, called Kasahara that corresponds to intestinal ALP and
has been identified in certain hepatomas.3 These isoforms
appear to be present in only some patients with these tumors,
therefore diagnostic sensitivity is not very high.
ALP has been studied in the diagnosis of mammary
neoplasia in the dog, but researchers found no significant
difference in serum ALP activity in dogs with malignant
tumors compared to those with benign tumors.94 The increases
in ALP activity in these dogs also did not correlate with the
presence of bone in the tumor or the histologic type of tumor.
A subsequent study found that compared with tumor-free
control dogs, serum activities of total ALP, bone ALP, liver
ALP, and steroid ALP were significantly higher in dogs with
malignant mammary tumors.95 No significant differences
were found in ALP isoform activities between dogs with
malignant epithelial mammary tumors and malignant mixed
mammary tumors containing bone.
Similarly, steroid ALP activity in serum has been investigated as a potential prognostic indicator in canine
lymphoma, however, no correlation between response rate
or duration of remission and steroid ALP activity was found.96

Page 230

The use of serum ALP activity as a prognostic indicator for

osteosarcoma in dogs was discussed above.

ALP Activity in Other Fluids as a Marker of Disease

Measurement of ALP activity in body fluids other than serum
has been investigated, particularly in pleural, peritoneal, and
reproductive tract fluids, and urine. As previously discussed,
ALP activity in joint fluid may reflect damage to articular
cartilage, and the presence or absence of ALP activity in cells
and tissues can help to distinguish osteosarcoma from other
types of sarcomas. Cytochemical staining for ALP activity in
leukocytes can be useful in identifying the lineage of poorly
differentiated cells, for example blast cells in leukemia.7 In
humans with neutrophilic leukocytosis, leukocyte ALP
activity can help differentiate a leukemoid response from
neoplasia. 7
In humans, measurement of fluid ALP activity may be
useful as a biochemical marker for differentiating pleural
transudates from exudates,97 although it remains to be seen
what advantage this conveys over traditional methods such as
cell counts, protein measurement, and cytologic examination.
In a study measuring enzyme activities in canine bronchoalveolar lavage fluid, ALP activity was significantly increased in
dogs with bronchopneumonia, but not in dogs with tracheobronchitis.98 The reason for this increase was unclear and ALP
isoforms were not determined, however, it was speculated
that the increase was caused by a pulmonary isoform of tissuenonspecific ALP, since this isoform is found in bronchoalveolar lavage fluid from humans with pulmonary disease.99
Several researchers have investigated increases in the
activity of peritoneal fluid ALP as a potential marker of small
intestinal degeneration or necrosis in naturally occurring and
experimentally induced equine colic.100103 This information
would be valuable in determining treatment of colic cases and
deciding whether surgical intervention is necessary. In 2
studies,100,102 serum or plasma ALP activity remained within
reference intervals while peritoneal fluid ALP activity increased. However, 12% of horses with naturally occurring
colic had severe small intestinal lesions without increases in
peritoneal fluid ALP activity,100 and in the experimental study,
results were similar between control and experimental
ponies.102 Thus, determination of peritoneal fluid ALP activity
is likely not a reliable means of predicting small intestinal
injury. However, it may be useful in predicting the severity of
intestinal insult and prognosis. Saulez et al103 reported that in
horses with naturally occurring colic, the highest peritoneal
fluid ALP activities were found with medical peritonitis and
with surgical lesions, including ulceration, strangulation,
rupture, and peritonitis. Horses with high peritoneal fluid
ALP activity had more severe intestinal lesions and increased
mortality compared to horses with lower ALP activity.
Interestingly, the ALP isoform responsible for the increased peritoneal fluid activity differed between the
studies. In the experimental study,102 ALP was of intestinal
origin, as expected, however, in the study of naturally occurring colic,100 ALP activity was primarily of granulocytic origin.
Granulocyte ALP is released from disintegrated granulocytes

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Fernandez, Kidney

and its activity correlates only roughly with total nucleated

cell counts from peritoneal fluid.100 It was suggested that this
difference occurred because there had not been time for
granulocyte accumulation in the experimental model,102 or
because intestinal ALP activity in peritoneal fluid increases
only after full thickness intestinal injury.100 The isoform results
from Saulez et al103 cannot be compared to the findings from
these studies.104
ALP activity in seminal fluid can be measured as part of
a breeding soundness evaluation. ALP is produced in the
epididymis in the dog, and can be used as a marker for patency
of the ductal system.105,106 In an aspermic or oligospermic dog,
decreased seminal fluid ALP activity suggests ductal blockage
rather than testicular failure, as sperm are not a major source of
ALP activity. This is also true of cats, pigs, and horses.5,107
Normal semen ALP activity ranges from 5000 to 40,000 U/L
in the dog, so sample dilution is required for laboratory
evaluation. In addition, complete semen collection is needed to
ensure the accuracy of this test. ALP is also present in low
concentrations in the canine testis and may have some
potential as a marker of germ cell function and fertility
status.106
In cows, measurement of ALP activity in cervical mucus
may be useful as a marker of normal mucus, and thus of
potential fertility.108 The composition of cervical mucus during
estrus is important in ensuring passage of sperm, since failure
of fertilization in cattle most often results from failure of sperm
to reach the ova.
In humans, atypical isoforms of ALP are reported in the
urine and serum of patients with chronic renal failure.109 These
isoforms are derived from both intestinal and tissuenonspecific ALP, and appear as creatinine clearance declines.
They may be useful markers of renal disease.

1. Syakalima M, Takiguchi M, Yasuda J, Hashimoto A. The canine

alkaline phosphatases: a review of the isoenzymes in serum, analytical
methods and their diagnostic application. Jpn J Vet Res. 1998;46:311.
2. OBrien PJ, Herschlag D. Functional interrelationships in the alkaline
phosphatase superfamily: phosphodiesterase activity of Escherichia coli
alkaline phosphatase. Biochemistry. 2001;40:56915699.
3. Harris H. The human alkaline phosphatases: what we know and dont.
Clin Chim Acta. 1989;186:133150.
4. Milne EM. The diagnostic value of alkaline phosphatase in canine
medicine: a review. J Sm An Pract. 1985;26:267278.
5. Turner RMO, McDonnell SM. Alkaline phosphatase in stallion semen:
characterization and clinical applications. Theriogenology. 2003;60:110.
6. Fonta C, Negyessy L, Renaud L, Barone P. Area and subcellular
localization of the ubiquitous alkaline phosphatase in the primate
cerebral cortex: evidence for a role in neurotransmission. Cerebral
Cortex. 2004;14:595609.
7. Raskin RE, Valenciano A. Cytochemistry of normal leukocytes. In:
Feldman BF, Zinkl JG, Jain NC, eds. Schalms Veterinary Hematology. 5th
ed. Philadelphia, PA: LippincottWilliams & Wilkins; 2000:337346.
8. Hank AM, Hoffmann WE, Sanecki RK, Schaeffer DJ, Dorner JL.
Quantitative determination of equine alkaline phosphatase isoenzymes in foal and adult serum. J Vet Intern Med. 1993;7:2024.
9. Kutzler MA, Solter PF, Hoffman WE, Volkmann DH. Characterization
and localization of alkaline phosphatase in canine seminal plasma and
gonadal tissues. Theriogenology. 2003;60:299306.
10. Saini PK, Peavy GM, Hauser DE, Saini SK. Diagnostic evaluation of
canine serum alkaline phosphatase by immunochemical means and
interpretation of results. Am J Vet Res. 1978;39:15141518.
11. Saini PK, Saini SK. Origin of serum alkaline phosphatase in the dog.
Am J Vet Res. 1978;39:15101513.
12. Hoffman WE, Renegar WE, Dorner JL. Serum half-life of intravenously injected intestinal and hepatic alkaline phosphatase isoenzymes in the cat. Am J Vet Res. 1977;38:16371639.
13. Panteghini M, Bais R, van Solinge WW. Enzymes. In: Burtis CA,
Ashwood ER, Burns DE, eds. Tietz Textbook of Clinical Chemistry and
Molecular Diagnostics. 4th ed. St Louis, MO: Elsevier Saunders;
2006:597644.
14. Mahaffey EA, Lago MP. Comparison of techniques for quantifying
alkaline phosphatase isoenzymes in canine serum. Vet Clin Path.
1991;20:5155.

Therapeutic Uses of ALP

ALP shows some potential as a novel therapeutic agent.
Beumer et al110 demonstrated that ALP derived from calf
intestine was able to detoxify lipopolysaccharide and enable
mice to survive lethal doses of Escherichia coli. Antibodies
directed against specific isoenzymes of ALP may also be
useful in targeting tumor cells that display these antigens111
and in treating inflammatory conditions such as inflammatory
bowel disease.112

15. Moore WE, Feldman BF. The use of isoenzymes in small animal
medicine. J Am Anim Hosp Assoc. 1974;10:420429.
16. Center SA, Randolph JF, ManWarren T, Slater M. Effect of colostrum
ingestion on gamma-glutamyltransferase and alkaline phosphatase
activities in neonatal pups. Am J Vet Res. 1991;52:499504.
17. Levy JK, Crawford PC, Werner LL. Effect of age on reference intervals
of serum biochemical values in kittens. J Am Vet Med Assoc. 2006;228:
10331037.
18. Crawford PC, Levy JK, Werner LL. Evaluation of surrogate markers
for passive transfer of immunity in kittens. J Am Vet Med Assoc.
2006;228:10381041.
19. Boyd JW. Serum enzyme changes in newborn calves fed colostrum.
Vet Clin Pathol. 1989;18:4751.

Conclusions
Although ALP is used primarily as a diagnostic indicator of
hepatobiliary disease, it is widespread in many tissues and
fluids throughout the body and thus has broad diagnostic
potential. Many opportunities exist for further research into
the use of ALP as a diagnostic enzyme and its multiple
isoenzymes and isoforms, especially in species other than dogs
and humans. Despite the extensive research that has been
conducted on ALP, there are still areas that have not been
thoroughly investigated, including the physiologic function
of ALP.

Vol. 36 / No. 3 / 2007

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