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CE U P D A T E -

James K. Klaassen, DVM, DACVP, PhD


VETERINARY MEDICINE III

Reference Values in
Veterinary Medicine

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| In veterinary medicine, as in human Reference values for immunologic and sero-
medicine, reference values are necessary for useful logic assays require use of species-specific reagents
interpretation of laboratory results. Without appropriate and kits. For example, anticanine immunoglobu-
lin is required to correctly perform a Coombs' test
reference values, the sensitivity and specificity of tests are
on a canine sample. (See DiUlio1 for a more in-
lowered, which can result in erroneous diagnoses and depth discussion of veterinary immunoassays.)
treatment protocols that may cause harm to or death of the
animal tested. Reference values specific for the species tested D e t e r m i n i n g Reference Values
and the equipment and reagents used are essential for "Reference values" is the preferred term, as recom-
accurate interpretation. Many veterinary reference mended by the International Federation of Clini-
laboratories use historic or published values rather than cal Chemistry, but most practitioners understand
the concepts of normal ranges, reference ranges,
establishing their own, because of the time and expense
and normal values. Some would argue that refer-
involved. In addition, limited availability of sufficient ence "range" is incorrect because in statistical jar-
numbers of many species (40-120 or more animals) makes gon, range is a single value that is the difference
establishment of laboratory-specific reference values between the highest and lowest value in a popula-
impractical. tion.2'3 However, most practitioners have long for-
gotten sophomore statistics, and believe that
This is the third article in a three-part continuing education series on veterinary
medicine. On completion of the series, the reader will be able to identify similarities
between human and veterinary immunologic testing in diagnostic laboratories,
describe several common assays used to detect immune and infectious disease in
animals, and discuss veterinary reference values and issues associated with
veterinary specimen analysis.

Many reference values in veterinary species differ


From Antech from those in humans. Some values overlap and
Diagnostics, Irvine, others do not, depending on the species. In dogs
Calif.
for example, the normal reference value for amy-
Reprint requests to
lase can be as high as 1,125 U/L, considerably
Dr Klaassen, Antech
Diagnostics, 17672
higher than in humans. In horses, the pancreas
Cowan Ave, Suite contains little amylase, and serum amylase levels
200, Irvine, CA are lower (0-24 U/L), closer to the value in
92614; o r e - m a i l : humans. Calcium levels in many avians and rep-
jklaassen@aol.com tiles can exceed 20 to 30 mg/dL, especially in
mature egg-laying females. Reference values estab-
lished at Antech Diagnostics (Irvine, Calif) for
common chemistry and hematology parameters in
adult dogs and cats (Fig 1) are included in Tables
1 and 2, respectively.
Fig 1. The same " n o r m a l " values do not apply t o all
cats, nor to all breeds of any other animal species.

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194 LABORATORY MEDICINE VOLUME 30, NUMBER 3 MARCH 1999
on 28 May 2018
Table 1. Reference Values for Common Table 2. Reference Values for Common Hematology
Chemistry Determinations in Adult Dogs Determinations in Adult Dogs and C a t s * 1
and Cats**
Parameter Canine Feline
Analyte Canine Feline WBC count (/u±) 4,000-15,500 3,000-14,800
ALT (U/L) 12-118 10-100 6
RBC count ( x 1 0 / u L ) 4.8-9.3 5.92-9.93
A l b u m i n (g/dL) 2.7-4.4 2.5-3.9
H e m o g l o b i n (g/dL) 12.1-20.3 9.3-15.9
A l b u m i n / g l o b u l i n ratio 0.8-2.0 0.35-1.5
Hematocrit (%) 36-60 29-48
Alkaline phosphatase (U/L) 5-131 10-50
Mean corpuscular v o l u m e (fL) 58-79 37-61
A m y l a s e (U/L) 219-1,125 100-1,200
M e a n corpuscular h e m o g l o b i n (pg) 19-28 11-21
AST (U/L) 15-66 10-100

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M e a n corpuscular h e m o g l o b i n 30-38 30-38
Bilirubin, total (mg/dL) 0.1-0.3 0.1-0.4 concentration (%)
BUN (mg/dL) 6-25 14-36 WBC, differential (/u.L)*
BUN/creatinine ratio 4-27 4-33 Segmented neutrophils 2,060-10,600 2,500-8,500
Band neutrophils 0-300 0-150
Calcium (mg/dL) 8.9-11.4 8.2-10.8
Lymphocytes 690-4,500 1,200-8,000
Chloride (mEq/L) 102-120 104-128 Monocytes 0-840 0-600
Cholesterol (mg/dL) 92-324 75-220 Eosinophils 0-1,200 0-1,000
Basophils 0-150 0-150
Creatine kinase (U/L) 59-895 20-400
3
Platelet count (x10 /u.L) 170-400 200-500
Creatinine (mg/dL) 0.5-1.6 0.6-2.4
"Conventional (rather than SI) units are given, as commonly used by
G l o b u l i n (g/dL) 1.6-3.6 2.3-5.3 veterinarians in the United States.
GGT (U/L) 1-12 1-10 'Reference values were established by Antech Diagnostics, Irvine, Calif.

Glucose (mg/dL) 70-138 50-170 *Absolute counts.

Lipase (U/L) 77-695 10^150


reference range and reference values are synony-
M a g n e s i u m (mg/dL) 1.2-1.9 1.1-2.3 mous. For the sake of scientific harmony, "refer-
O s m o l a l i t y (mOsm/L) 277-311 299-330 ence values" is used here.
Determination of reference values is best per-
Phosphorus (mg/dL) 2.5-6.0 2.4-8.2
formed in the laboratory for the population of
Potassium (mEq/L) 3.6-5.5 3.4-5.6 animals that will be tested. Local hospital or ref-
Protein, total (g/dL) 5.0-7.4 5.2-8.8 erence laboratories may be able to enlist the coop-
eration of area veterinarians to supply blood from
S o d i u m (mEq/L) 139-154 145-158
clinically healthy animals. It is important to know
Triglycerides (mg/dL) 29-291 25-160 the age of the animals so that age-specific refer-
ALT indicates alanine aminotransferase; AST, ence values can be established and too many
aspartate aminotransferase; BUN, blood urea young or old animals are not included. If general, Test Time!
nitrogen; GGT, 7-glutamyl transferase. if adult references values are being determined, Look for the CE
'Conventional (rather than SI) units are given, as animals younger than 5 to 6 months should not Update exam on
commonly used by veterinarians in the United Veterinary Medicine

i
be included in the reference population. Too
States. (902) in this issue of
many geriatric animals (older than 10 years) may Laboratory Medicine.
'Reference values were established at Antech shift the reference values for some analytes. There Participants will earn
Diagnostics, Irvine, Calif. can be important differences between various 3 CMLE credit hours.
breeds of animals. The same "normal" values do
not apply to all dogs, nor do the same reference
intervals apply to all breeds of cats, cows, horses,
or birds, for example. If possible, 40 to 120 clini-
cally healthy animals of various ages and breeds,
and both sexes should be included in the general

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by guest MARCH 1999 VOLUME 30, NUMBER 3 LABORATORY MEDICINE 195
on 28 May 2018
Fig 2. Because there
is significant
difference in
baseline values for
young vs mature
dogs and other
species, animals of
various ages should
be included in a
standardized
reference
population. Animals
younger than 5 to 6
months, however,
should not be

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included in a
reference
population.

reference population (Fig 2). Before samples are analyte. If the patient daily mean is near the high
obtained, specific written criteria should be given end of the reference value, more than half of the
the veterinarian to ensure that a standardized patients had elevated values. This should also be
"reference" population is selected for the study. obvious from the number of phone calls from
Without specific guidelines, too many variables practitioners about elevated results received.
will be introduced into the population, decreasing Calculation of reference values should include
sensitivity and specificity. the mean and standard deviation (SD)—and are
Reference values can also be determined by usually the mean value ±2 SDs, or 95% of the
using the laboratory patient database, on the population. However, most of the analytes in a
premise that the values for a large enough patient chemical profile (primarily enzymes) and CBC do
population (generally several thousand animals) not show Gaussian (normal or symmetric) distri-
yield a valid distribution if guidelines for accept- bution, and conventional procedures involving
able values are used. This assumes that most val- mean and standard distribution will yield skewed
ues from most patients are "normal" and that results. This would be obvious if the data were
values at both the high and low extremes do not plotted on a graph, or by lack of agreement
influence the calculations because of the large between the median, mode, and mean. Nonpara-
number of observations in the population. Values metric analyses or transformation analyses should
for the upper and lower limits depend on the be performed to obtain the most useful reference
mathematical method used and its assumptions.3 values. To make the data appear more Gaussian,
Even if the patient database is not used to percentile or square-root transformations of the
determine reference values, it may prove valuable data can be used.4 Most computer spreadsheet
for confirming that the reference values are rea- programs or statistical packages enable perfor-
sonable. Patient daily mean reports can be gener- mance of these statistical analyses.
ated by most laboratory computer systems. These Analysis of the reference population may result
reflect the average of the previous day's results for in narrower values than previously used or pub-
all patients for all tests. The mean value generally lished in the literature. Many of the references in
does not fluctuate significantly from day to day, the literature are outdated, and the analyses were
and is often helpful for quality assurance. The performed on older equipment and reagents. In
patient daily mean value for an analyte should be addition, some of the older values are broader. It
near the middle of the reference value for that

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196
by guest LABORATORY MEDICINE VOLUME 30, NUMBER 3 MARCH 1999
on 28 May 2018
may be that the older reagents yielded higher or methods that are set to begin checking for clot for-
lower results or were less precise (more variable), mation after veterinary samples would have
or the reference values were adjusted to fit clinical already clotted. These systems falsely report ele-
appearances. Use of these values perpetuates inac- vated prothrombin time (or noncoagulated speci-
curacies and hampers correct and early diagnosis. mens) when in fact the results are normal.
If the reference value is too broad the sensitivity of Knowledge of these differences enables determi-
a test will be decreased. For example, if the upper nation of reference values that are more accurate
limit of the reference value for amylase is too high, and similar to published values.
mild pancreatitis and enteritis, or renal insuffi-

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ciency may not be evident, resulting in excessive Conclusion
false-negative results. Conversely, if the reference In summary, published values may be helpful for
value is too narrow, excessive false-positive results determination of general results for a specific assay,
are produced, and the specificity of the analyte but accurate establishment of reference values
suffers. Sensitivity reflects the number of animals should be performed for each veterinary labora-
with disease that have abnormal test results (the tory, with its own equipment and reagents. Failure
test identifies all animals with disease), whereas to do so will affect the specificity and sensitivity of
specificity indicates the number of animals that test results.®
have abnormal results that have the disease.
References
Testing Procedures That May A f f e c t 1. DiUlio M. Veterinary immunology and serology: clinical
laboratory diagnostics. Lab Med. 1999;30:36-40.
Results 2. Solberg HE. Establishment and use of reference values. In:
Instruments and reagent systems can make a sub- Burtis CA, Ashwood ER, eds. Tietz' Textbook of Clinical Chem-
stantial difference in the results obtained on vet- istry. 3rd ed. Philadelphia, Pa: WB Saunders; 1999;336-356.
3. Lumsden JH. "Normal" or reference values: questions and
erinary samples. Many modern hematology comments. Vet Clin Pathol. 1998;27:102-106.
instruments do not analyze veterinary samples 4. Kaneko J J, ed. Clinical Biochemistry of Domestic Animals.
accurately. For example, often cell-lyse systems 4th ed. New York, NY: Academic Press; 1989;2-7.
designed for human cells do not work adequately
or similarly with veterinary specimens. Many ani-
c


mal species have considerably smaller RBCs Please let us know your opinion of the Veterinary 0
Vledicine (902) series. Place an X in one box for each ft'ifi 9
(mean corpuscular volume 30-70 fL), often less •
question. Return this form (or a photocopy) by fax to: WiPi U
than the RBC counting thresholds set for human 312) 850-8817; or, mail to: ASCP Press Administration, E
3
RBCs, therefore giving falsely decreased RBC 2100 W Harrison St, Chicago, IL 60612-3798. Thank you
'or your input. £
counts and hematocrit calculations and interfer- £
o
ing with calculation of RBC indices. Deficient Excellent o
In addition, certain reagent systems are not 1. The series met the objectives stated in the abstract.

appropriate for veterinary specimens and can •1 D2 D3 D4 D5


yield erroneous results. For example, thyroxine 2. The series provided useful technical data or original ideas. u
W
values in humans are in the range of 8 to 13 • 1 D2 D3 D4 D5
|xg/dL, compared with approximately 1 to 4 (xg/dL 3. The information provided in the series was new and timely. O
in most veterinary specimens. Reagent systems • 1 D2 D3 D4 D5 E
0
and kits or instruments designed to measure 4. Technical points were explained clearly and were easy to comprehend.

human thyroxine often do not have maximum • 1 D2 D3 D4 D5


sensitivity in the lower value to accurately deter- 5. The text was organized logically.

mine veterinary thyroxine values. The same is true •1 D2 D3 D4 D5


i. Illustrations, charts, and tables helped explain text and added to series value.
for coagulation systems. Many veterinary species
have short (5-9 seconds) prothrombin time, com- D1 D2 D3 D4 D5
Comments: (Attach additional pages, if necessary.)
pared with humans. Some coagulation systems
designed for use with human samples use optical

9905


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