Comp. Biochem. Physiol., 1969, Vol. 30, pp. 695 to 713. Pergamon Press.

Printed in Great Britain

HEMATOLOGY, BLOOD CHEMISTRY AND PROTEIN

POLYMORPHISMS IN THE WHITE-TAILED DEER
(ODOCOILEUS VIRGINIANUS)*
U L Y S S E S S. S E A L and A L B E R T W. E R I C K S O N t
Metabolic Research Section, Veterans Administration Hospital, Minneapolis,
and Department of Biochemistry, University of Minnesota, Minneapolis, U.S.A.
(Received 27 December 1968)

A b s t r a c t - - 1 . Blood samples from forty-six fetal and 100 adult and juvenile
deer, Odocoileus virginianus, were analyzed for twenty-five substances including
hematology, chemistries, hormones and individual proteins with significant
differences found between ages and sexes.
2. Hemoglobin polymorphism was found in the adult and juvenile animals, but
with a significantly different frequency than reported for southern populations.
3. None of the protein changes characterizing human pregnancy were
observed in the pregnant deer.
4. The fetal samples exhibited a protein polymorphism in the post-albumin
area, a hemoglobin polymorphism, but only a single transferrin.

INTRODUCTION
THE WHITE-TAILeDdeer (Odocoileusvirginianus)is the major game animal on the
North American continent. As such, it is the most common large mammal in its
range and is subject to intensive management. Despite this interest in the species,
knowledge of its normal hematology and blood chemistry is sparse. Little is known
about the individual plasma proteins and their quantitative variations in the deer
or any wild species. Extensive work with human blood proteins has established
the usefulness of such information for determining the physiological and health
status of individuals (Putnam, 1960; Shultze & Heremans, 1966). T h e fact that
the range of this species is extensive and that it is known to exhibit hemoglobin
polymorphism (Kitchen et aL, 1964; Weisberger, 1964) and transferrin poly-
morphism (Miller et al., 1965) also offers an opportunity to study the variation of
protein phenotypes in relation to separate populations, ecology and possible
selection factors. Identification of other serum protein polymorphisms would also
be of value for such population studies.
* A joint contribution from the Departments of Biochemistry, Ecology and Behavioral
Sciences and the Museum of Natural History, University of Minnesota, and the Metabolic
Research Section, Minneapolis Veterans Administration Hospital, Minneapolis, Minnesota
55417, and supported in part by USPHS Grant AM-11376-07 from the National Institute
of Arthritis and Metabolic Diseases, the V.A. Cooperative Urological Research Project and
the Graduate School of Biomedical Sciences Support Grant USPHS 415-0749-14.
t Department of Ecology and Behavioral Sciences and Museum of Natural History,
University of Minnesota, Minneapolis, U.S.A.
695
696 ULYSSES S. SEALAND ALBERT W. ERICKSON

MATERIALS AND METHODS

The opportunity to study a large sample of a relatively homogeneous herd came when a
decision was made to remove the bulk of a population of 400 animals from an enclosed 2500-
acre area near Fridley, Minnesota. This herd was derived from an estimated original
population of ten animals which were enclosed when a fencewas erectedin 1942. Essentially
no control has been exerted on the growth of the population beyond that of winter weather
conditions and food limitations, which had led to the death of 90-100 animals during the
winter of 1966-67. The current winter of 1967-68 was relatively mild and the ground had
only light snow-cover until the time of animal removal in late February and early March.
The general condition of the herd was good as demonstrated by body weight and kidney fat
measurements, although lesser comparative body sizes, antler development and reproduc-
tion rates compared with other Minnesota deer reflect previous stressful times.
The 100 adult and juvenile blood samples were drawn aseptically from the heart, within
5-20 min after the animals were shot, into vacutainers (Becton-Dickinson Co.) containing
either no additive for collection of serum or disodium E D T A for cellular hematology and
fibrinogen, or sodium fluoride and potassium oxalate for blood urea nitrogen (BUN) and
glucose. The remaining sample was drawn into a sterile evacuated 150-ml bottle containing
1 ml of heparin. Forty-six fetal samples were drawn from the umbilical vein into a heparin-
ized syringe.
The analytical methods used have been described elsewhere including cellular hematology
(Seal et al., 1967), total protein, paper electrophoresis, polyacrylamide electrophoresis,
haptoglobin, ceruloplasmin, transferrin and serum iron (Musa et al., 1967); corticosteroids
and corticosteroid-binding globulin (Seal & Doe, 1965); protein-bound carbohydrates
(Seal, 1964; Wintzler, 1955); thyroxine-binding globulin (Nikolai & Seal, 1966); and
fibrinogen (Swaim & Feders, 1967).
The assays for glucose, BUN, creatinine, cholesterol, uric acid and PBI were done with
automated methods using Technicon Autoanalyzer ® equipment and the methods supplied
with the manifolds. Statistical analyses were made by use of the Student's t-test.

RESULTS
Hematology
T h e hematology data on seventy-six deer samples, collected on 24 and 25
F e b r u a r y and 2 M a r c h 1968, are tabulated according to sex and age in T a b l e 1.
T h e difference in b o d y weight between the adult males and females was significant
with P < 0.002. Statistically significant differences in the hematology measurements
between groups included adult females and juvenile males with respect to h e m o -
globin ( P < 0 - 0 5 ) , hematocrit ( P < 0 . 0 5 ) and m e a n corpuscular volume (MCV)
( P < 0-01); adult males and juvenile females, with respect to red blood cells (RBC)
( P < 0.05); adult males and adult females, hematocrit ( P < 0.05); adult females and
juvenile females, M C V ( P < 0 . 0 2 ) ; and adult males and juvenile males, RBC
( P < 0.02). T h e m e a n corpuscular hemoglobin concentration ( M C H C ) was nearly
maximal (Schalm, 1965) and the same in all groups. N o instance of h y p o c h r o m i c
cells was found even in animals with a significant anemia.
F o u r animals with anemia whose results were not included in the statistics of
T a b l e 1 are tabulated in T a b l e 2. All were n o r m o c h r o m i c and normocytic,
eliminating m o s t specific deficiency diseases (Wintrobe, 1962). T h e presence of
haptoglobin m a d e the occurrence of intravascular hemolytic disease unlikely.
Present b o d y condition for this herd did not indicate generalized malnutrition.
T h e most likely cause of this anemia would be blood loss.
 TABLE l--HEMATOLOGY OF THE W H I T E - T A I L E D DEER ( O . virginianus)

Weight Hgb* RBC Hct MCV MCHC

Group n (kg) (g~/o) (10e/mm 3) (vol.%) (/~3) (%)

Adult male 18 69+11-3 13.5+1.9 10.8+1.2 40+5.7 37+3.5 33.7+1.2

Adult female 33 54+7"3 14.7+2"6 11.2+1"6 44+7.4 39+3.7 33"6+1"3
Juvenile malet 14 29+4.5 13.0+2"6 11"6+2"0 39+6"6 34+3"3 33"1+1"8
Juvenile femalet 11 27+3"6 14.5+1"8 11-8+1"0 43+5.6 36+3"6 33.7+0.7

* List of abbreviations used: BUN, blood urea nitrogen; EDTA, ethylenediamine tetracetic acid; PBI, protein-bound iodine;
MCV, mean corpuscular volume; RBC, red blood cells; M C H C , mean corpuscular hemoglobin concentration; A/G, albumin to
globulin ratio; CBG, corticosteroid-binding globulin; TBG, thyroxine-binding blobulin; Hgb, hemoglobin; Hct, hematocrit.
Juveniles are fawns of the year, approximately 9 months of age. All data in the tables are given as mean _+ standard deviation.

TABLE 2--BLooD STUDIES ON FOUR ANEMIC W H I T E - T A I L E D DEER

Serum
Weight Hgb RBC Hct MCV MCHC protein Haptoglobin
Animal (kg) (g%) (10e/ram s) (vol. ~/o) (/z3) (%) (g%) (mg ~o)

Adult female 58 7.9 6-12 23 37 34-3 5.3 8

Adult female 52 7"0 5"90 20 34 35"0 5"3 55
Juvenile female 28 7"8 6"53 24 37 32"7 5"0 24
Juvenile male 35 6.5 5.97 21 35 31.0 4"4 26
 T A B L E 3 - - H E M A T O L O G Y OF ADULT FEMALE DEER COLLECTED IN SUCCESSIVE SAMPLES

Weight Hgb RBC Hct MCV MCHC

Date n (kg) (g%) (106/mm 3) (vol%) (/z3) (%)

24 F e b r u a r y 13 58 + 7"7 14"2 + 2-6 10"8 + 1.9 43 + 7'2 40 + 3"6 33"2 + 1'2

25 February 11 51+6"8 14-3+2.2 11.2+1"4 42+4"9 38+3"5 33"9+1"5
2 March 9 51 + 6"4 15-8 + 3-1 11"9 + 1-4 47 _+9-7 39 + 4.0 33.7 _+ 1-1

TABLE 4----HEMATOLOGY OF TWO FETAL DEER SAMPLES COLLECTED 7 days APART*

OO
Weight Hgb RBC Hct MCV MCHC
Group n (g) (g%) (10e/mm 3) (vol.%) (/z3) (%)

24 F e b r u a r y
Male 15 294 + 75 8"1 + 0'8 5.4 + 0"6 36 + 3 68 + 7 22 + 1"8
Female 14 259 + 76 8"3 + 0'8 5.7 + 0.6 35 + 3 63 + 7 23 + 2'0
2 March
Male 14 375 _+73 8.7 _ 0.8 5.8 +_0.5 37 _ 4 63 __ 8 24_+ 1.4
Female 8 401 _+79 9"6 +_ 1"0 6"6 +_0"9 4l + 4 64 + 5 23 + 0"7

* T h e age of the fetuses was approximately 2-2" 5 months.

 BLOOD C H E M I S T R Y OF W H I T E - T A I L E D DEER 699
Since these samples were taken from animals collected successively from the
same herd, the possibility of a selective, sequential sampling bias existed and was
tested for in the adult female deer sample for which an adequate number of data
were available (Table 3). T h e only comparison yielding a statistically significant
difference was for body weight, P < 0.02 between days 1 and 2. T h e animals were
collected from several separate sections of the area. Analysis of the data on the
basis of area revealed no significant differences in cellular hematology, blood
chemistry or protein polymorphism frequencies. T h e hematology studies on the
fetuses (Table 4) are grouped according to the sex of the fetus and the date of
collection. The incidence of twins was the same on both dates; there were no
triplets. T h e mean body weights increased significantly in 1 week for the fetal
males (P<0.01) and for the fetal females (P<0.001). The differences in body
weight between sexes on each of the dates were not significant. T h e 7-day increase
in hemoglobin concentration was significant for the males (P< 0.05) and for the
females (P < 0.005). T h e increase in RBC was significant for the females (P < 0.02),
as was the increase in hematocrit ( P < 0.002). There was no significant change in
MCV but the M C H C increased significantly in the male fetuses ( P < 0.005). The
between-sex comparisons showed significant differences in the 2 March sample for
hemoglobin, RBC and hematocrit with P < 0.05 in each instance.

Blood chemistry
Analysis of the blood chemistry data revealed no significant differences between
groups for urea (BUN) only (Table 5). Uric acid was significantly lower in the
adult males than the adult females (P < 0.0001); juvenile males (P < 0.000001) and
juvenile females (P<0-02). The plasma glucose of the adult males was also
significantly lower than in the adult females (P < 0.005), juvenile males (P < 0.001)
and juvenile females ( P < 0.05). In contrast the plasma cholesterol values were
higher in the adult males than the adult females ( P < 0.0001), juvenile males
(P < 0.05) and juvenile females (P < 0.01). Also, the plasma creatinine levels were
higher in the adult males than in the adult females ( P < 0.001) or juveniles ( P <
0-001). T h e adult female levels were higher than the juveniles ( P < 0.01). Analysis
of ten mixed-sex fetal plasma samples yielded a mean creatinine of 0.8 + 0.2 mg°/o
and a mean plasma cholesterol of 42 + 5 rag% (Table 5).
Again, considering the possibility of successive sampling bias, the blood
chemistries of the adult females were analyzed for each day (Table 6). None of the
differences were statistically significant in this comparison.
Serum protein analyses were made by paper electrophoresis on ten samples
from each of the four groups and also for ten fetal samples (Table 7). Calculations
of albumin and immunoglobulin (gamma globulin) concentrations, and albumin to
total globulin ratios (A/G) revealed no statistically significant differences between
any of the juvenile and adult deer groups, in contrast to the report of Cowan &
Johnston (1962). Our mean total serum protein values are also lower than those
reported by Ullrey et al. (1967). T h e fetal samples contained significantly less
total protein, less albumin and less (probably none) gamma globulin (Fig. 1). The
 TABLE 5--BLOOD CHEMISTRY OF THE W H I T E - T A I L E D DEER

BUN U r i c acid Creatinine* Glucose Cholesterol

Group n (mg%) (mg%) (mg~/o) (nag~o) (mg%)

A d u l t male 18 16 + 7 0.2 + 0.1 2.6 + 0.5 149 + 62 60 + 10

A d u l t female 33 13 + 7 0.6 + 0.5 2.1 + 0.3 256 + 172 47 + 10
J u v e n i l e male 14 12 + 7 0"7 + 0"3 1"8 + 0"3 293 + 139 52 + 10
J u v e n i l e female 11 13 + 8 0-8 + 0"8 1"8 + 0"3 299 + 231 49 + 11
Fetal g r o u p 10 -- -- 0-8 + 0"2 -- 42 +_ 5

* n = 10 for each g r o u p .
g

TABLE 6--BLooD CHEMISTRY OF ADULT FEMALE DEER COLLECTED IN SUCCESSIVE SAMPLES

Serum
protein BUN U r i c acid Glucose Cholesterol
Date n (g%) (mg%) (mg~o) (mg%) (mg~o)

24 F e b r u a r y 13 5"3 + 0 , 8 11 + 7 0.9+0.7 243+175 46__+ 8

25 F e b r u a r y 10 5.3 +__0.4 14 +__3 0"5 +__0.2 205 + 122 47 + 11
2 March 12 5.2 + 0.8 15 __+9 0"5 __+0.3 269 + 145 48 +__11
 TABLE 7--PAPER ELECTROPHORETIC SERUM PROTEIN FRACTIONS OF W H I T E - T A I L E D DEER

Gamma
Serum protein Albumin globulin
Group n (g%) (g~o) (g~o) A/G*

Adult male 10 5.2 ± 0.3 3.25 ± 0.30 0.71 ± 0.15 1.7±0.3

Adult female 10 5"2 ± 0"6 3"34 ± 0"37 0"75 ± 0-10 1.8±0.3
Juvenile male 10 4"9 ± 0"8 3"14 ± 0"73 0"64 ± 0"16 1. 8±0. 4
Juvenile female 10 5"0 ± 0"4 3"20 ± 0"30 0"70 ± 0-19 1. 8±0. 4
Fetal 10 2"8 ± 0"2 1"12 ± 0"10 0"12 ± 0-05 0.7±0.1

* A / G = ratio of albumin to total globulin concentration. o

T A B L E 8 - - I N D I V I D U A L PLASMA PROTEINS IN WHITE-TAILED DEER

Transferrin Iron Haptoglobin Fibrinogen Ceruloplasmin

Group n (/~gFe bound/100 ml) (/~g%) (mg%) (mg~o) (AAs,0)

Adult male 17 424 + 83 234 + 50 29 + 29 266 + 50 0"22 + 0"08

Adult female 27 400 + 70 228 + 50 23 + 20 318 + 120 0"23 __+0"09
Juvenile male 10 399 + 53 290 + 68 17 + 10 232 + 61 0"26 + 0.05
Juvenile female 9 393 + 99 305 + 94 27 + 28 271 ± 142 0"27 + 0.07
702 ULYSSES S. SEAL AND ALBERT W . ERICKSON

absence of gamma globulin was reflected in the absence of the characteristic 7S

peak in the fetal serums by molecular weight analysis in the analytical ultra-
centrifuge (Fig. 2). The reversed or lower fetal A/G ratio on paper electro-
phoresis reflected not only the lower albumin concentration, but the presence of a
protein of alpha-one globulin mobility in a mean concentration of 1.1 + 0.1 g°/o.
This fraction probably contained a protein homologous with fetuin as identified
in bovine fetal serum (Engle & Woods, 1960).
Quantitative analysis of deer plasma for fibrinogen (Table 8) yielded levels in
adult females which were significantly higher than in juvenile males ( P < 0.01),
while the other differences were not significant. Mean serum levels of ceruloplasmin,
an index of acute and chronic inflammatory processes (Rice, 1960; Seal, 1964),
were the same in all groups as were the mean concentrations of haptoglobin. Serum
iron levels were higher in the juveniles than adults, with the four possible com-
parisons significant (0.02<P<0.05). The adult and juvenile groups did not
exhibit a sex difference within the age group. Transferrin, the iron-binding and
iron-transport protein of serum, was present in equivalent amounts in all four
groups with the result that relative saturation of the total iron-binding capacity was
higher in the juveniles.
Serum protein-bound carbohydrates are an index of serum glycoprotein levels
and changes (Wintzler, 1955; Seal, 1966). Each serum protein contains a charac-
teristic amount of each carbohydrate. Acute and chronic illness and pregnancy are
associated with changes in glycoproteins which may be detected by measurement
of the carbohydrates. The adult females had significantly higher hexose ( P < 0.02)
and hexosamine (P<0.05) than the juvenile males. The juvenile males had
significantly higher fucose ( P < 0.01) and sialic acid (< 0.01) than the adult males.
The adult females also had higher hexosamine levels ( P < 0.05) than the juvenile
females. The fetal sera had significantly higher hexose, hexosamine and sialic acid
levels and equivalent fucose levels. These values are particularly impressive in
view of the significantly lower serum protein levels in fetal serum (Table 9).

TABLE 9--SERUM PROTEIN-BOUND CARBOHYDRATES IN THE WHITE-TAILED DEER

Hexose Hexosamine Fucose Sialic acid

Group n (mg%) (mg%) (mg%) (mg%)
Adultmale 10 59_+ 5 61_+ 8 5-1_+1.1 49_+ 9
Adult female 10 55 -+ 10 66 -+14 5.9 _+1.4 54 _+ 7
Juvenile male 10 56 _+ 3 55 _+ 7 6"5 _+0"9 60 _+ 7
Juvenile female 10 58 _+ 6 55 _+ 8 5.7 _+1"3 56 _+ 9
Fetal 10 89_+ 9 83_+11 5'8_+1"6 97_+23

The hemoglobin data of Table 1, with the relatively large standard deviations,
suggested a heterogeneous distribution of the values. Presentation of the data in
a frequency distribution indicated the presence of a bimodal distribution of
hemoglobin levels in the adult females (Fig. 3). A heterogeneity of glucose data
 ~ ~rum 5.0g
F~t41 D66r Se~rum . . . . . . . . S.Og

A
FIc. 1. Scan records of serum protein paper electrophoresis strips from a pregnant
and fetus deer demonstrating absence of gamma globulins, the elevated alpha-1
component and the low albumin in the fetal sample relative to the adult doe.

FIc. 2. Ultracentrifuge patterns of fetal (bottom pattern) and deer serums (top
pattern) illustrating absence of 7S components (the smaller peak in the top pattern)
in the fetal serum. Protein concentration was 1 g~o, the centrifuge was operated at
60,000 rev/min and the picture was taken at 56 min after start of run.
F I c . 5. Polyacrylamide electrophoresis pattern of adult and fetal deer hemoglobins.
T h e respective samples were (1) human hemoglobin A, (2) deer fetal F2, (3) fetal
F1-F2, (4) deer type I - I I I , (5) deer type III, (6) deer type I - I I I .

F : c . 6. Polyacrylamide electrophoresis patterns of fetal and adult deer serums

illustrating post-albumin polymorphism in the fetal deer serums. T h e respective
bands indicated by the arrows from left to right are Ps, Py, and fetuin. The
respective samples are No. 1, adult deer and Nos. 2,3,4, fetal deer.
FIG. 7. Radioautograph of deer serums after electrophoresis with added laII-
thyroxine demonstrating presence of two thyroxine-binding components, as
compared to three in human serum. T h e dense band indicated by the left arrow
is albumin, the other is T B G . T h e respective samples were (1) human serum,
(2) adult female deer, (3) adult male deer and (4) fetal deer serum.
 BLOOD CHEMISTRY OF WHITE-TAILED DEER 703

(Table 5) was reflected in skewed frequency distributions (Fig. 4). For purposes
of further analysis, all data on deer with hemoglobins greater than 17"0 g% were
grouped and all data on deer with blood glucoses greater than 400 mg% were
grouped. These results (Table 10) indicate additional differences in these groups
when compared to the data in the other tables. Thus, the high glucose group had a

Juvenile ~ dov,nil~ 6'

¢, : ,, I,I,L
Adult o~
Z
4

10 12 14 10 18 10 12 14 1G 1B
Hemoglobin, g m / l o o ml
FiG. 3. Frequency distribution plots of hemoglobin data for the adult and juvenile
groups.

4t ,luv~,nlb duveniL~

t,,,,.
2

12
I I
L
I I I I I

10

8
E
Z G

200 400 (.-008OO

i i w i
[]
~
~00 400 G00 800
i J ,

G l u c o s e , r a g / t o o m[
FIG. 4. Frequency distribution plots of plasma glucose concentration data for the
adult and juvenile groups of deer.
704 ULYSSES S. SEAL AND ALBERT W. ERXCKSON

significantly higher serum iron (P< 0.02) and a higher uric acid (P< 0.001). The
high hemoglobin group, composed predominantly of adult females, had a high
BUN (P< 0.05) and a high serum iron (P< 0.01) compared to the remaining adult
females. The only overlap between the two high groups was a juvenile male who
had a hemoglobin of 17.6 g~o and a blood glucose of 486 mg%.
An additional set of comparisons with the adult female group of deer was made
with respect to pregnancy (Table 11). For this purpose the total group of adult
females was sorted into those not pregnant, those pregnant with hemoglobins less
than 17 g~o and those pregnant with hemoglobins greater than 17 g~o. The
anemic deer of Table 2 were not included. The non-pregnant group had a signi-
ficantly lower body weight (P< 0.002) and were probably yearlings. Comparison
with the pregnant, low hemoglobin group yielded one other significant difference--
lower fibrinogen (P < 0.002). Comparison of the low and high hemoglobin groups
demonstrated a significantly lower BUN and serum iron (P<0.05) in the low
hemoglobin group.
Protein polymorphisms
Hemoglobin samples from the deer and fetuses were analyzed by polyacrylamide
electrophoresis and the unstained gels photographed, as in Fig. 5. Two phenotypes
were found in the deer and two additional, different types were also identified in
the fetal samples. The relative proportions of each hemoglobin type, identified
according to the scheme of Kitchen et al. (1964), are tabulated in Table 12. The
relative amounts of the slower component, when present, varied in different deer.
The concentration of the slow component was always less than that of the fast
component in both the fetuses and the deer. No instance of a sample containing
only the slow component was found, either in the deer or their fetuses.
Comparison of the frequency of the type I hemoglobin in this population with
a Florida population (Kitchen et al., 1964; Kitchen, 1965) and a Missouri popula-
tion (Miller et al., 1965) demonstrated a significantly lower frequency in the
Minnesota populations (X2= 17, P < 0.01).
Comparison of adult and fetal deer serum protein gel electrophoretic patterns
demonstrated the presence of bands in fetal serum that were not evident in adult
patterns (Fig. 6). The band immediately behind albumin corresponded in mobility
and quantity to bovine fetuin. The polymorphic group of fetal proteins was
intermediate in mobility between the fetuin-like protein and the transferrin of fetal
serum. The three phenotypes observed in the forty-one fetal serums examined are
illustrated in Fig. 6. This post-albumin, here temporarily designated P with
subscriptf for the fast variant and s for the slow variant, was typed in the forty-one
fetal serums giving the results shown in Table 13. The data were also examined
relative to the fetal hemoglobin type. The frequency of t h e f g e n e was 0-659 in the
entire group and did not segregate with respect to hemoglobin type. The protein
has not been identified, but it did not bind iron or hemoglobin nor react as a
p-phenylenediamine oxidase, and hence was not a transferrin, haptoglobin or
ceruloplasmin.
 T A B L E 1 0 - - A N A L Y S E S O1: DEER SAMPLES SEPARATED ON THE BASIS OF H I G I I PLASMA GLUCOSE OR H I G H H E M O G L O B I N

Hgb MCV MCHC Protein BUN Uric acid

Group n (g % ) (p a) (% ) (g ~o) (rag %) (rag %)

High glucose* 15 13"6 _+2.2 37 _+3"3 33'9 + 1"4 5"0 _+0"7 9"5 _+5.7 1 '3 _+0"6
High hemoglobin t l0 18-2 _+1,0 41 + 3"4 33"9 + 1-6 5-7 _+0-7 18"5 + 6"3 0.5 + 0'4

Glucose Cholesterol Fibrinogen Ceruloplasmin Iron

(mg%) (mg%) (mg %) (A//5a0) (~ttg%)

High glucose 525 -+ 103 48 _+11 253 _+92 0"24 _+0"10 269 _+68
High hemoglobin 163_+116 51_+ 8 256+88 0-24_+0"11 288-+64

* The high glucose group (400 rag% or higher) contains one adult male, seven adult females, three juvenile females and four
juvenile males.
t The high hemoglobin group (17"0 g% or higher) contains seven adult females and one each of the other three groups.
tart

T A B L E 1 1 - - C O M P A R I S O N OF N O N P R E G N A N T AND PREGNANT FEMAI,E DEER GROUPS

Weight Hgb Protein BUN Cholesterol Fibrinogen Iron

Group n (kg) (g%) (g%) (mg%) (mg%) (mg%) (/~g%)

Total 33 54+7'3 14.7_+2.6 5"3+0'8 13-+7 47+_10 318+_120 228+50

Nonpregnant 5 46_+3'2 12.8_+1"9 5"0-+0.9 9_+4 42_+ 4 175_+ 30 234_+33
Hemoglobin ~<17"0 and 18 55_+7"7 13-7+-1'6 5"2_+0.5 12+-6 46+_ 8 300-+108 219+43
pregnant
Hemoglobin >17.0 and 7 57-+7"3 18-2+-1"0 5'7_+0'7 18+-6 51_+ 8 256+- 88 288-+64
pregnant
706 ULYSSES S . SEAL AND A L B E R T W . ERICKSON

The adult serums were examined for the transferrin types described by Miller
et al. (1965). We found only the two-band type, T F a, in our Minnesota sample,
again a significant difference from the Missouri population. Identifications of the
bands was accomplished by use of radioautography. The fetal serum samples

TABLE 12--HEMOGLOBIN TYPES IN FETAL AND ADULT DEER

Hemoglobin type n %

Fetal
F2 17 30
F1-F~ 40 70
Adult
III 30 30
I-III 70 70

contained primarily a single transferrin band corresponding in mobility to the

slower of the two adult transferrins. Small and variable amounts of the faster band
were occasionally evident.

TABLE 1 3 - - A FETAL SERUM PROTEIN P O L Y M O R P H I S M AND ITS FREQUENCY IN RELATION TO

FETAL H E M O G L O B I N TYPE

Serum protein type

Pf PaPs Ps
Group n % (n) % (n) % (n) Pf frequency*

Total 41 46"4 (19) 39"0 (16) 14-6 (6) 0-659

Hgb F, 12 41-7 (5) 50'0 (6) 8"3 (1) 0"667
Hgb FI-F2 25 44.0 (11) 36"0 (9) 20'0 (5) 0-620

/ %
Calculated according to formula: |%PI+
\ 2 7100"

Hormones and hormone-binding proteins

Assay of serums from each of the groups for fluorimetric corticosteroids, total
serum iodide and protein-bound iodide yielded the results summarized in Table 14.
The results of studies for corticosteroids indicated the presence of cortisol with the
values in the table representing maximal values since the methods used tend to
overestimate the steroid present. The low values for corticosteroid-binding
globulin were in the range found for other members of the Bovidae and Cervidae
(Seal & Doe, 1965). Comparison of CBG and TBG values in five pregnant and
five non-pregnant does yielded identical values in both groups.
 T A B L E 1 4 - - - - H O R M O N E S AND H O R M O N E - B I N D I N G PROTEINS IN WHITE-TAILED DEER

ll-OHCS CBG TSI PBI TBG*

Group n (beg%) (beg% cortisol bound) (beg 1 % ) (/xg 1 % ) (beg% T , bound)

Adult males 10 4+2 11.9+2.9 6"4+2'6 46±15

Adult females 10 6+3 5"6+1-1 9.6+2.3 6"1+1"6 34+ 4
Juvenile males 10 4+2 4'4+1"1 9.8+2'4 4"9+2"6 29+ 6
Juvenile females 10 10.7+4.5 7.0+2.7 37±12

* n = 5 in each group for these assays.

T S I = Total serum iodine. PBI = Protein-bound iodine.
708 ULYSSES S. SEAL AND ALBERT W . ERICKSON

The protein-bound iodide values represented thyroxine, as verified by an assay

with greater specificity (Murphy et al., 1966). The presence of two thyroxine-
binding components in deer serum was indicated by radioautography (Fig. 7). The
slower component appeared to be homologous with thyroxine-binding globulin
since it bound thyroxine and tri-iodothyroxine and it was saturated at relatively

TABLE ]5--SUMMARYOF SIGNIFICANT DIFFERENCES IN THE INTERGROUP COMPARISONS

Groups* Adult female Juvenile male Juvenile female

Adult male Weighff RBC~ RBC~
Hematocrit~ Uric acid~ Uric acid~
Uric acid~ Glucose~ Glucose~
GlucoseS. Cholesterol~ Cholesterol~'
CholesteroH' Creatinine~' Creatinine'~
Creatinine~ Iron~ Iron~
Fucose~
Sialic acid~
Adult female Hemoglobin~ MCV~'
HematocritJ' Creatinine~
MCV~ Iron~
Creatinine~' Hexosamine~'
FibrinogenJ~
Iron~
Hexose~
Hexosamine~
* The direction of the significant difference as indicated by the arrow is with respect to
the group in the left-hand column. Thus, the adult males weighed more than the adult
females, etc.

low thyroxine levels. The faster component was not saturated and migrated
identically with deer albumin. Support for a high-affinity thyroxine-binding
globulin was provided by a lalI-thyroxine half-life study which yielded a value of
53 hr (Burke, 1966). A summary of the statistically significant comparisons
between groups is presented in Table 15.
DISCUSSION
The major limitations and assets of this study are that the sample represented
one time of the year and a relatively restricted breeding population. The sample
studied represented about 20 per cent of the herd.
The studies of cellular hematology yielded only four anemic deer or 5 per cent
of the sample. The anemia did not appear to be the result of nutritional deficiency
or malnutrition, but rather of unexplained blood loss. This interpretation will
require further study of living animals including red-cell life-span measurements
(Noyes et al., 1966). The majority of significant differences found were between
the adult and juvenile (9-10-month-old) groups, with the juveniles having a larger
number of red cells of slightly smaller size. The hemoglobin content of the red
 BLOOD CHEMISTRY OF WHITE-TAILED DEER 709

ceils was nearly maximal [35-36 per cent is the maximum attainable value according
to Schalm (1965)]. The complete lack of any hypochromic samples would indicate
that iron-deficiency anemias are unlikely in free-ranging deer. This was borne out
by the relatively high serum iron levels found in all the adult and juvenile groups.
Hematology data available in the literature on O. virginianus include samples
from two does, with values close to those reported here, and their four newborn
fawns (Youatt et al., 1965). Seven malnourished and sick male fawns (Teeri et al.,
1958) of the same age as our juvenile group had very high hemoglobin and RBC
values, greater than 17 g% and 16 x 106 per mm 3, relative to the present data.
Another study of fourteen captive-born pregnant deer, being utilized for winter
browse digestibility studies, also yielded high values including hemoglobins greater
than 20 g°/o, hematocrit greater than 54 per cent and RBC of 17 x 106 per mm 3
(Ullrey et al., 1964). Data on O. hemionus includes three reports with results
generally comparable to those described here (Browman & Sears, 1955; Rosen &
Bischoff, 1952; Kitts et al., 1956).
The hematology of the fetal blood samples was very different from that of the
juvenile and adult groups, including lower hemoglobin levels, fewer red cells of
nearly twice the size and containing a very low proportion of hemoglobin, i.e. they
are macrocytic and hypochromic. Comparable data are available on domestic
ungulates (Altman& Dittmer, 1961). The fetal and newborn hematology data
available (Wintrobe & Schumacher, 1936; Seal et al., 1967) indicate that macro-
cytic cells are the rule, but the evidence concerning hemoglobin content shows
essentially normochromic cells, except for the mouse (Schribner et al., 1968).
Calculation of the data of Youatt et al. (1965) on four newborn fawns yields a mean
MCHC of 24 per cent and an MCV of 31, suggesting that the hypochromic state
continues until birth in the white-tailed deer.
The blood chemistry studies of this herd yielded significant differences,
primarily in the comparisons between the adult males and juvenile groups, in-
cluding lower uric acid, higher creatinine, higher cholesterol and lower glucose in
the adults. There were no differences ill urea. Our mean cholesterol values
(47-60 mg%) are higher than the 30 mg% reported by Wilber & Robinson (1958)
on ten samples collected in November. The plasma glucose values are unusual with
respect to the range of values and the frequent very high levels encountered in this
sample. The data in the literature on deer give values ranging from 25 to 125 mg°,o
(Bandy et al., 1957; Teeri et al., 1958; Youatt et al., 1965). We find similar values
in deer captured with immobilizing agents. It appears likely that the levels found
in this sample reflect the response of the white-tailed deer to severe acute stress,
including fatal trauma. Adrenaline infusions in the dog are reported to produce
hyperglycemia (Shin et al., 1968) as well as hypophosphatemia. Plasma glucose
levels are also increased by cortisol and the hyperglycemia in turn produces an
increase in plasma histamine (Kovacs & Sufliad, 1968). Glucagon administration
also effectively elevates blood glucose levels by mobilization of liver glycogen
(Perkoff et al., 1962). It may well be that these actions of adrenaline, eortisol and
glucagon summate in the acutely, severely traumatized deer to produce the levels
710 ULYSSES S. SEAL AND ALBERT W . ERICKSON

of hyperglycemia found in many of these animals. It is notable that the adult

males exhibited a lesser response.
Ten specimens from each of the groups were analyzed by paper electro-
phoresis for each of the conventional serum protein fractions. No significant
differences were found between the juvenile and adult groups with respect to any
of the measurements, including albumin or albumin to globulin ratio. Our values
for total serum protein are 0.5-1-0 g% lower than those reported for O. hemionius
(Bandy et al., 1957). However, their season of collection was summer and the
effects of season remain unexplored in the white-tailed deer. The fetal samples had
lower total serum proteins, lower albumins and probably absent gamma globulin,
although this will require immunochemical study. The fetuin-like component has
been described in a number of ungulate species (Engle & Woods, 1960).
Measurement of serum protein-bound carbohydrates provides a sensitive,
nonspecific index of growth, acute and chronic disease processes, cancer, trauma
and protein abnormalities, whose usefulness has not been explored in non-domestic
species. The major group differences found in this study included high values for
hexose, hexosamine and sialic acid in the fetal samples and differences between the
adult and juvenile groups. No sex differences within the age groups were evident.
Quantitative measurement of individual plasma proteins allows more specific
assessment of disease processes, alterations in hormonal status including pregnancy
(Musa et al., 1967), and perhaps changes in response to chronic psychosocial
disorder (Seal & Eist, 1966; Beasley & Seal, 1969). The applicability of these
concepts and this methodology to problems of population biology deserves
exploration. In this study, we have quantitated gamma globulin, serum albumin,
transferrin (and iron), haptoglobin, fibrinogen, ceruloplasmin, corticosteroid-
binding globulin and thyroxine-binding globulin (and thyroxine). The concentra-
tions of these specific proteins were the same in the four groups, except for the
higher fibrinogen in the pregnant adult females as compared to the juvenile males.
Localization of these proteins on polyacrylamide-gel or starch-gel electrophoresis
patterns would be useful to identify and interpret the changes recorded by Tets &
Cowan (1966) in their starch-gel electrophoresis study of deer serums. Specific
identification of individual proteins also greatly increases the power of electro-
phoresis as a tool for comparing populations with respect to protein polymorphisms
and variants. Evidence that these proteins in the deer may respond to the stimulus
of inflammation was provided by an adult female animal with a fibrinogen of
700 mg%, a haptoglobin of 250 mg% and a decreased serum iron of 136 m g % - -
all changes which are consistent with an acute, infectious process.
The lack of any notable pregnancy effects, at this stage of gestation in the deer,
when compared to human changes, was notable. During human pregnancy there
is a decrease in serum protein, albumin, hemoglobin and serum iron, and an increase
in fibrinogen, ceruloplasmin, transferrin, CBG, TBG, serum corticosteroids and
PBI (Musa et al., 1967; Seal & Doe, 1966). The only one of these changes at all
evident in this study of deer was an increase in fibrinogen. Possible further changes
later in pregnancy are being explored.
 BLOODCHEMISTRYOF WHITE-TAILEDDEER 711

Another area of interest to population biology is the assessment of genetic

variability in individuals and in separate populations of a species. Protein poly-
m o r p h i s m s furnish one example of such genetic heterogeneity and further suggest
that active selection m a y be occurring with respect to a particular gene as expressed
in its ultimate protein product. White-tailed deer hemoglobins have been shown
to be heterogeneous with identification of seven adult phenotypes and two fetal
phenotypes (Kitchen et aL, 1964; Miller et aL, 1965). W e have found b o t h fetal
phenotypes b u t only two adult phenotypes in our sample. I t will be of interest and
value to type deer f r o m others parts of Minnesota and the surrounding states to
determine if the frequencies of these hemoglobin chain alleles are different f r o m
those reported for the Florida and Missouri populations or if the distribution in our
sample is the consequence of a " f o u n d e r effect" and relative isolation during the
growth of the herd. T h e same considerations apply to the analysis for transferrin
types. T h e discovery of a p o l y m o r p h i c fetal s e r u m protein system is of interest
and deserves further exploration to identify and characterize the protein since
demonstration of such a system in fetal s e r u m has not been described.

Acknowledgements--Appreciation is extended to the following agencies and persons for

assistance rendered during the field collection phase of this study: Messrs. R. D. Wettersen,
P. D. Karns and J. Idstrom, Division of Game and Fish, Minnesota Department of Con-
servation; Major F. Von Gortler, Military Commander, New Brighton Arsenal; Mr. J. Mayo,
Veterans Administration Hospital, and Mr. K. Keenlyne, Graduate Student, University of
Minnesota.
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Key Word Index--White-tailed deer; Odocoileus virginianus; comparative hematology;

Blood proteins; hemoglobin; transferrin; blood chemistry; protein polymorphisms;
pregnancy; thyroxine; cortisol; fetal proteins; ungulates; population biochemistry; protein-
bound carbohydrates.