Normal Blood Leukocyte Concentration Values
N. G. ORFANAKIS, B.A.,
R. E. OSTLUND, B.S.,
AND J. W. ATHENS,
Department
of Medicine,
University
C. R. BISHOP,
M.D.,
M.D.
of Utah College of Medicine,
Salt Lake City, Utah S4112
ABSTRACT
Orfanakis, N. C , Ostlund, R. E., Bishop, C. R., and Athens, J. W.: Normal
blood leukocyte concentration values. Amer. J. Clin. Path. 53: 647-651, 1970.
Absolute blood leukocyte counts were determined in 291 normal individuals
(226 Caucasian and 65 Negro), using modern, electronic leukocyte counting
methods. Two hundred cell differential counts of coverslip smears stained
with Wright's stain were done. Normal mean values and 95% limits expressed
as cells per cu. mm. were: band neutrophils 500 (100 to 2,100); segmented
neutrophils 3,000 (1,100 to 6,050); total neutrophils 3,650 (1,830 to 7,250);
lymphocytes 2,500 (1,500 to 4,000); monocytes 430 (200 to 950); eosinophils
150 (0 to 700); basophils 30 (0 to 150). No clinically significant differences
between Negroes and Caucasians were found.
of our neutrophil kinetic
studies 1 ' 2 it became necessary to determine
the range of blood neutrophil values in
normal subjects as a standard of comparison for patients with disease. Although Osgood and co-workers 8-11 defined normal
blood leukocyte concentrations in considerable detail, the recent development of
electronic cell counting methods has introduced a new variable. In addition, different
values for Negroes and Caucasians have
been reported. 3 For these reasons the reevaluation of blood leukocyte concentrations in normal subjects was undertaken.
State Prison (15 Caucasian and 14 Negro
men) and trainees at the Clearfield, Utah,
Job Corps Center (23 Caucasian and 51
Negro men). The age range was 16 to 44
years for the Caucasians and 16 to 49 years
for the Negroes.
The individuals from the university and
the prison recently had undergone physical
examinations. The hematologic studies of
the Job Corpsmen were performed as part
of their entrance medical examination.
Sickle cell preparations were made for all
Negroes, and persons with positive tests
were excluded from the study.
Leukocyte Counts. Venous blood antiMaterials and Methods
coagulated with double-oxalate was used
Subjects. Only healthy persons not tak- for all leukocyte counts. The blood samples
ing medications were studied. The subject were obtained in the morning after breaksources were students at the University of fast (Job Corps trainees and prison inUtah Medical Center (188 Caucasians, 22 mates) or soon after lunch (students). Leuwomen and 166 men), inmates of the Utah kocyte counts were performed with a
Coulter Counter. 5 Two aliquots per blood
Received July 24, 1969; accepted for publication
October 10. 1969.
sample were prepared using an autodiluSupported by research and training grants AM
ter*; cetrimide-citrate was the lytic dilu04489 and AM 5098, and by a General Research
I N THE COURSE
support grant FR 05428 from the National Institutes of Health.
' Scientific Products, Evanston, Illinois.
647
648
Vol. 53
ORFANAKIS ET AL.
ent. Each aliquot was counted three times
on a model A or F Coulter Counter and the
six counts were averaged.
Coverslip blood films were prepared from
freshly-drawn blood to which no anticoagulant had been added.
Differential Counts. Two hundred cells
were counted under oil immersion after
staining with Wright's stain. Two separate
coverslips were examined, and 100 cells
were counted on each. If there was good
agreement between the two counts, 4 the
results were averaged. If agreement was
not good, two more coverslips were stained,
an additional 200 cells were counted, and
the four counts averaged. Periodically,
slides were counted by each of several
observers, including two hematologists
(C. R. B. and J. W. A.). No significant differences were observed. Since the absolute
concentration of each cell type gives more
useful information than percentage values,
the absolute concentration values were calculated from the total leukocyte count and
the differential count, and the data are reported in this form.
A segmented cell is defined as one with
a filament connecting the lobes. The filament must demonstrate length but no
breadth as one focuses through the field.
This is the same definition used by Osgood,7 except that he included those cells
with coiled or folded nuclei in which filaments could not be demonstrated (doubtful
cells) with the segmented forms. The distinction between band neutrophils and segmented neutrophils is arbitrary and has
been variously defined. Cartwright 4 defines
a segmented neutrophil as a cell in which
two or more lobes are separated by a strand
of chromatin of lesser width than either
lobe. Others have required that the connecting isthmus must have a diameter at
least a third that of either lobe. These definitions are tedious if they are to be carried
out strictly, and require measurements of
many cells. Consistency with either of these
definitions is hard to obtain. Thus, the
more easily learned and more precise definition that a filament must have length
but no breadth has been used in these studies. Since it is often useful to be able to
detect increased concentrations of young
neutrophils (band or juvenile forms) in the
blood,6 we have elected to place doubtful
cells into the band or juvenile category,
thus establishing more clearly the upper
limit of band forms in normal subjects.
Methods of Analysis. Osgood and coworkers 8 have reported normal leukocyte
and differential count values for subjects
four to seven years of age, eight to 14 years
old,9 15 to 18 years old,10 and 19 years old
and older,11 and have demonstrated higher
counts in children but no differences between the sexes. Since our subjects ranged
from 16 to 49 years in age, we examined
Osgood's data and ours for evidence of
differences between the 15- to 18-year-old
group and the older subjects. Except for
a slightly higher 95% range for total leukocyte count in the younger subjects, the
values were almost identical. Therefore, no
division of the data into subgroups according to age or sex has been made.
Frequency plots of the blood cell concentration values for band neutrophils, segmented neutrophils, lymphocytes, monocytes, eosinophils, and basophils were made.
Because a skewed distribution toward the
higher values was apparent for each cell
type, the data were replotted on logarithmic probability paper (Codex No. 3228)
and median values and 95% limits obtained from these graphs.
For comparison of values in Caucasians
and Negroes, logarithmic transformation of
each cell concentration value was determined and group comparison made using
the t test, according to the method of
Woolf.12
Results
The median and 95% limits for absolute
concentration (cells per cu. mm.) of the
several blood leukocyte types in Negroes
May 1970
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LEUKOCYTE VALUES
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and Caucasians are shown in Table 1, lines
1 and 2. T h e distribution of the values for
the two groups are shown in Figures 1
and 2. It can be seen in Figures 1 and 2
and Table 1 that the frequency distribution and median and 95% limit values are
essentially identical for lymphocytes, monocytes, eosinophils and basophils in the two
racial groups. The frequency distribution
and the median values appear to be slightly,
but not significantly, different for segmented neutrophils, but are clearly different for band neutrophils as a result of a
larger proportion of low values in Negroes.
However, since the 95% limit values for
band forms are very similar for the two
groups there is little reason to complicate
clinical interpretation by delineating two
sets of normal values. Therefore, the data
were combined and the median and 95%
values for the 291 subjects are given in
line 4 of Table 1.
It seemed of interest to compare our
values with those reported by Osgood and
co-workers.11 This comparison can be seen
in the last two lines of Table 1. It is apparent that values obtained at 4,500-foot altitude using an electronic cell counter were
essentially the same as values obtained
using the hemocytometer at sea level 30
years earlier. T h e differences in band neutrophil and segmented neutrophil values
appear to be due to the different classification of band and segmented forms (see
"Materials and Methods").
Discussion
It has been customary for many years to
determine the total leukocyte count and
the different cell count of patients and to
derive useful clinical information therefrom. However, there has often been considerable confusion about "relative" and
"absolute" changes in leukocyte count, especially in recent years with the increase
in the incidence of drug-induced cytopenias. Although Osgood and co-workers8_1X
reported normal ranges for a number of
650
Vol. 53
ORFANAKIS ET AL.
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CUMULATIVE FREQUENCY IN %
-i—i—i—i—i
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CUMULATIVE
40 60
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FREQUENCY IN %
FIG. 1. Frequency distribution of blood leukocyte concentrations in 226 Caucasian (O) and
65 Negro ( • ) normal individuals. In the graph on the left, the values for band and segmented
neutrophils are shown; on the right, the values for lymphocytes and monocytes are shown.
hematologic parameters, including "absolute" leukocyte concentrations, many years
ago, confusion still arises. Consider a patient whose leukocyte count is 2,500 cells
per cu. mm. with 5% neutrophils, 7 5 %
lymphocytes, 10% monocytes, and 10%
eosinophils. Such a patient has neutropenia with normal numbers of other cell
types present, but there is often confusing
discussion of relative lymphocytosis and
monocytosis and considerable equivocation
about eosinophilia in such cases. This confusion is readily avoided if the absolute
number of each leukocyte type is calculated and reference is made to the normal
ranges for cell concentration (Table 1).
900
700H
500
400
300-|
5
200
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FIG. 2. Frequency distribution of blood eosinophil and basophil concentrations in 226 Caucasian
(O) and 65 Negro ( • )
normal individuals.
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May 1970
651
LEUKOCYTE VALUES
Because leukocyte counts are now usually
done by electronic counting methods using
blood that has been collected in one of
several anticoagulants, it seemed worth
while to reassess normal values. It is apparent from the present study that with
appropriate attention to methodologic details the normal blood concentration ranges
for the several leukocyte types are the same
with modern methods as with the heraocytometer.
It has been suggested in a recent retrospective study that about 25% of Negroes
may have leukopenia if judged by accepted
standards. 3 In the present prospective study
it is clear that lymphocyte, monocyte, eosinophil, and basophil concentration median and 95% values are identical in Negroes and Caucasians, but differences in
band neutrophil values can be demonstrated. The reasons for the difference are
not clear but presumably they are not dietary or environmental because the difference is found in persons living in the same
environment and eating very similar diets
(Job Corpsmen and prison inmates).
The differences between band and segmented neutrophil values in the Salt Lake
City and Portland groups is the result of
a different definition of the dividing line
between these cell types. The requirement
used here, namely, that only a filament connects lobes in a segmented cell, is favored
because it is less subjective and the values
obtained are more reproducible than with
other definitions. With past definitions of
the band cell, doubtful cells were included
with the more plentiful segmented forms.0
By including doubtful cells with the band
forms, the maximum band form value is
more clearly defined and a minimal shift to
the left can be detected more easily. Marsh
and co-workers6 demonstrated the usefulness of such a definition in detecting a
"shift to the left" as the earliest sign of
infection in dogs with induced bacterial
pneumonia.
With this strict definition of segmented
neutrophils, mean lobe count values are
considerably lower than those reported elsewhere. In 20 normal students examined to
date, the mean neutrophil lobe count was
2.04 ± 0.38 (2 S.D.).
Acknowledgment.
Dr. Alley Flinders of the
Clearfield Job Corps Center and Dr. William Knott
and Warden John Turner at the Utah State Prison
helped arrange this study.
References
1. Athens, J. W., Haab, O. P., Raab, S. O., Boggs,
D. R., Ashenbrucker, H., Cartwright, G. E.,
and Wintrobe, M. M.: Leukokinetic Studies.
XI. Blood granulocyte kinetics in polycythemia vera, infection, and myelofibrosis. J. Clin.
Invest. 44: 778-788, 1956.
2. Athens, J. W„ Raab, S. O., Haab, O. P., Boggs,
D. R., Ashenbrucker, H., Cartwright, G. E.,
and Wintrobe, M. M.: Leukokinetic Studies.
X. Blood granulocyte kinetics in chronic myelocytic leukemia. / . Clin. Invest. 44: 765-777,
1965.
3. Broun, G. O., Herbig, F. K., and Hamilton,
J. R.: Leukopenia in Negroes. New Eng. J.
Med. 275: 1410-1413, 1966.
4. Cartwright, G. E.: Diagnostic Laboratory Hematology. Ed. 4. New York, Grune & Stratum,
1968, p. 146.
5. Gagon, T. E., Athens, J. W., Boggs, D. R.,
and Cartwright, G. E.: An evaluation of the
variance of leukocyte counts as performed
with the hemocytoineter, Coulter and Fisher
instruments. Am. J. Clin. Path. 46: 684-691,
1966.
6. Marsh, J. C , Boggs, D. R., Cartwright, G. E.,
and Wintrobe, M. M.: Neutrophil kinetics in
acute infection. / . Clin. Invest. 46: 1943-1953,
1967.
7. Osgood, E. E., and Ashworth, C. M.: An Atlas
of Hematology. San Francisco, J. W. Stacey,
Inc., 1937, pp. 46-48.
8. Osgood, E. E., Baker, R. L., Brownlee, I. E.,
Osgood, M. W., Ellis, D. M., and Cohen, W.:
Total, differential and absolute leukocyte
counts and sedimentation rates of healthy
children four to seven years of age. Am. J.
Dis. Child. 58: 61-70, 1939.
9. Osgood, E. E., Baker, R. L., Brownlee, I. E.,
Osgood, M. W., Ellis, D. M., and Cohen, W.:
Total, differential and absolute leukocyte
counts and sedimentation rates for healthy
children. Standards for children eight to
fourteen years of age. Am. J. Dis. Child. 68:
282-290, 1939.
10. Osgood, E. E., Baker, R. L., Brownlee, I. E.,
Osgood, M. W., Ellis, D. M., and Cohen, W.:
Total, differential and absolute leukocyte
counts and sedimentation rates of healthy
adolescents fifteen to eighteen years of age.
J. Lab. Clin. Med. 24: 905-912, 1939.
11. Osgood, E. E., Brownlee, I. E., Osgood, M. W.,
Ellis, D. M., and Cohen, W.: Total, differential and absolute leukocyte counts and sedimentation rates determined for healthy persons nineteen years of age and over. Arch.
Int. Med. 64: 105-120, 1939.
12. Woolf, C. M.: An Introduction to Biochemistry
(syllabus). Tempe, Arizona, Arizona State University, 1965, p. 128.