242
LETTERS TO THE EDITOR
A.J.C.P. • August 1981
What's an RDW?
To the Editor:—In their interesting
article on red cell distribution, Hammersley and colleagues 5 leave three
questions unresolved. First, do hemoglobins S-A and C-A refer strictly to
S- or C- trait, conventionally designated AS or AC 6 or do these groups
include double heterozygotes for S or
C and for /3 + -thalassemia? Did any of
these patients have concurrent athalassemia, or iron, folate, or vitamin
B12 deficiency? Were any of these patients actually SS or AS patients given
AA blood, or AA patients given AS or
AC blood? Second, how many patients with the same abnormal hemoglobin electrophoresis did not have
abnormal RDW (red cell distribution
width)? Third, what was the RDW
measurement in normal subjects?
We have found that the RDW as
measured in the Coulter Counter Model
S-Plus in 1980 had a duplicate error
range of 1.1 U 2 ; the precision trial of
Hammersley and colleagues 5 showed
2 SD = 0.7 U. Our studies with 683
normal subjects showed RDW of 7 . 9 12.3 U: the higher values included no
subjects with AS or AC, while there
were three with AS who had RDW
< 11.5 U. Therefore, RDW < 12.5 U
should be considered ambiguous for
normal versus abnormal: this range
would include 42 of the 111 with
SA and 16 of the 63 with CA. The
RDW as measured by the Coulter
Counter S-Plus II is expected to have
greater precision. 4 Our smaller groups
with only S- or C- trait had heterogeneity of red cell size indistinguishable from AA normals, while subjects
with SS, SC, and S-thalassemia did
have increased red cell volume heterogeneity: in hemoglobinopathies, anisocytosis seems to correlate with anemia.'
In contrast, patients with iron, folate,
or vitamin B 12 deficiency all have increased heterogeneity of red cell volume. Such anisocytosis is an effective discriminant of iron deficiency vs
heterozygous thalassemia in subjects
with low mean cell volume. 3
Even allowing for probable duplicate
samples of the same patient, it seems
likely that among the patients providing 40,850 blood specimens, far
more had AS or AC than the 174 found
by the 800 electrophoreses prompted
by high RDW. The patients with SA or
CA and high RDW may include:
the high-side distributional tail of RDW
among a far greater number of patients
with S- or C- trait; transfused patients; patients with the trait and nutritional deficiency; and those doubly
heterozygous for S or C and for aor /3+-thalassemia (the last would have
slight anemia and therefore be expected
to have slight anisocytosis). The authors
appropriately caution that high RDW
should not be the primary screen for
hemoglobinopathy. 5 Only a small fraction of AS or AC patients would be
detected, with very poor sensitivity.
Nutritional deficiency or transfusion,
the most common causes of anisocytosis, would have to be ruled out 1
before heterozygous hemoglobinopathy
can be accepted as the cause of
anisocytosis; therefore, specificity of
high RDW for hemoglobinopathy also
would be poor. AS and AC could not
reliably be inferred from the peripheral
blood film. Hemoglobins SS, SC, CC,
and S-j8°-thalassemia are associated
with anisocytosis and do have abnormal
blood films suggesting the diagnosis.
However, such patients were only 9%
of all with high RDW. Therefore, not
only is heterozygous hemoglobinopathy
unlikely to usefully correlate with high
RDW, but hemoglobin electrophoresis
probably should be deferred (in the absence of a suggestive smear) until
likelier causes of the high RDW had
been evaluated.
greater number had concurrent folate
deficiency. As far as we could ascertain, none of these patients had athalassemia or vitamin B 12 deficiency.
Some of the S-S and S-C patients
were given A-A blood. Our S-S and S-C
ladies from the obstetrics department
were erythrocytopheresed and exchange
transfused periodically during their
pregnancies to enable them to deliver
full-term healthy babies. These ladies
had hemoglobin electrophoreses to
monitor the amount of A-A blood remaining as they progressed through their
pregnancies and the values would
change each time.
As far as we know, no A-A patients
were transfused with A-S or A-C blood
during the time of this study. We do
know that it has happened occasionally.
DAVID BESSMAN^
M.D.
Department of Medicine (HematologyOncology)
University of Texas Medical Branch
Galveston, Texas
References
1. Bessman JD: Heterogeneity of red cell
volume: Quantitation, clinical correlation, and possible mechanisms.
John Hopkins Med J 146:226-230,
1980
2. Bessman JD: Evaluation of whole blood
platelet counts and particle sizing.
Am J Clin Path 74:257-262, 1980
3. Bessman JD, Feinstein DI: Quantitative
anisocytosis as a discriminant between iron deficiency and thalassemia
minor. Blood 53:288-294, 1979
4. Coulter Electronics, Inc, Coulter Counter*
Model S-Plus II Operator's Reference
Manual; Hialeah, Florida, Coulter
Electronics, 1980
5. Hammersley M, King RV, Sullivant
RE, et al: High erythrocyte distribution values and probabilities of
hemoglobinopathies. Am J Clin Pathol
75:370-372, 1981
6. McCurdy PR: Sickle cell trait. N Eng
J Med 282:1158, 1970
The Authors' Reply
In his interesting comments on our
article on red cell distribution, Dr.
Bessman poses several questions.
Hemoglobin SA and CA do refer
strictly to S- or C- trait. We found two
patients doubly heterozygous for sickle
trait and /3-thalassemia minor which we
included in our paper.
Many of these patients had concurrent
iron deficiency anemia and an even
Vol. 76 • No. 2
We have no way of knowing how
many patients with normal RDW's had
abnormal electrophoreses, as our study
only included patients with high RDW's
(over 12.0).
During this time, we had two people,
one a secretary with A-S and a normal
RDW, the other a medical technology
student with A-C and a normal RDW.
The student with A-C had her fiance's
LETTERS TO THE EDITOR
hemoglobin electrophoresed and happily
he was A-A.
The RDW measurement in normal
subjects were:
Mean 10.6 with a
243
only another tool to add to our incidental
detection of hemoglobinopathies.
MARY W. HAMMERSLEY, MT(ASCP)
Standard Deviation of 1.6
As we stated, this is not a primary
method of detecting hemoglobinopathies,
P A U L I. Liu,
M.D.,
PH.D.
Department of Laboratory Medicine
Medical University of South Carolina
Charleston, South Carolina
Fluorescent Nuclear Antibodies
To the Editor:—Your
article "A
comparison of the sensitivities and
specificities of different substrates for
the fluorescent antinuclear antibody
test" by F. Kozin, M. Fowler, and S. M.
Koethe (Am J Clin Pathol 74:785-790,
1980) was of considerable interest to us
as it contrasts with our own experience,
some of which has been previously
published. 3 Referral to the data indicates
a relatively minor difference in frozen
mouse liver, rat liver, and the human
epithelial cell line as supplied by ElectroNucleonics. Our comparative studies
between frozen rat liver and the human
epithelial cell line supplied by ElectroNucleonics, have shown that the human
epithelial cell line is more sensitive than
frozen rat liver in the fluorescent antinuclear antibody test (FANA) utilizing
the identical conjugate on both substrates. We used sera from six patients
with collagen vascular diseases and
tested them simultaneously on both
substrates. These sera were supplied by
Dr. Thomas K. Burnham and had been
previously characterized by pattern.
Three observers viewed the slides and
the titers shown represent a consensus.
More importantly, the human epithelial
cell line was used as the substrate to
detect antinuclear antibodies in 150
patients with various collagen vascular
diseases plus 150 age and sex-matched
controls. We found that 22% of normals
had positive FANA's if serum was
diluted 1:20 as is suggested in the kit
Pattern
Coarsely
speckled
Coarsely
speckled
Nucleolar
Peripheral
Homogeneous
Finely
speckled
Frozen
Rat Liver
Human
Epithelial
Cell Line
1:1280
1:10240
1:80
1:2650
1:640
Neg at 1:20
1:320
1:10240
1:640
1:5120
Neg at 1:20
1:640
supplied by Electro-Nucleonics, whereas
only 4% were positive at serum dilutions of 1:80 or greater. All except
one of 50 SLE patients were positive at
1:80 or greater. This one patient was
also negative at 1:20 but had previously
had a positive FANA on another serum
specimen. Thus the reports that 2% of
normal individuals have positive FANA's
on frozen mouse or rat liver substrates
at serum titers of only 1:42 would also
indicate the increased sensitivity of the
human epithelial cell line of antinuclear
antibodies. Our observations are also in
line with those presented by Hahon
and colleagues who showed rat liver
was less sensitive than all of eight
different cultured cell lines. 1 It has also
been our experience as discussed in the
article by Kozin and associates that the
ease of interpreting patterns is greater
when utilizing the human epithelial cell
line, especially for less experienced
medical technologists.
It is clear physicians must be careful
in interpreting low titer positives on
any substrate. Regardless of which substrate they choose, it is extremely important for each laboratory doing FANA's
to establish it's own meaningful titers
for normal individuals (who may have
low levels of antinuclear antibody) as
well as for diseased patients.
MARY F. LIPSCOMB,
M.D.
Department of Pathology
University of Texas, Southwestern
Medical School
JAMES N. G I L L I A M ,
M.D.
Chairman, Dermatology
University of Texas, Southwestern
Medical School
A N D R E W CHUBICK,
M.D.
Medical Director
Baylor Arthritis Center
Dallas, Texas
References
1. Hahon N, Eckert HL, Stewart J: Evaluation of cellular substrates for antinuclear antibody determinations. J
Clin Micro 2:42, 1975
2. Nakamura RM, Immunopathology clinical
laboratory concepts and methods,
Little, Brown and Company, cl974,
p 261
3. Prystowsky S, Gilliam JN: Antinuclear
antibody studies in chronic discord
lupus erythematosus. Arch Dermatol
113:183, 1977
Anti-Kell
To the Editor:—The recent article
by Molthan and Strohm, 4 raised an
interesting question regarding the
possible inability of LISS procedures
to detect an IgG anti-Kell alloantibody.
However, before this report can be
seriously considered, there are some
points which need clarification.
The authors did not describe the
quality control performed on the original
LISS solution either by the hospital's
transfusion service or by the author's
laboratory. The pH, temperature, and
ionic strength of the LISS solution
significantly affects the rate of the anti-