JOURNAL OF CLINICAL MICROBIOLOGY, Aug. 1982, p. 286-290 Vol. 16, No.

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0095-1 137/82/080286-05$02.00/0

Evaluation of a New Rapid Plasma Reagin Card Test as a

Screening Test for Syphilis
MARY W. PERRYMAN,'* SANDRA A. LARSEN,' EDITH A. HAMBIE,' DEBORAH E. PETTIT,'
REBECCA L. MULLALLY,I AND WILLIAM WHITTINGTON2
Sexually Transmitted Diseases Laboratory Program, Center for Infectious Diseases, 1 and Venereal Diseases
Control Division, Center for Prevention Services,2 Centers for Disease Control, Atlanta, Georgia 30333
Received 22 February 1982/Accepted 6 May 1982

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This study evaluates the American Dade (Biokit Laboratories) rapid plasma
reagin (Dade RPR) card test, currently used in Spain for the diagnosis of syphilis,
which has been recently released to the U.S. market. Used as a basis for
comparison with the Dade card test were the 18-mm standard rapid plasma reagin
(standard RPR) card test and the Venereal Disease Research Laboratory (VDRL)
slide test, using both fresh sera obtained from 505 individuals and paired serum-
plasma specimens from 174 individuals. Results obtained proved the Dade RPR
card test with serum to be very similar to the standard RPR card test; sensitivity
was 92.3% and specificity was approximately 99% for both RPR card tests.
Although the sensitivity of the VDRL slide test was lower at 88.5%, its specificity
was also approximately 99%. Quantitatively, the agreement 1 dilution between
±

the two card tests was 93.6%. Agreement 1 dilution between the Dade RPR card
±

test and the VDRL slide test was 46.5% for sera, comparable to the standard RPR-
VDRL agreement of 50%. In the limited evaluation of the RPR card tests with
plasma, the specificity was 99.4% and the sensitivity was 100% for both tests.
Quantitative agreement 1 dilution between plasma and serum pairs was 100%
±

for the Dade RPR card test. Our results showed that the Dade RPR card test is as
sensitive and as specific as the standard RPR card test. Therefore, it was
concluded that one card test has no particular advantage over the other.

The standard rapid plasma reagin (standard
RPR) 18-mm circle card test developed by Port-
noy was one of the first modifications of the
Venereal Disease Research Laboratory (VDRL)
test (3). By addition of EDTA to stabilize the
VDRL antigen, choline chloride to eliminate
heat inactivation of the serum, and sized char-
coal particles to allow the technician to visualize
the antigen-antibody reaction, the VDRL test
was modified to become the RPR card test (4).
Although initial studies evaluated the use of the
RPR card antigen with plasma (4), thus suggest-
ing the name, the RPR card test has been
accepted as a standard test for syphilis only with
sera, but not with plasma.
Today in the United States, the RPR card test
manufactured by Hynson, Westcott and Dun-
ning is the most commonly used screening test
for syphilis. Some of the reasons for the popular-
ity of the RPR card test are as follows. (i) The
sensitivity and specificity of the RPR card test
are equal to or better than those of the VDRL
test (1, 3, 4, 5). (ii) The RPR card test provides
serological support for the diagnosis of syphilis
at the time the patient is still in the clinic (6). (iii)
Materials required for RPR card testing are easy
to use, inexpensive, and expendable. Test re-
sults are read with the naked eye (1). (iv) The
RPR card test can be performed in the clinic or
in the field and allows expeditious, large-scale
testing (3). Recently, other manufacturers have
introduced reagents similar to the product by
Hynson, Westcott and Dunning (2). The purpose
of this study was to compare a newer RPR (Dade
RPR) card test manufactured by Biokit Labora-
tories, Barcelona, Spain, and distributed by
American Dade, Division of American Hospital
Supply Corporation, Miami, Fla., with the stan-
dard RPR card test, using both plasma and sera
for the serodiagnosis of syphilis. For additional
comparison, all sera were also tested in the
VDRL slide test, and all nontreponemal test
reactivity was confirmed with the fluorescent
treponemal antibody absorption (FTA-ABS)
test.
MATERIALS AND METHODS
Specimens. Blood was obtained from 505 persons
who attended the DeKalb County (Ga.) Sexually
Transmitted Disease Clinic. A sample of blood from
174 of these 505 persons was immediately dispensed
from a 7-ml Vacutainer tube without anticoagulant

286
VOL. 16, 1982
TABLE 1. Qualitative agreement among
nontreponemal tests
No. of specimens with indicated
reaction by following test':
Test and reactiona VDRL stide Dade RPR
card
N R W N
Standard RPR card
N 449 1 5 452
R 6 39 5 2
VDRL slide
N 449 6 flocculation from slight to intense with the RPR card
R 1 39
W 4 6 degree of flocculation or only a slight roughness. The
a N, Nonreactive; R, reactive; W, weakly reactive
(considered reactive in tabulations).
b Agreement was 99.0% between the standard and
Dade RPR card tests, 97.8% between the VDRL slide
and Dade RPR card tests, and 97.6% between the
standard RPR card and VDRL slide tests.
used to originally obtain the blood to a 4-ml Vacutainer
tube containing 6.0 mg of EDTA in the potassium
form. All blood samples were centrifuged, and the
serum or plasma was separated from the cells and
tested within 18 h unless otherwise indicated. Samples
of sera were dispensed to be heated for testing in the
VDRL slide test or FTA-ABS test or both. Histories
were obtained on all individuals whose serum or
plasma exhibited any degree of reactivity in any of the
nontreponemal tests.
Reagents. Antigen for the standard RPR card test
was supplied by the Biological Products Program,
Center for Infectious Diseases, Centers for Disease
Control, Atlanta, Ga. A single lot of antigen was used
throughout the study. Brewer diagnostic cards for the
standard RPR card test, needles, dispensing vials, and
stirrers were supplied by Hynson, Westcott and Dun-
ning.
The Dade RPR card antigen was supplied as part of
a kit containing two 5-ml ampules of antigen; one vial
each of reactive, moderate to weakly reactive, and
nonreactive control sera; plastic-coated Dade RPR
test cards; disposable plastic capillary tubes; rubber
bulbs; wooden stirrers; a dispensing vial; and a 20-
gauge needle. A single lot of antigen, 2681, was used
during the study.
The VDRL slide and FTA-ABS test reagents and
control sera for these tests, as well as those for the
standard RPR card test, were reference reagents fur-
nished by the Biological Products Program.
Testing. Each nontreponemal test was performed
daily on serum or plasma by a technician who had no
knowledge of the results of the other reagin tests. Test
assignments were rotated among the technicians to
eliminate test preference bias. Quantitative nontrepo-
nemal tests were performed on any serum or plasma
exhibiting any degree of reactivity in the qualitative
test. The FTA-ABS test was performed on any single
serum or serum from a plasma pair which was reactive
in any of the nontreponemal tests. The VDRL slide,
standard RPR card, and FTA-ABS tests were per-
NEW RPR CARD TEST 287
formed according to the 1969 Manual of Tests for
Syphilis. The Dade RPR card test was performed
according to the directions of the manufacturer. Brief-
ly, for the qualitative test, 0.05 ml of serum or plasma
and 1 drop of antigen were placed in a single 18-mm
circle of a 10-place, plastic-coated card. The serum
and antigen were mixed and spread with a stirrer
R within the circle. The card, when filled, was placed
under a humidifier cover and rotated for 8 min at 100
rpm. Immediately after mechanical rotation, the card
3 was briefly rotated manually and read under a high-
48 intensity, incandescent light. The readings were re-
ported as either "reactive" or "nonreactive." Reac-
tive indicated a specimen showing any degree of
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antigen. Nonreactive indicated a specimen giving no
quantitative test was reserved for sera exhibiting any
degree of reactivity or roughness. To perform the
quantitative test, 0.05 ml of saline was placed in the
second through fifth circles of the card, and then 0.05
ml of serum was placed in each of the first two circles.
Serial twofold dilutions were prepared in the second
through fifth circles. Then, the Dade RPR card antigen
was dropped next into the diluted serum or plasma,
and the specimen plus antigen was mixed and spread
within the circle. The remainder of the quantitative
procedure was exactly the same as for the qualitative
procedure. Results for the quantitative test were re-
ported as the highest dilution of serum or plasma
giving any degree of reactivity.
RESULTS
The Dade RPR card is similar to the Brewer
diagnostic card except for the color of the cir-
cles. The Dade RPR card has Kelly-green cir-
cles, which may aid in providing a contrast to
the gray charcoal color of the antigen. The Dade
antigen is similar in color and texture to the
standard RPR card antigen.
The qualitative comparisons of the results
from the 505 sera tested in the Dade and stan-
dard RPR card tests and the VDRL slide test are
shown in Table 1. Qualitative agreement be-
tween the Dade and standard RPR card tests
was 99.0%. If the VDRL slide test results that
were weakly reactive were considered as reac-
tive, then agreement between the Dade RPR
card test and the VDRL slide test was 97.8%,
whereas agreement between the standard RPR
card test and the VDRL slide test was 97.6%.
Fifty-seven sera were reactive in at least one
of the nontreponemal tests. The distribution of
the reactive sera among the three nontrepone-
mal tests and the FTA-ABS test is shown in
Table 2. The agreement of the FTA-ABS test
results with the standard RPR card test, the
Dade RPR card test, and the VDRL slide test
was 87.0% (47 of 54 results in agreement) for
both RPR card tests and 77.8% (42 of 54 results
in agreement) for the VDRL slide test. The three
sera giving borderline FTA-ABS test results
were not entered into tabulation. The diagnoses
288 PERRYMAN ET AL. J. CLIN. MICROBIOL.
TABLE 2. Distribution of reactive sera among the for the standard RPR card test, and 98.9% for
three nontreponemal tests and the FTA-ABS test the VDRL slide test.
No. of sera Quantitative agreement among the three non-
Nontreponemal test with following treponemal tests is shown in Table 4. The results
pattern of reactivity No. of result by FTA- of only those sera which were reactive in both of
RPR card test
serawith
pattem
ABS test': the tests compared are tabulated. Agreement +
1 dilution (dil) between the two RPR card tests
was 93.6%. Again, assuming weakly reactive
Dade Stan-
dard slide R B N
VDRL slide test readings as equivalent to reac-
R R R 43 38 2 3 tive 1 dil, then agreement 1 dil between the
±

N R R 1 1 standard RPR card test and the VDRL slide test

N N R 4 2 2 was 50%, and agreement between the Dade RPR
N R N 1 1 card test and the VDRL slide test was 46.5%.

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R R N 5 5 Of the 174 plasma and serum pairs tested,
R N N 1 1 qualitative agreement between the results with
R N R 2 1 1
a R, Reactive; B, borderline; N, nonreactive.
associated with reactive nontreponemal tests are
shown in Table 3. The sensitivity of all three
nontreponemal tests, based on the 21 untreated
cases of syphilis, was 95.2% (20 of 21 results in
agreement). With the 31 sera from individuals
with treated syphilis, both the Dade and stan-
dard RPR card tests were 90.3% reactive (28 of
31 results in agreement). On the the other hand,
the sensitivity of the VDRL slide test with
treated cases was 83.9% (26 of 31 results in
agreement). Therefore, overall sensitivity, i.e.,
treated and untreated, was 92.3% for both RPR
card tests and 88.5% for the VDRL slide test.
Comparative specificities of the three non-
treponemal tests based on the 453 sera drawn
from individuals presumed not to have syphilis
were 99.3% for the Dade RPR card test, 99.6%
plasma and serum was 98.9% for the standard
RPR card test and 99.4% for the Dade RPR card
test. Agreement between the two RPR card tests
when plasmas were tested was 100%, whereas
agreement between the two tests for the 174 sera
was 99.4%. In the standard RPR card test, 14
specimens were reactive and 158 specimens
were nonreactive when either plasma or serum
was used; 2 specimens were nonreactive for
serum and reactive for plasma. In the Dade RPR
card test, 15 specimens were reactive and 158
specimens were nonreactive when either plasma
or serum was used; 1 specimen, drawn from an
individual diagnosed as having treated second-
ary syphilis, was nonreactive for serum and
reactive for plasma. A total of 16 specimens
were reactive and 158 specimens were nonreac-
tive for plasma by either test, whereas 15 speci-
mens were reactive and 158 specimens were
nonreactive for serum by either RPR card test.
One specimen was reactive for serum by the

TABLE 3. Diagnoses associated with sera reactive in nontreponemal tests

No. of sera with indicated reaction by following testa:
RPR card test
Stage of syphilis VDRL slide FTA-ABS
Dade Standard
R N R N R W N R B N
Primary
Untreated 5 1 6 4 2 6
Treated 7 1 7 1 5 1 2 8
Secondary
Untreated 6 6 6 6
Treated 5 5 5 5
Latent
Untreated 9 8 1 8 1 8 1
Treated 15 1 15 1 10 3 3 14 2
Past history of syphilis 1 1 1 1 2 2
Other sexually transmitted diseases 3 2 2 3 3 2 2 3
a R, Reactive; N, nonreactive; W, weakly reactive (considered reactive in tabulations); B, borderline.
VOL. 16, 1982 NEW RPR CARD TEST 289
TABLE 4. Quantitative comparisons of the three nontreponemal tests
No. of indicated type of reaction (dil) obtained by following
RPR card test':
Comparison Dade Standard
--2 -1 Equal +1 . +2 .-2 -1 Equal +1 - +2
Dade and standard RPR card tests 3 11 29 4
Dade RPR card and VDRL slide tests 1 8 11 23
Standard RPR card and VDRL slide tests 7 15 22
a Agreement ± 1 dil was 44 of 47 results in agreement (93.6%) between the Dade and standard RPR card tests,
20 of 43 results in agreement (46.5%) between the Dade RPR card and VDRL slide tests, and 22 of 44 results in
agreement (50%o) between the standard RPR card and VDRL slide tests.

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Dade test and nonreactive for serum by the
standard test. The diagnoses for the 16 reactive
plasmas are shown in Table 5. Both RPR card
tests were 100% sensitive with plasma. The
specificity of the card tests with plasma was
99.4%, based on the 159 specimens from individ-
uals without syphilis.
Quantitative agreement + 1 dil (Table 6) be-
tween the plasma and serum pairs within the
same test was 92.9% for the standard RPR card
test and 100% for the Dade RPR card test. The
between-test quantitative agreement with plas-
ma was 93.8% 1 dil.
± repeatable results + 1 dil, some increased nega-
All of the above plasma results are based on
results of tests performed on plasma which had
been separated from the cellular components of
the blood by centrifugation. In addition to these
tests, 33 of the plasmas were tested after 18 h at
25°C but before centrifugation and separation
from the cellular material. The plasma portion of
the unspun blood contained fibrous material that
was avoided when transferring the plasma to the
card. Of these plasmas, 32 gave results that were
identical, even to titer, to those obtained on the
plasma after centrifugation, for a repeatability of
97%. One plasma was reactive 1 dil in the Dade
RPR card test before centrifugation but nonreac-
tive in both RPR card tests after centrifugation.
The serum pair of the plasma was nonreactive in
all three nontreponemal tests and the FTA-ABS
test.
Two groups of centrifuged separated plasmas
were frozen after initial testing. One group of 39
specimens was frozen for 1 week. Upon thaw-
ing, these plasmas were clear in appearance.
Although retesting of the specimens gave 100%
tive roughness was noted in both card tests. Five
plasmas were read as negative rough with the
standard RPR card test, whereas three were
read with this result in the Dade RPR card test.
The second group of 33 specimens was retested
after 3 weeks of storage at -20°C. A fibrous
precipitate was seen in these plasmas. The
amount of fibrous material transferred to the
card was proportional to the amount of distur-
bance of the fibrous clot. Negative roughness
was especially increased with the Dade RPR

TABLE 5. Diagnosis associated with plasmas reactive in the RPR card tests
No. of specimens with indicated reaction by following test':
RPR Card Test
Stage of syphilis FTA-ABS
Dade Standard
R N R N R B N
Primary
Untreated 1 1 1
Treated 3 3 3
Secondary
Untreated 2 2 2
Treated 2 2 2
Latent
Untreated 3 3 3
Treated 4 4 4
Other sexually transmitted diseases 1 1
a R, Reactive; N, nonreactive; B, borderline.
290 PERRYMAN ET AL. J. CLIN. MICROBIOL.
TABLE 6. Quantitative comparison
No. of indicated type of reaction (dil)a
Comparison Serum Dade RPR card test
-2 -1 Equal +1 -2 -1 Equal +1
Serum and plasma in standard RPR card test 1 2 9 2
Serum and plasma in Dade RPR card test 7 6 2
Plasma in Dade and in standard RPR card tests 1 2 8 5
a Agreement ± 1 dil was 13 of 14 results in agreement (92.9%) between serum and plasma in the standard RPR
card test, 15 of 15 results in agreement (100%) between serum and plasma in the Dade RPR card test, and 15 of 16
results in agreement (93.8%) between plasma in the Dade and in the standard RPR card test.

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card test. Repeatability for the Dade card test
was 100%, whereas that for the standard RPR
card test was 97%. One sample, which had been
reactive 1 dil in both RPR card tests with the
fresh plasma, was reported as nonreactive in the
standard test on repeat after freezing. This was
the same plasma whose serum pair had been
nonreactive in both the standard RPR card and
the Dade RPR card tests.
DISCUSSION
Our evaluation indicates the Dade RPR card
test to be as sensitive and as specific as the
standard RPR card test for the serodiagnosis of
syphilis. Since one card test has no particular
advantage over the other, the choice of product
will be a matter of personal preference.
In a previous evaluation of the use of EDTA
plasma in the standard RPR card test, we found
that the plasma may be stored at either 25 or 4°C
for up to 24 h without an increase in negative
roughness. Results from the current study with
plasma indicated that (i) when testing plasma
which has not been centrifuged or separated or
both from the cellular constitutents, care must
be taken to avoid the fibrous material in the
plasma portion; and (ii) separated plasma may
be frozen for up to 1 week without the formation
of an additional fibrous clot. Only plasma from
blood drawn with EDTA as the anticoagulant
has been evaluated in the RPR card tests; there-
fore, we caution against the testing of plasma
when the anticoagulant used is other than EDTA
or is unknown. The use of plasma to follow the
efficacy of treatment or in the treponemal tests
has not been evaluated. If a plasma is reactive in
the RPR card test and confirmation is necessary,
then we suggest that a serum be drawn, the
quantitative RPR card test be repeated with the
serum, and a treponemal test be performed.
LITERATURE CITED
1. Falcone, V. H., G. W. Stout, and M. B. Moore, Jr. 1964.
Evaluation of rapid plasma reagin (circle) card test. Public
Health Rep. 79:491-495.
2. March, R., and G. E. Stiles. 1980. The reagin screen test: a
new reagin card test for syphilis. Sex. Transm. Dis. 7:66-
70.
3. Portnoy, J. 1963. Modification of the rapid plasma reagin
(RPR) card test for syphilis for use in large scale testing.
Am. J. Clin. Pathol. 40:473-479.
4. Portnoy, J., W. Garson, and C. A. Smith. 1957. Rapid
plasma reagin test for syphilis. Public Health Rep. 72:761-
766.
5. Reed, E. L. 1965. The rapid plasma reagin (circle) card test
for syphilis as a routine screening procedure. Public Health
Lab. 23:96-103.
6. Tiedemann, J. H., and J. Mullins. 1966. The RPR card test
as a diagnostic aid in a venereal disease clinic. Public
Health Lab. 24:12-15.