Rapid Plasma

Reagin(RPR) Test

Ram Prashad Ojha

Introduction

 The RPR test is a nontreponemal testing

procedure for the serologic detection of
syphilis.
Principle of RPR
 The RPR Card antigen suspension is a carbon
particle containing cardiolipin antigen that
detects reagin.

 Reagin is an antibody like substance present in

serum or plasma from individuals with syphilis.

 The reagin binds to the test antigen which

consists of cardiolipin-lecithin coated particles
that cause macroscopic flocculation.
Cont…
 When a specimen such as serum or plasma
contains antibody, flocculation occurs with
the resulting aggregation of the carbon
particles.

 The flocculation appears as black clumps

against the white background of the plastic
coated card.
Cont…
 Antibodies associated with syphilis begin to
appear in the blood 4 to 6 weeks after
infection.

 Nontreponemal tests determine the presence

of reagin.

 Reagin is a nontreponemal autoantibody

directed against cardiolipin antigens.
Materials for RPR
 RPR Test Cards
 RPR Controls(Reactive and Non-Reactive)
 RPR Antigen

◦ (cardiolipin-lecithin-cholesterol,choline
chloride,EDTA & carbon particle)
 Dispenstirs
 Rotator
Reactions for Controls

The following reactions should be observed to

compare against the test results:
 Reactive control - characteristic strong

clumping.
 Reactive moderate control - moderate

clumping.
 Non-reactive control - smooth, grayish

appearance of unclumped particles

Specimen Collection
 Unheated serum- centrifuge for
sedimentation of cellular elements, serum
may be frozen until time of testing.

 Unheated Plasma - specimen should be

collected with an anticoagulant such as EDTA
or heparin, plasma must be stored at 2°C to
8°C. Plasma must be tested within in 24 hrs
of collection.
 Theaddition of choline chloride,(block
inhibitors in serum,eliminate the heat
inactivation & allow testing of plasma)

 EDTA(stabilizethe antigen &allow it to be

used for upto 6month at 4-10◦c)
Procedure for Test
1. Label rings on test card with numbers of
samples to be tested
2. Use Dispenstir to draw up serum sample.
3. Hold Dispenstir in a perpendicular position
directly over the test circle to which the
specimen is to be delivered.
4. Squeeze Dispenstir to allow 1 drop to fall on
to each circle.
5. Spread the specimen in the confines of the
circle.

6. Reconstitute the antigen bottle, by shaking.

Holding the bottle in a straight vertical position,
place one “free falling” drop on each test area.
7. Rotate card for 8 minutes on a mechanical
rotator at 100 rpm.

8.After rotating mechanically, the test card

should be rotated manually by hand 3 to four
rotations and then read immediately
macroscopically in the “wet” state under a high
intensity lamp.
 The test results should be reported as
reactive (even if minimally reactive) or non-
reactive.

 All test results that are reactive should be

quantitated.
Causes of Negative test result

1. The patient does not have syphilis.

2. The infection is too recent for antibodies to
be produced. (Repeated tests should be
administered at 1 week, 1 month, and 3
month intervals to establish presence or
absence of disease).
3. The syphilis is latent or inactive
4. Faulty immunodefense mechanism
5. Faulty lab techniques
 A positive reaction is not conclusive for
syphilis. Several conditions produce biologic
false positive results for syphilis. (False
positive means that the test revealed a
positive reaction when it was actually
negative).
 False positives may reveal the presence of

other serious diseases.

Non-syphilitic Conditions Giving
Biologic False-Positive Results
 Malaria
 Leprosy
 Relapsing fever
 Infectious Mononucleosis
 Atypical pneumonia
 Viral pneumonia
 Lupus erythematosus
 Measles
 pregnancy
 Lymphoma
 Tuberculosis
 Connetive tissue disease
 Endocarditis
 Drug abuse
 Contamination
Resolving False Positive RPR Tests

 False positive RPR tests may be resolved by

testing the patient’s serum with a specific
treponemal antigen test.
Conformation of result
 Diagnosis of syphilis requires correlation of
geographical area or country, patient history, physical
findings, and results of syphilis antibody tests.

 T. pallidum is diagnosed when both the screening and

confirmatory test are reactive.

 Treponemal tests are more specific than non-

treponemal tests

 Treponemal tests confirm syphilis when a reactive or

positive non-treponemal result is obtained.
Confirmatory Tests for Syphilis
 FTA-ABS (Fluorescent Antibody Absorbed
test)
 TPHA (T.pallidum Haemaggultination Assay)
 EIA (Enzyme Immunoassay)
 Treponemal tests are used to confirm reactive
non-treponemal procedures.

 A positive FTA-ABS test almost always

remains positive and therefore is not
recommended for monitoring therapy.
Theory of Antigen-Antibody Reaction
 It is theorized that reagin is an antibody
against tissue lipids.
 Lipids are presumed to be liberated or free

from body tissue in the normal course of

activity.
 As a result of being free in the body, these

lipids may induce antibody formation.

Interfering Factors
 Alcohol decreases reaction intensity in tests
and therefore should be avoided for a least
24 hrs before blood is drawn.
 Avoid drawing the blood sample immediately

after the patient has eaten.

Clinical Information
 Sexual partners of patients with syphilis
should be evaluated for the disease.
 After treatment, patients with early stage

syphilis should be tested at 3 month intervals

for one year to monitor declining reactivity of
the syphilis.
Summary
 RPR (Rapid Plasma Reagin) is Non specific (screening) test
for syphilis and detect reagin antibody present in serum.
 It is modification of VDRL in which cardiolipin Ag is absorb

on finely divided carbon particles, which are suspended in

choline chloride which blocks inhibitory factor in serum,
thus eliminating the need of heating serum sample before
testing.
 The Ag gets clump and flocculated with serum containing

Reagin antibody.
 Report:

No clump- Non reactive

Clump - Reactive with titer (eg: Reactive, titer1:8)
Thank You…….