One drop of freshly prepared antigen is added with a syringe delivering 60 drops from one ml. Theslide is shaken in a VDRL shaker for 4 minutes at 180 rotations. It is then examined under the micros-cope with low power objective. Formation of clumps indicates positive reaction designated asreactive while uniform distribution of crystals in the drop indicates that the serum is non-reactive. Aweakly positive reaction may also occur (weakly reactive).The reactive sera can be diluted and results quantified. VDRL test can also be done in tubes as tubeflocculation test.
Advantages of VDRL test:
a. Simple and rapid testb. Reasonably sensitive and specificc. Requires small quantity of serumd. Can be easily quantifiede. Results are reproduciblef. Reagents are cheap, easily available and have long shelf lifeg. Quality control of tests can be performed.
Biological False Positive Results
In spite of simplicity of test and numerous advantages there are various biological false positive(BFP) results. These can be classified as acute (when they disappear in six months time) or chronicwhen they are positive for more than six months), include the following:Systemic lupus erythematosus, Malaria, Lepromatous leprosy, Infectious mononucleosis, Tropicaleosinophilia, Relapsing fever, Hepatitis, Tissue regeneration, Collagen disorders, Severe trauma,Coronary artery disease, Repeated blood loss, Menstruation, Immunization, Pregnancy, Haemolyticanaemia and Heroin addiction.
Treponema pallidum haemagglutination (TPHA) test is performed by using extract of T. pallidum.Red blood cells are treated to adsorb treponemes on their surface. When mixed with serumcontaining antitreponemal anti bodies, the cells become clumped. This test is similar to FTA-ABS testin specificity and sensitivity, but become positive somewhat later in the infection.Micro haemagglutination -T. pallidum (MHA-TP) is automated version of this test and so also ishaemagglutination treponemal test for syphilis (HATTS). However, these two tests lack sensitivity inprimary syphilis.
Fluorescent antibody test uses an indirect immunofluorescence test in which Nichol's strain of T.pallidum is employed as antigen. The patient's serum is diluted 1:5 and reacted with the antigenwhich has been smeared on the slide. The combination is covered with antihuman immunoglobulinfluorescent conjugate. After proper washing, the smear is examined under fluorescent microscope.In a positive test fluorescent treponemes are observed. When patient serum is diluted to 1:200 insteadof 1:5 (as is done now a days), the test is called as FTA200. Patient's serum is adsorbed with an extractof Reiter's treponemes to avoid false positive results and then the test is performed (FTA-ABS test).This is extremely sensitive and specific test.