RAPID PLASMA REAGIN (RPR) TEST RPR is use in the non-treponemal flocculation test for the qualitative and semiquantitative determination of regain antibodies in serum or plasma Materials: RPR Antigen: Suspension of carbon approximately 0.2 g/L, 0.003% Cardiolipin, 0.02% Lecithin and 0.09% Cholesterol. Positive Control: Serum containing antibodies against Treponema pallidum. Negative Control: Serum free of antibodies against Treponema pallidum. Stirrers Disposable test slides Micropipettes capable of dispensing 50 ul and 16 ul Disposable serum pipets Test tubes Test cards Rotator (set at 100 rpm) Isotonic saline: 0.9% NaCl Principle: RPR is a modified form of VDRL Antigen which contains carbon particles to improve the visual reading of the result. When binding occurs between cholesterol / cardiolipin / lecithin in the reagent and the reagin antibodies in the sample, the results can be seen macroscopically in the form of black clumps. No visual flocculation indicates a negative result. Procedure: Qualitative Method: 1. Dispense one drop (50 ul) of patient sample onto a glass slide or RPR Test Card. 2. Spread the sample over the entire area of the test circle using the flat end of the pipette / stirrer. 3. Using the dispensing bottle / needle assembly allow one free – falling drop (16 ul) of antigen to drop onto the test specimen. Do not restir. 4. Rotate the test card at 100 RPM for 8 minutes only on an automatic rotator. 5. Immediately after the 8 minutes inspect the result visually in good light. Semi Quantitative Method: 1. Using isotonic saline prepare serial dilutions (50 ul) of the patients serum (1:2, 1:4, 1:8, 1:16, 1:32, 1:64 and so on) 2. Transfer one drop (50 ul) of each serum dilution to the test circle on the slide. 3. Add one drop (16 ul) of shaken antigen to the sample. (There is no need to mix these two components) 4. Rotate the slide / cards for 8 minutes at 100 rpm 5. Immediately after the 8 minutes, inspect the result visually in good light. Results and Interpretation: Qualitative Method: Medium and large aggregates Reactive Finely dispersed aggregates Weak – Reactive No aggregates visible, smooth grey appearance Non – Reactive Semi – Quantitative Method: The titer of the last dilution that produces a reactive result. Questions: 1.Why is a qualitative test performed before a semi-quantitative test? 2. Why is it necessary to rotate the slide/card?