Question

Lab Activity: Testing for Syphilis with the Arlington Scientific (ASI) RPR test kit

How is a titer performed using this assay kit? Under what circumstances would this be done?

Answer 1

ASI RPR (rapid plasma reagin)  is a nontreponemal flocculation test to detect syphilis which is caused by Treponema pallidum. Infection by this spirochete induces the production of two types of antibodies, the first one is anti-treponemal antibodies which are detected by FTA-ABS antigen and the second is anti-nontreponemal antibodies(reagin) that can be detected by ASI RPR test kit. This ASI RPR is a qualitative and semi-quantitative text in which the reagin antibody binds to the carbon antigen that is a complex of cardiolipin(0.003%), lecithin(0.020–0.022%), cholesterol(0.09%) with activated charcoal which acts as a visual enhancer between reactive and non-reactive results.

The titer is performed in the following manner-

1. using a volumetric pipet dispense a drop of 0.05ml saline on the circles(2 to 5) present on the ASI RPR card( DO NOT SPREAD)
2. dispense a drop of 0.05ml plasma sample on the 1st circle on the test card( DO NOT SPREAD)
3. dispense a drop of 0.05ml test sample on the 2nd circle and resuspend the mixture 5-6 time. Avoid the formation of bubbles
4. transfer 0.05ml of this mixture to the 3rd circle and resuspend properly. in the same way serial dilution is performed by repeating this for 4th and 5th circle. Discard 0.05 ml from this last circle. Following is the dilution series

CIRCLE 1 2 3 4 5

DILUTION 1:1 1:2 1:4 1:8   1:16

5. spread all the dilutions in the test circles starting from the 5th

6. add one drop of carbon antigen to each test circle after agitating the bottle properly to ensure reagent homogeneity. Put the test card on automatic rotator at approx 100 rpm for 8minutes. Tilt the card for 3-4 times and then read results macroscopically in the wet state under high intensity light

RESULT- If the test sample is positive, aggregates will be observed at center or periphery of the test circle ranging from slight to marked and intense. the highest dilution is marked as endpoint titer.

This semi-quantitaive test provides more accuracy than the qualitative test which only depicts the results as reactive or non-reactive accurately at the later stages of the infection.