Remel RPR Card Test Qualitative Procedure PDF

33 Remel RPR CARD TEST INTENDED USE Remel RPR Card Test is a nontreponemal flocculation test intended for detection of reagin (anti-lipoidal antibodies) in human serum for presumptive serological diagnosis of syphilis when used in conjunction with a treponemal test. SUMMARY AND PRINCIPLE RPR Card...

33 Remel RPR CARD TEST INTENDED USE Remel RPR Card Test is a nontreponemal flocculation test intended for detection of reagin (anti-lipoidal antibodies) in human serum for presumptive serological diagnosis of syphilis when used in conjunction with a treponemal test. SUMMARY AND PRINCIPLE RPR Card Test is a nontreponemal test for serological detection of syphilis,recommended for use when venous blood is collected. RPR Antigen consists of antigens derived from sources not directly associated with Treponemal microorganisms1. 3 In this method, carbon-particle cardiolipin antigen detects reagin, a substance present in sera of syphilitic persons and occasionally in sera of persons with other acute or chronic conditions.4 Reagin is an antibody-like substance produced from the reaction of treponemal microorganisms with body tissue. The detection of reagin in the diagnosis of syphilis. A fourfold decrease in titer following syphilis treatment indicates that treatment has been successful; a fourfold increaseindicates either treatment failure or reinfection. In serum containing reagin, flocculation occurs with agglutination of the carbon particles in the RPR Antigen. Black clumps appear against a whitebackground which can be read macroscopically. In contrast, nonreactive specimens appear to have a uniform light-gray color. PRECAUTIONS This product is for /n Vitro diagnostic use and should be used by properly trained individuals. Precautions should be taken against the dangers of microbiological hazards by properly sterilizing specimens, containers, and test materials after use. Carefully read the entire procedure prior to performing any tests. 1. Potential Biohazardous Material: All human serum should be considered potentially infectious and handled accordingly. Refer to the current edition of Biosafety in Microbiological and Biomedical Laboratories for information on handling human specimens.8 2. RPR Antigen contains thimerosal as a preservative, which may be toxic if ingested. 3. Refer to Material Safety Data Sheet for detailed information on reagent chemicals. STORAGE Store product in its original container at 2-8°C until used. Once the kit is opened, store the RPR Antigen at 2- 8°C and all other kit components in a dry place at room temperature. Once the RPR Antigen is placed in the plastic dispensing bottle it is stable for 3 months or until the expiration date on the kit, provided it is stored at 2-8°C. Do not freeze or expose to temperature extremes. PRODUCT DETERIORATION This product should not be used if (1) the appearance of the reagents has changed, (2) there is evidence of contamination, (3) the expiration date has passed, or (4) there are other signs of deterioration. SPECIMEN COLLECTION AND HANDLING Use serum prepared from whole blood collected without anticoagulant. Allow blood to fully clot before centrifuging. Serum should be clear and separated from cells as soon after collection as possible. Hemolyzed specimens are not acceptable for testing when printed matter cannot be read through them.  34 MATERIALS AND REAGENTS PROVIDED Tests/Kit Components: 150 500 RPR Antigen: 0.003%Cardiolipin, 0.09% Cholesterol, 1 x3 ml 3x3 ml 0.021% Lecithin, 0.0125 M EDTA, 0.01 M Na2HP0 4, 0.01 M KH2P0 4, 0.01875% Charcoal, 0.1% Thimerosal (preservative), 10.0% Choline Chloride, w/v, Demineralized Water Plastic Dispensing Bottle 1 each 1 each 20 Gauge, Galvanized Needle, Blunt Cut 1 each 1 each White. 10 well Test Cards 15 each 50 each Pioette/Stirrers. 50 ul 150 each 500 each REF Number R16302 R16303 MATERIALS REQUIRED BUT NOT SUPPLIED (1) Mechanical rotator with fixed speed or adjusted to 100 rpm, circumscribing a ¾" circle, (2) Humidifier cover with moistened blotter or sponge, (3) 1 ml pipette, (4) High intensity incandescent lamp, (5) RPR Liquid Controls {REF R16307) or suitable alternative, (6) Calibrated pipette, 100 µI, 50 µI, (7) Test lubes (12 X 75), (8) Normal saline (0.85%), (9) Human serum, nonreactive for syphilis (for quantitative procedure). PROCEDURE Preparation of Reagents: Check the delivery of the needle by placing it firmly on a syringe. Fill the pipette with 1ml of water. Hold it in a vertical position and dispense 1.0 ml of water while simultaneously counting the drops. Aneedle delivery rate of 60 drops +/- 2 drops is acceptable. If the needle does not meet this specification it should not be used. Allow RPR Antigen to equilibrate to room temperature. Gently shake the bottle for 10-15 seconds to resuspend the antigen. Attach the needle (provided) to the tapered fitting on d i s p e n s i n g b o t t l e. Upon completion of the daily tests, remove the needle from the dispensing bottle and rinse with demineralized water. This will help maintain clear passage of the suspension. Do not wipe the needle as this may affect the accuracy of the antigen drop as it is dispensed. Allow RPR Antigen, RPR Liquid Controls, and test specimens to equilibrate to room temperature prior to use. The temperature of the room and all test materials, including specimens, must be maintained in the range of 23-29°C. Temperatures below this range will cause false-negative reactions and lower titers, while temperatures above the range have the opposite effect.7 Verify the speed of the mechanical rotator (100 +/- 2 rpm) to ensure reproducible results. Handle test cards by grasping the edge of the card; do not touch the surface of the test wells. RPR controls with established patterns of reactivity should be included in each test run. RPR Liquid Controls are available under a separate reference number (R16307).  35 Qualitative Test Procedure: 1. Label the test wells on the card with the specimen identification 2. Use a separate pipette/stirrer for each test specimen or control. Pre- squeeze the pipette/stirrer and draw up the specimen or control. Dispense 1 free-falling drop (50 µI) into the appropriate well. 3. Using the opposite, flattened end of the pipette/stirrer gently spread the specimen or control over the entire circle using a circular motion. 4. Gently shake RPR Antigen suspension in the dispensing bottle. Holding the bottle in a vertical position, dispense several drops into the cap to verify the needle passage is clear. Dispense 1 free-falling drop into each well containing specimen or control. Do not stir; mixing of the antigen suspension and the sample is accomplished during rotation. 5. Immediately place the test card on the mechanical rotator, cover withthe humidifier cover, and rotate for 8 minutes at 100 rpm. Note: False-positive reactions may occur due to evaporation if samples are not properly covered during rotation. 6. Following the 8-minute rotation, briefly rotate and tilt the card back and forth by hand 3-4 x to aid in differentiating nonreactive from minimally reactive results. Immediately read the card macroscopically in the wet state under a high intensity incandescent lamp. Avoid glare when reading reactions. Reading Report Small to large clumps (R) or slight but definite clumps (Rm) Reactive (R) No clumping or very slight roughness Nonreactive (NR) Note: All specimens with a reactive or rough reaction in the Qualitative Test should be tested according to the Quantitative Test procedure to provide a baseline from which changes can be determined, particularly for evaluating the efficiency of treatment\. Initial reports should only be made on specimens that are nonreactive. QUALITY CONTROL RPR controls with established patterns of reactivity should be included in each test run. A test run can be defined as a period of approximately 24 hours. Use reactive, minimally reactive, and nonreactive controls and test according to the Qualitative Test procedure. Remel RPR Liquid Controls can be obtained under a separate reference number (REF R16307). Each laboratory should establish endpoint titers for the quantitative controls used. If controls do not perform as expected, patient results should not be reported. Quality control testing should be performed according to established laboratory quality control procedures following the guidelines and recommendations of applicable federal, state, and local regulatory agencies. LIMITATIONS OF PROCEDURE 1. A diagnosis of syphilis should not be based on a single reactive serologic test. All historical information, clinical findings, and laboratory results should be taken into consideration. · 2. Biologic false-positive (BFP) reactions occurring with nontreponemal tests may be acute or chronic. Acute false- positives (titers lasting 6 months) have been observed with connective tissue diseases (e.g., systemic lupus erythematosus), narcotic addiction, aging, leprosy, and malignancy. Unusually high false-positive titers may be seen in patients diagnosed with lymphoma. 3. The RPR test cannot be used to test cerebrospinal fluid.