Intended Use
Novabios Leishmania Ab Test is a rapid chromatographic immunoassay for the qualitative detection of antibodies  to the subspecies of the Leishmania donovani (L.donovani), the Visceral leishmaniasis causative protozoans, in Whole Blood /Serum / Plasma.
It is intended to be used as a screening test and as an aid in the diagnosis of the disease of Visceral leishmaniasis.
Visceral leishmaniasis, or Kala-azar, is a disseminated infection caused by several subspecies of the L.donovani. The disease is estimated by the World Health Organization (WHO) to affect approximately 12 million people in 88 countries1. It is transmitted to humans by bites of the Phlebotomus sandflies, which acquire infection from feeding on infected animals. Though it is a disease found in poor countries, in Southern Europe, it has become the leading opportunistic infection in AIDS patients2-3.
Identification of L. donovani organism from the blood, bone marrow, liver, lymph nodes or the spleen provides a definite mean of diagnosis. Serological detection of anti-L. donovani IgM is found to be an excellent marker for the acute Visceral leishmaniasis. Tests used in clinic are included ELISA, fluorescent antibody or direct agglutination tests 4-5. Recently, utilization of L. donovani specific protein in the test has improved the sensitivity and specificity dramatically6-7. One step Leishmania Ab Test is a simple, visual qualitative test that detects L. donovani antibodies in human Whole Blood/serum/plasma. The test is based on immunochromatography and can give a result within 15 minutes.
 
Principle
Novabios Leishmania Ab Test is a qualitative membrane strip based immunoassay for the detection of L. donovani antibodies (IgG and IgM) in Whole Blood /Serum / Plasma. The test device consists of: 1) a burgundy colored conjugate pad containing L. donovani recombinant envelope antigens conjugated with Colloid gold (L. donovani conjugates) and rabbit IgG-gold conjugates,2) a nitrocellulose membrane strip containing two test bands (T1 and T2 bands) and a control band (C band). The T1 band is pre-coated with the antibody for the detection of IgG anti- L. donovani, T2 band is coated with antibody for the detection of IgM anti- L. donovani, and the C band is pre-coated with goat anti rabbit IgG. When an adequate volume of test specimen is dispensed into the sample well of the test cassette, the specimen migrates by capillary action across the cassette. IgM anti- L. donovani, if present in the specimen, will bind to the L. donovani conjugates. The immunocomplex is then captured by the reagent pre-coated on the T2 band, forming a burgundy colored T2 band, indicating a Leishmania IgM positive test result and suggesting a fresh infection. IgG anti- L. donovani if present in the specimen will bind to the L. donovani conjugates. The immunocomplex is then captured by the reagent coated on the T1 band, forming a burgundy colored T1 band, indicating a Leishmania IgG positive test result and suggesting a recent or repeat infection. Absence of any T bands (T1 and T2) suggests a negative result. The test contains an internal control (C band) which should exhibit a burgundy colored band of the immunocomplex of goat anti rabbit IgG/rabbit IgG-gold conjugate regardless of the color development on any of the T bands. Otherwise, the test result is invalid and the specimen must be retested with another device.
 
Storage and Stability
·            Store as packaged in the sealed pouch at room temperature or refrigerated (4-30℃ or 40-86℉). The test device is stable through the expiration date printed on the sealed pouch.
·            The test must remain in the sealed pouch until use.
 
Additional Special Equipment
Materials Provided
Ÿ    Test devices                         Ÿ    Disposable specimen droppers   
Ÿ    Buffer                              Ÿ    Package insert
Materials Required But Not Provided
Ÿ    Timer                                Ÿ    Centrifuge
Ÿ    Specimen collection containers        
 
Precautions
·            For professional in vitro diagnostic use only. Do not use after expiration date.
·            Do not eat, drink or smoke in the area where the specimens and kits are handled. 
·            Handle all specimens as if they contain infectious agents.
·            Observe established precautions against microbiological hazards throughout all procedures and follow the standard procedures for proper disposal of specimens.
·            Wear protective clothing such as laboratory coats, disposable gloves and eye protection when specimens are assayed.
·            Follow standard biosafety guidelines for handling and disposal of potential infective material.
·            Humidity and temperature can adversely affect results.
 
Specimen Collection and Preparation
1.          Novabios Leishmania Ab Test can be performed used on Whole Blood /Serum / Plasma.
2.          To collect whole blood, serum or plasma specimens following regular clinical laboratory procedures.
3.          Testing should be performed immediately after specimen collection. Do not leave the specimens at room temperature for prolonged periods. For long term storage, specimens should be kept below -20℃. Whole blood should be stored at 2-8℃ if the test is to be run within 2 days of collection. Do not freeze whole blood specimens.
4.          Bring specimens to room temperature prior to testing. Frozen specimens must be completely thawed and mixed well prior to testing. Specimens should not be frozen and thawed repeatedly.
 
Test Procedure
Allow the test, specimen, buffer and/or controls to reach room temperature 15-30℃ (59-86℉) prior to testing.
1.          Bring the pouch to room temperature before opening it. Remove the test device from the sealed pouch and use it as soon as possible.
2.          Place the test device on a clean and level surface.
3.          For serum or plasma specimen: Hold the dropper vertically and transfer 3 drops of serum or plasma (approximately 100μl) to the specimen well(S) of the test device, then start the timer. See illustration below.
4.          For whole blood specimens: Hold the dropper vertically and transfer 1 drop of whole blood(approximately 35μl) to the specimen well(S) of the test device, then add 2 drops of buffer (approximately 70μl) and start the timer. See illustration below.
5.          Wait for the colored line(s) to appear. Read results at 15 minutes. Do not interpret the result after 20 minutes.

Notes: 
Applying sufficient amount of specimen is essential for a valid test result. If migration (the wetting of membrane) is not observed in the test window after one minute, add one more drop of buffer   to the specimen well.
 
Interpretation of Results
Positive: Control line and at least one test line appear on the membrane. The appearance of T2 test line indicates the presence of Leishmania specific IgM antibodies. The appearance of T1 test line indicates the presence of Leishmania specific IgG antibodies. And if both T1 and T2 line appear, it indicates that the presence of both Leishmania specific IgG and IgM antibodies. The lower the antibody concentration is, the weaker the result line is.
Negative: One colored line appears in the control region(C).No apparent colored line appear in the test line region.
Invalid: Control line fails to appear. Insufficient specimen volume or incorrect procedural techniques are the most likely reasons for control line failure. Review the procedure and repeat the test with a new test device. If the problem persists, discontinue using the test kit immediately and contact your local distributor.

Limitations
1.          Novabios Leishmania Ab Test is for in vitro diagnostic use only. The test should be used for the detection of L. donovani antibodies in Whole Blood /Serum / Plasma specimens only. Neither the quantitative value nor the rate of increase in L. donovani antibodies can be determined by this qualitative test.
2.          Novabios Leishmania Ab Test will only indicate the presence of L. donovani antibodies in the specimen and should not be used as the sole criteria for the diagnosis of Visceral leishmaniasis infection.
3.          As with all diagnostic tests, all results must be interpreted together with other clinical information available to the physician.
4.          If the test result is negative and clinical symptoms persist, additional testing using other clinical methods is recommended. A negative result does not at any time preclude the possibility of Visceral leishmaniasis infection.
 

25T/Kits