Beijing Kinghawk CO.,LTD

INTENDED USE

Kinghawk One Step Malaria P.f/Pan Whole Blood Test Cassette is a rapid self-performing, qualitative, two site sandwich immunoassay, utilizing whole blood for the detection of P.falciparum specific histidine rich protein-2 (P.f HRP-2) and Pan specific pLDH. The test may also be used for differentiation of P.falciparum and other malarial species in whole blood samples and for the follow up of anti-malarial therapy.
For in vitro diagnostic use only. For healthcare professional use only.

SUMMARY

Malaria remains one of the most serious tropical and subtropical   diseases in many countries of the world. It is rampant in most areas of the tropics. Malaria is caused by a parasite that is transmitted from one person to person by the bite of infected Anopheles mosquitoes. There are four kinds of malaria that can infect humans: Plasmodium falciparum, P.vivax, P.ovale and P.malariae. Malaria also has been reported from blood transfusions or congenitally from mother to child. It is estimated to affect more than 500 million people causing between one and three million deaths every year.

PRINCIPLE

Kinghawk One Step Malaria P.f/Pan Whole Blood test utilizes the principle of immunochoromatograhpy. As the test sample flows through the membrane assembly of the device after addition of the buffer, the colored monoclonal P.f specific HRP-2, Pan specific pLDH colloidal gold conjugate antibodies complexes the proteins in the lysed sample. This complex moves further on the membrane to the test region where it is immobilized by the P.f specific HRP-2 antibody / Pan specific pLDH which will lead to a formation of purple color band/s. While both the bands will appear at the test region in falciparum positive samples, only one band would appear in non-falciparum malaria positive sample. Absence of this color band/s in the test region indicates a negative test result.

The unreacted conjugate along with the rabbit globulin colloidal gold conjugate and unbound complex if any, move further on the membrane and are subsequently immobilized by anti-rabbit antibodies coated on the membrane at the control region, forming a pink / purple band. This control band serves to validate the test performance.

PRECAUTIONS AND WARNINGS
 
This kit is for in vitro use only. Do not swallow.
Discard after first use. The test cannot be used more than once.
Do not use test kit beyond the expiration date.
Do not use the kit if the pouch is punctured or not well sealed.
Decontaminate and dispose of all specimens, reaction kits and potentially contaminated materials, as if they were infectious waste, in a biohazard container.
Wear protective gloves while handling specimens. Wash hands thoroughly afterwards. Avoid splashing or aerosol formation. Clean up spills thoroughly using an appropriate disinfectant.
Keep out of the reach of children.
DISPOSAL OF THE DIAGNOSTIC: The used-device has the infectious risk. The process of disposing the diagnostic must follow the local infectious disposal law or laboratory regulation.
CONTENT OF THE KIT

1.  Individual pouches, each containing:
 Test Device
Desiccant pouch
5μl sample dropper
2.  Alcohol pads (optional)
3.  Lancets (optional)
4.  Clearing buffer in a dropper bottle.
5.  Leaflet with instructions for use.

STORAGE AND STABILITY 
 
Stored at 4ºC to 30ºC in the sealed foil pouch up to the expiration date.
Keep away from sunlight, moisture and heat.
DO NOT FREEZE.
SPECIMEN COLLECTION AND PREPARATION

Collection by venipuncture

1. Collect the whole blood into the collection tube (using the suitable anti-coagulant)by venipuncture.
2. If specimens are not immediately tested, they should be refrigerated at 2°C ~ 8°C. For storage periods greater than three days, freezing is recommended. They should be brought to room temperature prior to use.

Collection using a lancet

1. Clean the area to be lanced with an alcohol swab.
2. Squeeze the end of the fingertip and pierce with a sterile lancet. Wipe away the first drop of blood with sterile cotton.
3. Take a dropper provided, while gently squeezing the tube, immerse the open end in the blood drop and then gently release the pressure to draw blood into the dropper.

TEST PROCEDURE
  
Allow the device, buffer and specimen to equilibrate to room temperature (10°C ~30°C) prior to testing.
 
Remove the test cassette from the foil pouch by tearing at the notch and place it on a level surface.
Slowly add 5 µl of whole blood to the sample well(A) and then add 3 drops (about 80ul) of clearing buffer to the buffer well (B).
As the test begins to work, you will see purple color move across the result window in the center of the test device.
Wait for 15 minutes and read results. Do not read results after 30 minutes.









Caution: The above interpreting time is based on reading the test results at room temperature of 10 ºC ~30 ºC. If your room temperature is significantly lower than 10 ºC , then the interpreting time should be properly increased.

INTERPRETATION OF RESULTS

Positive (+) 
The presence of two color band  ("T1" and "C") indicates a positive result for a infection with P. falciparum 




2.  The presence of two color bands ("T2" and "C") indicates a positive result for Non-P. falciparum (P. vivax, P. malariae or P.ovale) infection.




 
The presence of three color bands("T1", "T2"and "C") indicates a positive result for P. falciparum monoinfection or "mixed" infection.
 



Negative (-)
The presence of only one band ("C") within the result window indicates a negative result.


Invalid 
If the color band ("C") is not visible within the result window after performing the test, the result is considered invalid. The directions may not have been followed correctly or the test may have deteriorated. It is recommended that the specimen be re-tested.



















QUALITY CONTROL

Though there is an internal procedural control line in the test device of Control region, the use of external controls is strongly recommended as good laboratory testing practice to confirm the test procedure and to verify proper test performance. Positive and negative control should give the expected results. When testing the positive and negative control, the same assay procedure should be adopted.

LIMITATIONS OF PROCEDURE

The test is limited to the detection of P.falciparum specific histidine rich protein-2 (P.f HRP-2) and Pan specific pLDH. Although the test is accurate in detecting antigens of Malaria P.f/Pan, a low incidence of false results can occur. Other clinically available test are required if questionable results are obtained. As with all diagnostic tests, a definitive clinical diagnosis should not be based on the results of a single test, but should only be made by the physician after all clinical and laboratory findings have been evaluated.

PERFORMANCE CHARACTERISTICS

1. Kinghawk One Step Malaria P.f/Pan Whole Blood Test have a sensitivity of >90% at densities above 40-100 parasites /ul blood
 
Sensitivity and Specificity
Collect 325 patients' samples from several medical institutions. Test these samples using microscopic examination method and colloidal gold method. Compare sensitivity and specificity between. These samples were obtained from patients attending infection clinics.

Table 1 Result of the sensitivity and specificity between (Malaria P.f):

Table 2 Result of the sensitivity and specificity between (Malaria Pan):

B.     Precision

1.  Within run precision was determined by using 10 replicates of four different specimens containing different concentrations of antigen. The negative and positive values were correctly identified 100% of the time.

2.  Between run precision was determined by using the four different specimens containing different concentrations of antigen in 3 different lots of test devices. Again negative and positive results were correctly identified 100% of the time.

BIBLIOGRAPHY OF SUGGESTED READIN
 
David R. and et. Al. A Longitudinal Study of Type-Specific Antibody Responses to Plasmodium falciparum Merozoite Surface Protein - 1 in an Area of Unstable Malaria in Sudan, Journal of Immunology, 161 : 347-359 (1998).
 
Helen L.Gibson, Jeffrey E.Tucker : Structure and expression of the gene for Pv200, a major blood-stage surface antigen of Plasmodium vivax. Molecular and Biochemical Parasitology, 50 (1992) 325-334.
 
Alon Warburg and Imogene Schneider. In Vitro Culture of the Mosquito Stages of Plasmodium falciparum. Experimental parasitology 76, 121-126 (1993).
BEIJING KINGHAWK PHARMACEUTICAL CO., LTD