How-To – HIV Screening Test

14 May 2016 by Macco, Comments Off on How-To – HIV Screening Test

Intended Use

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test is a rapid chromatographic immunoassay for the qualitative detection of antibodies to Human Immunodeficiency Virus type 1 and 2 (HIV-1/2) in human saliva. All positive specimens must be confirmed with Western Blot or other qualified EIA.

Principle

It has been shown that the acquired immunodeficiency syndrome (AIDS) is caused by viruses which transmitted by sexual contact, transfusion, using contaminated blood products and sharing contaminated needles. HIV-1 and 2 viruses have been isolated from patients with AIDS and AIDS-related complex (ARC), high risk persons for AIDS. HIV-1/2 viruses deplete T helper cells, a subpopulation of T cells for body defense, thus causing AIDS patients susceptible to opportunistic infections and developing malignant tumors. The incidence of specific antibodies to HIV-1/2 is high in AIDS, ARC and persons with high risk for AIDS. The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test was designed to detect antibodies to HIV-1/2 in AIDS patients, ARC or high risk persons and identify any potential donors carrying these antibodies in specimens.

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test is a chromatographic immunoassay for the detection of antibodies to HIV-1/2 in human saliva. HIV-1/2 antibodies binding protein are percolated onto membrane as capture reagents in the Test region (T). During the testing, specimen is allowed to react with the colloidal gold reagents which have been labeled with HIV-1/2 specific antigens. Antibodies to HIV-1/2, if present, a pink colored band will develop on the membrane in proportion to the amount of HIV-1/2 antibodies present in the saliva specimen. Absence of this pink colored band in the test region (T) suggests a negative result. to serve as a procedural control, a pink colored band in the control region (C) will always appear regardless of the presence or absence of antibodies to HIV-1/2 in human saliva.

Materials Provided

Each OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test contains:

  1. 1 Foil Pouche. Each pouch contains one device (assay test strip) and one desiccant.
  2. 1 Dropper bottle of Saliva Dilution Buffer (1ml per bottle).
  3. 1 Saliva Collector.
  4. 1 Package Insert.

Materials Required but Not Provided

  1. Timer or clock
  2. Disposable gloves

Storage and Stability

The test device is to be stored refrigerated or at room temperature (2-30°C) under dry conditions for the duration of its shelf life. The test device is stable through the expiration date printed on the sealed pouch. The test device must be kept sealed in the pouch until use. DO NOT FREEZE!

Precaution

  1. For in vitro diagnostic use only.
  2. Do not use a test device beyond the expiration date imprinted on the back of the foil pouch.
  3. Nothing should be placed into the mouth of the subject for at least 10 minutes prior to saliva collection. Those include food, drink, tobacco products or any other materials.
  4. Avoid cross contamination of saliva samples by using a new saliva Collector and Dropper bottle for each sample.
  5. Saliva specimens may be potentially infectious. Avoid contacting with skin by wearing gloves and proper laboratory attire. Properly handle and dispose of all used test devices in an approved biohazard container.
  6. All specimens should be treated as if capable of transmitting diseases. Observe established precautions against microbiological hazards throughout the procedure and follow the standard procedures for proper disposal of specimens.
  7. Wear protective clothing such as laboratory coats, disposable gloves or eye protector when specimens are being tested.
  8. Immediately clean up any spillage of materials with a 1:10 dilution of 5% sodium hypochlorite. Autoclave any contaminated materials for 60 minutes at 121°C before disposal. All liquid waste should be treated with 5% sodium hypochlorite solution (the final concentration should not be lower than 0.5% sodium hypochlorite) for 30 minutes before disposal.
  9. High humidity and temperature can adversely affect results.

Specimen Collection and Storage

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test is designed for the use of buffered human fresh saliva as the specimen. Collecting specimen must follow standard clinical procedure. If specimens are to be shipped, they should be packed in compliance with federal regulations covering the transportation of etiologic agents. Do not place anything into the mouth including food, drink, gum, or tobacco products for at least 10 minutes prior to collection of oral fluid specimen.

  1. Remove the saliva collector out of the pouch.
  2. Pull out the collection swab from the collection chamber.
  3. Place the collection swab into mouth and gently move around root of the teeth for about 5 minutes to let sufficient saliva be collected in the swab.
  4. Remove the swab out of the mouth and place it inside the chamber.
  5. Push the swab to squeeze out 1ml saliva to a dropper bottle which contains 1ml saliva dilution buffer. Make sure the total volume is around 2ml. Mixing well before using.

*Saliva specimens can be kept at room temperature if a test is performed within 24 hours after collection. The specimens must be stored at 2-8°C for two weeks before using. If the test is not performed within 2 weeks, saliva specimens should be frozen for prolonged storage. Frozen specimens must be completely thawed and mixed well prior to testing. Avoid repeatedly freezing and thawing of specimens.

Assay Procedure

  1. Do not open foil pouch until ready to begin testing. Refrigerated test devices should be allowed to come to room temperature (15- 28°C) before opening the pouch.
  2. Remove the device from the protective pouch and label the device with specimen identification. The product is sensitive to the presence of moisture. After opening the package, the test device should be used immediately.
  3. Add 6 drops (250μl) of fresh saliva specimen to the sample well.
  4. Read the result within 10 minutes. Observe the colored band developed over the control region indicating the assay is complete.

IMPORTANT: In order to prevent an incorrect reading, do not read the test results after 15 minutes. If the test is read after 15 minutes, the intensity of the colored bands may change or a new band may appear in test region (T). to avoid confusion, discard all test devices after interpreting results.

Interpretation of Result

POSITIVE: Presence of two visible, pink-colored bands, one in the control region (C) and the other in test region (T), indicates presence of HIV-1/2 antibodies in sample.

NEGATIVE: Presence of a single colored band in the control region (C) indicates the absence of HIV-1/2 antibodies or that the concentration of antibodies in sample is below the detection cut-off level.

INVALID: Results are invalid if no band appears in the control region, or a band appears only in the test region after 10 minutes. An invalid result may be due to improper assay procedures or damage to the device. The assay is inconclusive and the specimen should be re-tested using a new test device.

NOTE: Do not interpret results after 15 minutes. The test band intensity may be weaker or stronger than that of the control band, but a very faint band in the test region indicates that the concentration of HIV-1/2 antibodies in the sample is lower than the detection cut-off level. The sample should be re-tested or confirmed with a more specific method before a positive determination is made.

After Testing

The saliva specimens may be potentially infectious. Avoid contacting with skin by wearing gloves and proper laboratory attire. Properly handle and dispose of all test devices in an approved biohazard container. Residual sample should be disposed of in a medically approved manner after completion of all testing.

Quality Control

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test has a built-in process Control Region. A pink band should always appear in this region regardless of the presence or absence of antibodies to HIV-1/2 in human saliva.. This pink process Control band verifies: 1) that sufficient sample volume was added, 2) that proper flow was obtained, and 3) that the reagents worked properly. If this band is missing, the test was not performed correctly or failed to function correctly. Such a test is inconclusive and the specimen should be re-tested using a new test device. Good laboratory practice requires use of control materials to ensure proper test device performance and reliability. Quality control standards are available for the validation of device functionality from certified labs. When testing the quality control standards, use the same assay procedure as with a oral fluid sample. It is recommended that control specimens be used for each new lot of kits.

Assay Comparisons & Equivalency

Accuracy and equivalency comparisons of OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test were evaluated against 600 individual external WB-certified clinical laboratory samples. The results have been tabulated below.

OraWellUSA Rapid Screen Test vs. WB

WB HIV 1/2 Test(+) HIV 1/2 Test(-) Rowtotals
(+) 300 0 300
(-) 1 299 300
Col.totals 301 299 600

When compared to WB, the percent agreement with OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test, positive samples were 300/300 or 100%. Negative samples recovered at 299/300 or 99.7%, while the overall relative accuracy obtained was 599/600 or 99.8%.

Limitations

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test procedure and the interpretation of results must be followed closely. A test result read after 15 minutes may not be consistent with the original reading obtained within the 15 minutes test period. This assay is designed for testing antibodies against HIV-1/2 in human saliva. Any results derived from the test of other body fluids may not be interpreted correctly based on the current criteria. Other body fluids and pooled samples are not recommended in this assay.

The OraWellUSA HIV-1/2 Antibody Saliva Rapid Screen Test is designed to detect antibodies to HIV-1/2 in saliva and thus is useful in screening blood donated for transfusion and further manufacture, in establishing prior infection of HIV or in evaluating patients with AIDS symptoms and signs. For positive specimens, it is recommended that more specific supplemental test should be conducted and clinical evaluation on patient’s situation should be performed before final conclusion is made. HIV-1/2 rapid testing alone cannot be used to diagnose AIDS even if the antibodies against HIV-1/2 are present in patient’s saliva. The test result will only indicate the qualitative level of antibodies of HIV 1/2 in the specimen and should not be used as the sole criteria for the diagnosis of HIV. Even if the test results are positive, further clinical evaluation should be considered with other clinical information available to the physician. A negative result does not rule out infection by HIV because the antibodies to HIV may be absent or may not be present in sufficient quantities to be detected at a very early stage of infection or could result from improper sample collection.

References

  1. M Popovic, MG Sarngadharan, E Read, and RC Gallo. Detection, isolation and continuous production of cytopathic retroviruses (HTLV-III) from patients with AIDS and pre-Aids. Science 1984, 497-500.
  2. RC Gallo, SZ Salahuddin, M Popovic, et al. Frequent detection and isolation of cytopathic retroviruses (HTLV-III) from patients with AIDS and at risk for AIDS . Science 1984, 224:500-503.
  3. JW Curran, WM Morgan, AM Hardy, et al., The epidemiology of AIDS: current status and future prospects. Science 1985, 229:1352-1357.
  4. Clavel, F. HIV-2, The West African AIDS virus. AIDS 1987, 1:135-140.
  5. P Piot, FA Plummer, FS Mhalu, et al. AIDS: an international perspective, Science 1988, 239:573-579.
  6. Provisional Public Health Service Interagency recommendations for screening donated blood and plasma for antibody to the virus causing Acquired Immunodeficiency Syndrome. MNWR. 1985,34: 1-5.

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