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U.S. Global AIDS Coordinator, Randal Tobias, is being publicly tested for HIV/AIDS in Ethiopia, in an effort to reduce the stigma of being tested.

HIV (Human Immunodeficiency Virus) test kits used both to screen donor blood, blood components and cellular products, and to diagnose, treat and monitor persons with HIV and Acquired Immuno Deficiency Syndrome (AIDS) are regulated in the United States by the Food and Drug Administration (FDA).

HIV tests to detect antibodies, antigens or ribonucleic acid (RNA) in serum, plasma, oral fluid, dried blood spot or urine have been approved by FDA for donor screening, diagnosis, prognosis and patient monitoring.

Terminology

The window period is the time from infection until a test can detect any change. The average window period with antibody tests is 22 days. Antigen testing cuts the window period to approximately 16 days and NAT further reduces this period to 12 days. FDA 2001

Antibody tests are reported as positive or negative. Performance of these tests is described in terms of:

sensitivity the percentage of the results that will be positive when HIV is present
specificity the percentage of the results that will be negative when HIV not present.

All diagnostic tests have limitations, and sometimes their use may produce erroneous or questionable results.

false positive results indicate that HIV is found to be present when, in fact, it is not.
false negative results do not identify HIV that is present.

Nonspecific reactions, hypergammaglobulinemia, or the presence of antibodies directed to other infectious agents that may be antigenically similar to HIV can produce false positive results. Autoimmune diseases, such as systemic lupus erythematosus, can also cause false positive results.

Principles

Screening donor blood and cellular products

Tests selected to screen donor blood, blood components and cellular products provide a high degree of confidence that HIV is not present. A combination of antibody, antigen and nucleic acid tests are used by blood banks in Western countries. The WHO estimated that in 2000 inadequate blood screening in some countries resulted in 1 million new HIV infections.

Diagnosis of HIV infection

Different tests are selected for the diagnosis of HIV infection in a particular person, which requires a high degree of confidence that HIV is present. In the United States, this is achieved by an algorithm using two tests to detect antibodies, which are the body’s response to infection. If antibodies are detected by an initial test based on the ELISA method, then a second test based on the Western blot procedure determines the size of the antigens in the test kit binding to the antibodies. According to the CDC, there are 1.1 million persons infected with HIV in the United States as of 2005.

Human Rights

The UNAIDS/WHO policy statement on HIV Testing (PDF) states that conditions under which people undergo HIV testing must be anchored in a human rights approach, which pays due respect for ethical principles. According to these principles, the conduct of HIV testing of individuals must be:

confidential
accompanied by counselling
conducted with informed consent, meaning both informed and voluntary

The reality is that stigma and discrimination continue to stop people, who may have been exposed to HIV infection, from consenting to HIV testing.

HIV testing and counselling has a pivotal role in preventing HIV infection around the world.

Antibody tests

Antibody tests are specifically designed for the routine testing of HIV in adults, are inexpensive, and are very accurate. If a person does not have a realistic risk of infection, then these tests are not necessary.

Antibody tests give false negative results during the window period of between three weeks and six months from the time of HIV infection until the immune system produces detectable amounts of antibodies. The vast majority of people have detectable antibodies after three months. A six month window is extremely rare with modern antibody testing. During this window period an infected person can transmit HIV to others, without their HIV infection being detectable using an antibody test. Antiretroviral therapy during the window period can delay the formation of antibodies and extend the window period beyond 12 months. C B Hare, B L Pappalardo, M P Busch, B Phelps, S S Alexander, C Ramstead, J A Levy, F M Hecht (2004). "Negative HIV antibody test results among individuals treated with antiretroviral therapy (ART) during acute/early infection". The XV International AIDS Conference. pp. Abstract no. MoPeB3107. {{cite conference}}: Unknown parameter |booktitle= ignored (|book-title= suggested) (help)CS1 maint: multiple names: authors list (link)

ELISA

The ELISA test, or the enzyme immunoassay (EIA), was the first screening test commonly employed. It has a high sensitivity.

The test proceeds by the general ELISA method: the person's serum is diluted 400 fold and applied to a plate to which HIV antigens have been attached. Some of the antibodies in the serum may bind to these HIV antigens. The plate is then washed to remove all other components of the serum. Then a specially prepared "secondary antibody"—an antibody that binds to human antibodies—is applied to the plate, followed by washes. This secondary antibody is chemically linked in advance to an enzyme. Thus the plate will contain enzyme in proportion to the amount of secondary antibody bound to the plate. A substrate for the enzyme is applied, and catalysis by the enzyme leads to a change in color or fluorescence. As the ELISA results are reported as a number, the most controversial aspect of this test is deciding the "cut off" point between positive and negative.

Western blot

The Western blot test uses the general Western blot procedure. HIV-infected cells are opened and the contained proteins are entered into a slab of gel to which a voltage is applied. Different proteins will move with different velocities in this field, depending on their size, while their electrical charge is levelled by a substance, called sodium lauryl sulfate. Once the proteins are well separated, they are transferred to a membrane and the procedure continues similar to ELISA: the person's diluted serum is applied to the membrane and antibodies in the serum may attach to some of the HIV proteins. Antibodies which do not attach are washed away, and enzyme-linked antibodies with the capability to attach to the person's antibodies first detect to which HIV proteins the person has antibodies.

There is no universal criteria for interpreting the Western blot test: the number of viral bands which must be present vary from definition to definition. If no viral bands are detected, the result is negative. If at least one viral band for each of the GAG, POL, and ENV gene-product groups are present, the result is positive. Thus the three-gene-product approach to Western blot interpretation has not been adopted for public health or clinical practice. Tests in which less than the required number of viral bands are detected are reported as indeterminate: a person who has an indeterminate result should be retested, as later tests may be more conclusive. Almost all HIV-infected persons with indeterminate Western-Blot results will develop a positive result when tested in one month; persistently indeterminate results over a period of six months suggests the results are not due to HIV infection.

Rapid or point-of-care tests

Rapid Antibody Tests are qualitative immunoassays intended for use as a point-of-care test to aid in the diagnosis of HIV infection. These tests should be used in conjunction with the clinical status, history, and risk factors of the person being tested. The specificity of Rapid Antibody Tests for in low-risk populations has not been evaluated. These tests should be used in appropriate multi-test algorithms designed for statistical validation of rapid HIV test results.

If no antibodies to HIV are detected, this does not mean the person has not been infected with HIV. It may take several months after HIV infection for the antibody response to reach detectable levels, during which time rapid testing for antibodies to HIV will not be indicative of true infection status. A comprehensive risk history and clinical judgement should be considered before concluding that an individual is not infected with HIV.

OraQuick is an antibody test that provides results in 20 minutes. The blood, plasma or oral fluid is mixed in a vial with developing solution, and the results are read from a sticklike testing device.

Orasure is an HIV test which uses mucosal transudate from the tissues of cheeks and gums. It is an antibody test which first employs ELISA, then Western Blot.

There is also a urine test; it employs both the ELISA and the Western Blot method.

Home Access Express HIV-1 Test is a FDA-approved home test: the patient collects a drop of blood and mails the sample to a laboratory; the results are obtained over the phone.

There have been a number of cases of fraudulent tests being sold via mail order or the Internet to the general public. In 1997, a California man was indicted on mail fraud and wire charges for selling supposed home test kits. In 2004, the US Federal Trade Commission asked Federal Express and US Customs to confiscate shipments of the Discreet home HIV test kits, produced by Gregory Stephen Wong of Vancouver, BC. In February 2005, the US FDA issued a warning against using the rapid HIV test kits and other home use kits marketed by Globus Media of Montreal Canada.

Interpreting antibody tests

ELISA testing alone cannot be used to diagnose AIDS, even if the recommended investigation of reactive specimens suggests a high probability that the antibody to HIV 1 is present. In the United States, such positive tests are not reported out unless confirmed by a Western Blot test.

The ELISA antibody tests were developed to provide a high level of confidence that donated blood was NOT infected with HIV. It is therefore not possible to conclude that blood rejected for transfusion because of a positive ELISA antibody test is in fact infected with HIV. Usually, retesting the donor in several months will produce a negative ELISA antibody test.

It is known that false positive results due to factors unrelated to exposure to the HIV virus are common with the ELISA test. False positives can be caused by antibodies to viruses other than HIV, antibodies produced by pregnancy, and other medical conditions. A false positive results DOES NOT indicate that you are infected with the AIDS virus, nor does it indicate a condition of significant risk to your health.

The evidence for regarding the risks and benefits of HIV screening was reviewed in July 2005. ("Screening for HIV: A Review of the Evidence for the U.S. Preventive Services Task Force", Annals of Internal Medicine, Chou et. al, Volume 143 Issue 1, pp. 55-73. [1]): "The use of repeatedly reactive enzyme immunoassay followed by confirmatory Western blot or immunofluorescent assay remains the standard method for diagnosing HIV-1 infection. A large study of HIV testing in 752 U.S. laboratories reported a sensitivity of 99.7% and specificity of 98.5% for enzyme immunoassay, and studies in U.S. blood donors reported specificities of 99.8% and greater than 99.99%. With confirmatory Western blot, the chance of a false-positive identification in a low-prevalence setting is about 1 in 250 000 (95% CI, 1 in 173 000 to 1 in 379 000)."

Antigen tests

The p24 antigen test detects the presence of the p24 protein of HIV (also known as CA), a major core protein of the virus. Monoclonal antibodies specific to the p24 protein are mixed with the person's blood. Any p24 protein in the person's blood will stick to the monoclonal antibody and enzyme-linked antibody to the monoclonal antibodies to p24 causes a color change if p24 was present in the sample.

This test is now used routinely to screen blood donations, thus reducing the window to about 16 days. It is not useful for general diagnostics, as it has very low sensitivity and only works during a certain time period after infection before the body produces antibodies to the p24 protein.

Nucleic acid based tests

Nucleic acid based tests amplify and detect a 142 base target sequence located in a highly conserved region of the HIV gag gene. Since 2001, donated blood in the United States, has been screened with nucleic acid based tests, shortening the window to about 12 days. Since these tests are relatively expensive, the blood is screened by first pooling some 10-20 samples, testing these together, and if the pool tests positive, each sample is retested individually. A different version of this test is intended for use in conjunction with clinical presentation and other laboratory markers of disease progress for the clinical management of HIV-1 infected patients.

In the RT-PCR test, the viral RNA is extracted from the patient's plasma and is treated with reverse transcriptase so that the RNA of the virus is transcribed into DNA. The polymerase chain reaction (PCR) is applied, using two primers thought to be unique to the virus's genome. After the PCR amplification process is completed, which takes some time, the resulting amplified segments bind to specific oligonucleotides bound to the vessel wall and are then made visible with a probe that's bound to an enzyme. The amount of virus in the sample can be quantified with sufficient accuracy to detect three fold changes.

In the Quantiplex bDNA or branched DNA test plasma is centrifugated to concentrate the viruses, which are then opened to release the RNA. Special oligonucleotides are added which bind to viral RNA and to certain oligonucleotides bound to the wall of the vessel. In this way, viral RNA is fastened to the wall. Then new oligonucleotides are added which bind at several locations to this RNA; and other oligonucelotides which bind at several locations to those oligonucleotides. This is done to amplify the signal. Finally, oligonucleotides that bind to the last set of oligonucleotides and that are bound to an enzyme are added; the enzyme action causes a color reaction which allows to quantify the viral RNA in the original sample. Monitoring the effects of antiretroviral therapy by serial measurements of plasma HIV-1 RNA with this test has been validated for patients with viral loads greater than 25,000 copies per millilitre. reference

Other tests used in HIV/AIDS treatment

The CD4 T-cell count is not an HIV test, but rather a procedure where the number of CD4 T-cells in one microlitre of blood are counted in a standard medical lab test after a blood draw.

This test does not check for the presence of HIV. It is used monitor the immune system function in HIV+ people. Declining CD4 T-cell counts are considered to be a marker of the progression of HIV infection. In HIV+ people, AIDS is officially diagnosed when the count drops below 200 cells or when certain opportunistic infections occur. This use of a CD4 count as an AIDS criterion occurred in 1992; the value of 200 was chosen because it corresponded with an increased likelihood of opportunistic infections. Lower levels of CD4 counts in people with AIDS are indicators that prophylaxis against certain types of opportunistic infections should be instituted.

Low CD4 T-cell counts are associated with a variety of conditions, including many viral infections, bacterial infections, parasitic infections, sepsis, tuberculosis, coccidioidomycosis, burns, trauma, intravenous injections of foreign proteins, malnutrition, over-exercising, pregnancy, normal daily variation, psychological stress, and social isolation.

This test is also used occasionally to estimate immune system function for people whose CD4 T cells are impaired for reasons other than HIV infection, which include several blood diseases, several genetic disorders, and the side effects of many chemotherapy drugs.

Generally speaking, the lower the number of T cells, the lower the immune system's function will be. Normal T4 counts are between 500 and 1500 CD4+ T cells per microliter and the counts may fluctuate in healthy people, depending on recent infection status, nutrition, exercise and other factors -- even the time of day. Women tend to have somewhat lower counts than men.

Symptoms of T4 cell immune collapse are almost never seen until the number drops below 200. Similar symptoms of immune collapse are generally seen in people with very low T4 cell counts, whether this immunosuppression is caused by HIV, cancer, or some other disease. However, the long-term treatment differs substantially, because it needs to address the cause of the immunosuppression.

External links

UNAIDS issues HIV testing policy June 2004
HIV Testing August 5, 2004
HIV Antibody Assays September 2001
http://www.hivtest.org/ - CDC HIV test resources
US FDA list of approved HIV tests
Not B4 We Know The strategy of testing for STDs together before having sex

@@ Line 17: / Line 17: @@
 * '''[[false negative]]''' results do not identify HIV that is present.
-Nonspecific reactions, hypergammaglobulinemia, or the presence of antibodies directed to other infectious agents that may be antigenically similar to HIV can produce [[false positive]] results.  Autoimmune diseases, such as [[systemic lupus erythematosus]], can also cause [[false positive]] results. Indeed, the scientific literature has documented more than 60-70 different factors other than past or present infection with HIV that can cause a positive reaction on HIV tests - including flu, flu vaccination, herpes simplex, rheumatoid arthritis, malaria, tuberculosis, antibodies to leprosy - and even pregnancy. A list of the factors known to cause false-positive results, together with references to the medical literature for each factor, is available at http://www.aliveandwell.org/html/questioning/66reas.html.
+Nonspecific reactions, hypergammaglobulinemia, or the presence of antibodies directed to other infectious agents that may be antigenically similar to HIV can produce [[false positive]] results.  Autoimmune diseases, such as [[systemic lupus erythematosus]], can also cause [[false positive]] results.
 ==Principles==
@@ Line 25: / Line 25: @@
 ===Diagnosis of HIV infection===
-Different tests are selected for the diagnosis of HIV infection in a particular person, which requires a high degree of confidence that HIV is present. In the United States, this is achieved by an [[algorithm]] using two tests to detect antibodies, which are the body&#8217;s response to infection. If antibodies are detected by an initial test based on the [[ELISA]] method, then a second test based on the [[Western blot]] procedure determines the size of the antigens in the test kit binding to the antibodies. This testing [[algorithm]] is claimed to have an accuracy of 99.50%, which means that testing 100,000 people not infected with HIV would be expected to produce 500 [[false positive]] results. The [[Prosecutor's fallacy|predictive value]] of this testing [[algorithm]] depends on the prevalence of HIV infection in the group being tested. If 10% of the group being tested is infected with HIV, less than 2% of the positive results will be [[false positive]]. But if less than 1 in 7,500 people in the group being test are infected with HIV, then over 90% of the positive results will be [[false positive]].
+Different tests are selected for the diagnosis of HIV infection in a particular person, which requires a high degree of confidence that HIV is present. In the United States, this is achieved by an [[algorithm]] using two tests to detect antibodies, which are the body&#8217;s response to infection. If antibodies are detected by an initial test based on the [[ELISA]] method, then a second test based on the [[Western blot]] procedure determines the size of the antigens in the test kit binding to the antibodies.
+According to the CDC, there are 1.1 million persons infected with HIV in the United States as of 2005.
-According to the CDC, there are 1.1 million persons infected with HIV in the United States as of 2005.  Out of a population of 298,098,020 (the population of the US as of February 13 2006,) this means that a random sample of 100,000 Americans, as described above, would include 369 HIV-positives.  As noted above, if this group were tested for HIV, 500 of them would test false-positive, which means that '''false positives in the group would outnumber true positives by a factor of 35%'''.
 === Human Rights ===
@@ Line 45: / Line 44: @@
 Antibody tests give [[false negative]] results during the ''window period '' of between three weeks and six months from the time of HIV infection until the [[immune system]] produces detectable amounts of antibodies. The vast majority of people have detectable antibodies after three months. A six month window is extremely rare with modern antibody testing. During this window period an infected person can transmit HIV to others, without their HIV infection being detectable using an antibody test. [[antiretroviral drug|Antiretroviral therapy]] during the window period can delay the formation of antibodies and extend the window period beyond 12 months. {{cite conference | author=C B Hare, B L Pappalardo, M P Busch, B Phelps, S S Alexander, C Ramstead, J A Levy, F M Hecht | url=http://www.iasociety.org/ejias/show.asp?abstract_id=2172342 | title=Negative HIV antibody test results among individuals treated with antiretroviral therapy (ART) during acute/early infection | booktitle= The XV International AIDS Conference | year=2004 | pages=Abstract no. MoPeB3107}}
-A person exposed to [[HIV]] would be expected to produce antibodies that specifically bind to HIV. The development of antibody tests for HIV has been complicated because all human [[blood]] has large quantities of antibodies that bind to components of the antibody test kits. For this reason, the blood to be tested is diluted in an attempt to detect antibodies that have been produced after exposure to HIV. Manufacturers of all these test kits state that, "There is no single recognised standard for establishing the presence or absence of [[antibodies]] to [[HIV]] in human blood."
 ===ELISA===
-The '''ELISA test,''' or the enzyme immunoassay (EIA), was the first screening test commonly employed. It has a high sensitivity. The low specificity of the test is because antibodies attach to antigens in the test kits "by accident", even though the person has never been exposed to HIV. About 80% of positive ELISA tests are followed by a negative Western-Blot test, and therefore regarded as [[false positive]].
+The '''ELISA test,''' or the enzyme immunoassay (EIA), was the first screening test commonly employed. It has a high sensitivity.
 The test proceeds by the general [[ELISA]] method: the person's [[blood serum|serum]] is diluted 400 fold and applied to a plate to which HIV antigens have been attached. Some of the antibodies in the serum may bind to these HIV antigens. The plate is then washed to remove all other components of the serum.  Then a specially prepared "secondary antibody"&mdash;an antibody that binds to human antibodies&mdash;is applied to the plate, followed by washes.  This secondary antibody is chemically linked in advance to an [[enzyme]].  Thus the plate will contain enzyme in proportion to the amount of secondary antibody bound to the plate.  A [[substrate (biochemistry)|substrate]] for the enzyme is applied, and catalysis by the enzyme leads to a change in color or fluorescence. As the ELISA results are reported as a number, the most controversial aspect of this test is deciding the "cut off" point between positive and negative.
@@ Line 55: / Line 54: @@
 The '''Western blot test''' uses the general [[Western blot]] procedure. HIV-infected cells are opened and the contained [[protein]]s are entered into a slab of gel to which a [[voltage]] is applied. Different proteins will move with different velocities in this field, depending on their size, while their electrical charge is levelled by a substance, called sodium lauryl sulfate. Once the proteins are well separated, they are transferred to a membrane and the procedure continues similar to ELISA: the person's diluted serum is applied to the membrane and antibodies in the serum may attach to some of the HIV proteins. Antibodies which do not attach are washed away, and enzyme-linked antibodies with the capability to attach to the person's antibodies first detect to which HIV proteins the person has antibodies.
-There is no universal criteria for interpreting the Western blot test: the number of viral bands which must be present vary from definition to definition.  If no viral bands are detected, the result is negative. If at least one viral band for each of the GAG, POL, and ENV gene-product groups are present, the result is positive. Such a criterion, however, is not sensitive enough for clinical use, as the absence of antibodies to p24 or to p31 is relatively common even in patients who are clearly infected with HIV.  Thus the three-gene-product approach to Western blot interpretation has not been adopted for public health or clinical practice. The Association of State and Territorial Public Health Laboratory Directors (ASTPHLD) and CDC standard ignores p31 results, and interprets the presence of gp41 and gp120/160 bands as a positive result. Tests in which less than the required number of viral bands are detected are reported as indeterminate: a person who has an indeterminate result should be retested, as later tests may be more conclusive. Almost all HIV-infected persons with indeterminate Western-Blot results will develop a positive result when tested in one month; persistently indeterminate results over a period of six months suggests the results are not due to HIV infection.
+There is no universal criteria for interpreting the Western blot test: the number of viral bands which must be present vary from definition to definition.  If no viral bands are detected, the result is negative. If at least one viral band for each of the GAG, POL, and ENV gene-product groups are present, the result is positive. Thus the three-gene-product approach to Western blot interpretation has not been adopted for public health or clinical practice. Tests in which less than the required number of viral bands are detected are reported as indeterminate: a person who has an indeterminate result should be retested, as later tests may be more conclusive. Almost all HIV-infected persons with indeterminate Western-Blot results will develop a positive result when tested in one month; persistently indeterminate results over a period of six months suggests the results are not due to HIV infection.
-The Western blot test is usually performed after a positive ELISA test, because many samples that are ELISA negative have antibodies to one or more of the proteins in the Western blot test, and would give inaccurate results, especially when interpreted according to criteria which require a complete absence of all viral ''and non-viral'' bands to report a Western-Blot result as negative rather than indeterminate.
-In Africa, if the Western-Blot test shows 2 bands - the test is considered HIV Positive. In Australia - if the Western-Blot test shows 2 bands - the test is considered HIV Negative.
-According to data presented in Lundberg et al. (JAMA 260:674-679) when the US FDA criteria are used to interpret the HIV Western blot less than 50% of US AIDS patients are HIV positive whereas 10% of persons not at risk of AIDS are also positive by the same criteria.
-For further reading:
-http://www.nature.com/nbt/journal/v11/n6/abs/nbt0693-696.html
 ===Rapid or point-of-care tests===
@@ Line 87: / Line 76: @@
 The ELISA antibody tests were developed to provide a high level of confidence that donated blood was ''NOT'' infected with HIV. It is therefore not possible to conclude that blood rejected for transfusion because of a ''positive'' ELISA antibody test is in fact infected with HIV. Usually, retesting the donor in several months will produce a ''negative'' ELISA antibody test.
-All laboratory tests can produce 'false alarms'. This is a positive result in the screening test, which, on confirmatory testing is shown to be negative. False reactions are a recognised complication of all biological tests and are perfectly normal. They are of no significance for the health of the donor.
 It is known that false positive results due to factors unrelated to exposure to the HIV virus are common with the ELISA test. False positives can be caused by antibodies to viruses other than HIV, antibodies produced by pregnancy, and other medical conditions. A false positive results DOES NOT indicate that you are infected with the AIDS virus, nor does it indicate a condition of significant risk to your health.
@@ Line 122: / Line 109: @@
 == See also ==
-* [[Raven paradox]]
+* [[AIDS testing]]
-* [[Prosecutor's fallacy]]
-* [[Bayesian inference]]
 ==External links==