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Diagnostic tests for HIV/AIDS

HIV/AIDS also called as Human immunodeficiency virus/Acquired immune deficiency syndrome. One of the most dangerous STD’s, significantly challenging the gobal world with its devastating effects.HIV is caused by the “LENTI”virus[a class of retrovirus].HIV virus mainly acts on the imunne system of the body,by attacking the CD4 cells.Destruction of these cells results in the loss of defensive actions aganist various types of infections.Over the time,HIV leads to AIDS[Accquired immune deficiency syndrome]. If HIV is left untreated,it leads to AIDS stage.Well,early detection of the infection,using appropriate diagnostic tests helps the patients to live longer with the infection, without reaching to the AIDS stage.

The diagnostic tests for HIV includes;ELISA,westernblot test,polymerease chain reaction,[NAT test].ELISA is the primary test for HIV detection,if it is found to be positive,then western blot test is preffered for the further confirmation.If the ELISA is negative,then the individual should be retested again after a month.While PCR is a technique used to detect virus in the body,which is accurate method.Some of the other tests include antigen tests,antigen-antibody combinations.

 

ELISA:

Enzyme linked immune sorbent assay,was the first diagnostic test employed for the HIV. It has a high sensitivity.It involes the immune system component i,e.antibodies[igG,igM].Firstly serum from the person is collected,and it is diluted upto 400 folds.The diluted serum is applied on the plate to which HIV antigens are attached.If the person is having HIV antibodies,they may bind to the antigens.Then the plate is washed to remove the all other serum components.”SECONDARY ANTIBODY”-an antibody that binds to the human antibodies itself,is applied to the plate followed by another wash.This secondary antibody is chemically linked to enzyme.Thus the secondary antibody bound to the plate is in proportion with the enzyme on the plate.A substrate is added for the enzyme,and the catalysis by the enzyme leads to fluorescene or in colour of the substances.The most controversial aspect of this test is determining the”cut-off” point between a positive and negative result.

 

WESTERN BLOT:

Similar to the ELISA,western blot is also an antibody detection test.But unlike ELISA, first the viral proteins are separated and immobilised.Binding of serum antibodies to specific HIV proteins is visualised.

Specific cells of HIV infection are opened and the proteins within are palced into a slab of gel,to which the electric current is applied.Different proteins have different speeds, depending on their size.While the electric charge is levelled by a sodium lauryl sulphate, surfactant. Once the proteins are well separated,they are transferred to the membrane and the further process is continued similar to ELISA.The diluted serum is applied to the membrane and the antibodies in the serum may attached to the the HIV proteins.Antibodies that are not attached are washed away,where as the enzyme linked antibodies attaches to the secondary antibodies and determines to which HIV proteins the person is having antibodies.Commercially prepared western blot test kits contains the HIV proteins alredy on a cellulose acetate strip.

There is no particular specific criteria for interpreteting the western blot results.The number of viral bands present may vary.If viral bands are detected,then the test result is negative.If atleast one viral band is detected for each of the GAG,POL,and ENV gene product groups,then the reult is positive.But the 3-gene product approach of interpretation is NOT For clinical practice.For tests in which less than the required number of viral bands are detected,they are reported as indeterminate.Persons with the indeterminate results are retested for the conclusive results.Almost all the persons with the indeterminant results are found to develop a positive result.

The HIV proteins used in western blotting can be produced by using recombinant DNA techniques called recombinant immunoblot assay[RIBA].

If no antibodies are detected to HIV,that doesn’t mean the person has not be infected.It may take several months for infection to show antibody response of detectable levels. Rapid testing for antibodies to HIV will not give a indicative of true infection.In most the cases,it takes  two to six weeks to reach a detectable level of infection.Even these tests are having the high speficity,false positives may do occur.The positive results should be confirmed by using a lab western blot test.

Even ELISA is having high probability to diagnose HIV,it can’t be used alone,should not be reported as ‘positive’ unless confirmed by a western blot.

Rare false results are due to factors unrelated to HIV,found to be more often with the ELISA than western blot.False positives may be asosicated with the medical conditions such as recent acute illness and allergies.

 

PCR:

Polymerease chain reaction is used to detect genetic material i,e. RNA.It is used for the amplification of the DNA sequences. PCR technique is developed by KARY MULLIS in 1983 and wsa awrded nobel prize in 1993 in chemistry.

The basic underlying principle in PCR is’extensive modifications can be done  to perform the wide array of genetic manipulations.Generally PCR employs a originally isolated heat-stable polymerease enzyme i,e. TAQ POLYMEREASE.This polymerease enzymatically assembles a new DNA strands,using single stranded DNA as template.This method is also known as VIRAL LOAD TEST.Viral load assays measure HIV-RNA from virus particles in the blood plasma.This test is also called NUCLEIC ACID TEST [NAT].IT is of 2 types DNA-PCR,RNA-PCR.

DNA-PCR: Used for screening of babies of HIV+ mothers

RNA-PCR: Used to screen blood donations, organ donations.

PCR tests have been developed that can detect the infection,as little as one viral genome among the DNA of vast group of host cells.Infections can be detected earlier. The donated blood can be directly screened for the virus.Unlike ELISA and western bloting, used for antibodies detection,PCR is is more accurate with early detection.

PCR can be used for the earliest detection, but ELISA and western blot are not effective in early detection, as the antibodies takes time to appear in the circulation.

 

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