A-306 Standardization of Automated Methodology for Qualitative Molecular Detection of HIV-1 in Dried Blood Spots (DBS)

Abstract

Abstract Background Human Immunodeficiency Virus (HIV) is the etiological agent of acquired immunodeficiency syndrome (AIDS) with more than 35 million people infected worldwide. Typically, after seroconversion, individuals enter a clinically stable and relatively asymptomatic phase that can last for years. Without antiretroviral treatment, individuals usually progress to AIDS, which is characterized by a decrease in CD4+ cells in the immune system, which increases the susceptibility to opportunistic infections and eventual death. Historically, HIV tests have relied on the antibody response that patients produce due to the virus infection. Although these antibodies are ineffective in combating the virus, they have been found in most chronically infected patients. In newborns, nucleic acid amplification tests can diagnose HIV infection during the first 18 months of a baby's life, while their blood still contains maternal antibodies and can result in a false-positive interpretation of serological tests. the availability of a diagnostic test capable of detecting HIV infection that offers flexibility, stability and ease of collection is of great importance. In this sense, the detection of HIV-1 in dried blood spots (DBS) represent a good strategy, with simple collection of sample and provides evidence of current infection using nucleic acid amplification technologies such as polymerase chain reaction (PCR). In this way, this work aimed to evaluate a qualitative test for detetion of HIV 1 in DBS. Methods Were selected 103 whole blood samples. The samples were tested from the qualitative HIV test (PCR In house) from the Pardini Group's Genetics sector. With the aid of a pipette, approximately 70 µl of blood from each sample were dispensed in duplicate, within the circle outlined on the DBS filter paper. DBS was previously identified with the sample data. The DBS with the sample was placed in a tray on a flat surface at room temperature (between 18 and 25°C) for at least 4 hours protected from direct light to dry. After drying, the pre-extraction of the sample was performed and later the samples were placed in the equipment on the cobas® 4800, using the COBAS HIV 1 kit (Roche diagnostics), (qualitative test) for extraction. After the extraction step was complete, the plate was transferred to the cobas z 480 analyzer for detection. All steps were performed according to the manufacturer's instructions. At the end of the tests, the results were compared between the in-house methodology and the cobas® HIV-1 qualitative test. Results The test cobas® HIV-1 qualitative test showed 100% agreement with the in-house methodology for 90/90 negative and 13/13 positive samples. When comparing the test matrix (whole blood or DBS), no significant differences were observed in relation to test sensitivity and specificity. In addition, test automation provided a substantial reduction in test execution time. Conclusion The availability of tests that use DBS as a matrix offer important advantages, especially for children and the elderly, due to greater stability, ease of sample collection and transport. The qualitative cobas® HIV-1 test in DBS has been shown to be a good choice for detecting HIV-1 in infected patients.