What is HIV and Why Monitor Viral Load?

HIV (Human Immunodeficiency Virus) is the causative agent of AIDS, a disease that severely compromises the immune system. HIV attacks and replicates within CD4+ T cells, weakening the host’s ability to fight infections. If left untreated, HIV can lead to life-threatening opportunistic infections and malignancies.

HIV is transmitted through infected blood, semen, vaginal fluids, and breast milk. It is a global public health concern, and accurate monitoring of HIV viral load is essential for treatment planning and disease progression tracking.

The HIV viral load reflects the amount of viral RNA in the bloodstream. Routine viral load testing helps clinicians:

• Assess disease progression

• Monitor antiretroviral therapy (ART) effectiveness

• Detect treatment failure early

• Guide treatment switching decisions

Role of gag and pol Targets in HIV Diagnostics

The TRUPCR® HIV Viral Load Kit amplifies two highly conserved regions of the HIV-1 genome:

• gag gene: encodes structural proteins such as the p24 core protein, a critical marker for viral replication

• pol gene: encodes enzymes like reverse transcriptase and integrase, key targets for ART

Targeting both gag and pol enhances the assay’s sensitivity and specificity. By reducing the risk of false negatives from sequence variability or mutations, this dual-target approach strengthens the reliability of patient monitoring over time.

About the TRUPCR® HIV Viral Load Kit

This quantitative real-time PCR test is designed to provide clinicians with precise and reproducible measurement of HIV-1 RNA levels. It uses TaqMan hydrolysis probe technology, which allows for fluorescence-based quantification of amplified cDNA, with real-time tracking of each amplification cycle.

Each test tube contains:

• Primers and probes targeting HIV-1 gag and pol

• Internal control probe (to detect PCR inhibition)

• cDNA synthesis reagents

• Real-time PCR master mix with hot start Taq polymerase and Uracil-DNA Glycosylase (UDG) to prevent carry-over contamination

This design ensures a closed-tube system, minimizing hands-on steps and reducing contamination risks. The kit is RUO-labeled, ideal for research settings and laboratory-developed tests (LDTs).

Principle and Workflow

The assay workflow follows a two-stage RT-PCR process:

1. RNA Extraction: HIV-1 RNA is extracted from serum or plasma samples.

2. cDNA Synthesis: Using reverse transcriptase, RNA is converted to complementary DNA (cDNA).

3. Amplification: The cDNA is amplified using Taq polymerase, with fluorescence generated by oligonucleotide probes.

4. Detection: Fluorescence signals (FAM, HEX, etc.) are measured in each cycle.

5. Quantification: The Ct value correlates with viral load and is analyzed against standard controls.

The internal control verifies that the reaction occurred correctly, ruling out inhibitors or extraction errors.

Clinical Application

The TRUPCR® HIV Viral Load Kit is ideal for:

• Routine HIV-1 viral load monitoring in diagnostic laboratories

• Research and clinical trials evaluating ART efficacy

• Baseline viral load assessment for newly diagnosed patients

• Monitoring post-treatment rebound or virological failure

Its low detection limit (100 IU/mL) makes it suitable for early detection of rebound or residual viremia, supporting aggressive monitoring in patients under suppressive therapy.

The kit is validated across major real-time PCR platforms, including:

• ABI 7300 / 7500

• QuantStudio™ 3 & 5

• Bio-Rad CFX96™

• Rotor-Gene Q

• StepOne™ / StepOnePlus™

This compatibility ensures flexible integration into existing molecular diagnostic workflows.