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Isolation of HIV from cultures of purified CD4+ lymphocytes
C Nielsen1, C M Nielsen, J L Petersen
1Department of Virology, Statens Seruminstitut (SSI), Copenhagen, Denmark.
Insights
Optimized HIV isolation from CD4+ lymphocytes achieved 98% success in a single attempt. Culture time was independent of CD4 count, with most positive results within 14 days.
Area of Science:
- Virology
- Immunology
- Cell Biology
Background:
- Human Immunodeficiency Virus (HIV) infection requires sensitive diagnostic methods.
- Efficient isolation of HIV from patient samples is crucial for research and clinical monitoring.
- CD4+ lymphocytes are a primary target for HIV replication.
Purpose of the Study:
- To optimize and evaluate a method for isolating infectious HIV from CD4+ lymphocytes.
- To assess the efficiency of HIV isolation across different patient groups.
- To investigate the impact of media additives on HIV propagation.
Main Methods:
- Isolation of CD4+ lymphocytes from peripheral blood using indirect panning.
- Culturing of lymphocytes for HIV detection.
- Evaluation of media additives including amphotericin-B, interleukin-2, and polybrene.
Main Results:
- Successful HIV isolation in 98% of 102 HIV-antibody-positive patients (including those with AIDS/ARC and clinically healthy individuals).
- Culture time to positive isolation was largely independent of patient CD4 count.
- 87% of positive cultures became positive within 14 days.
- Amphotericin-B suppressed HIV replication, while interleukin-2 and polybrene showed beneficial effects on HIV propagation.
Conclusions:
- Optimized indirect panning provides a highly efficient method for HIV isolation from CD4+ lymphocytes.
- The method is effective across a spectrum of HIV-positive patients.
- Specific media additives can significantly influence HIV isolation and propagation success.
Abstract:
Isolation of HIV from cultures of CD4+ lymphocytes purified from peripheral blood by indirect panning was optimized and evaluated. Infectious HIV was isolated by single isolation attempts in 98% of 102 HIV-antibody-positive patients (55 had AIDS or ARC and 47 were clinically healthy). The average culture time required for positive cultures was largely independent of the CD4 count of the patients and 87% of the positive isolation cultures from both groups of patients became positive within 14 days of culture. An evaluation of the possible influence of media additives on propagation of HIV showed that: amphotericin-B had a suppressive effect on HIV replication at concentrations recommended for anti-fungal activity; recombinant and human interleukin-2 were equally suitable for both isolation cultures and for propagation of HIV, and polybrene, at a concentration of 2 micrograms/ml in the culture medium had a beneficial effect.