Giant Human T-Lymphocytes (Jurkat Cells) Expressing HIV-1 Tat-72: LM and TEM.

Abstract The human immunodeficiency virus (HIV-1) is dependent upon the synthesis of TAT for efficient transcriptional activation and replication. Following translocation to the nucleus TAT interacts with a cis-acting TAT-responsive element located between nucleotides +1 and +42 within the HIV-1 long terminal repeat. This region contains an RNA stem loop structure TAR which interacts with TAT and cellular proteins to increase transcription 1000 fold We have transfected human T-lymphocytes with an expression vector containing the HIV-1 LTR driving the synthesis of TAT-72. A stable cell line was generated by selection with G418. Cells were grown on round glass coverslips coated with a solution of fibronectin overnight (at 37 °C). In addition to light microscopy (LM) observations, cells on coverslips were fixed with a 3.5% glutaraldehyde buffered solution (0.1 M Na cacodylate) for 5 min at room temperature and gently centrifliged in a microfuge (400 rpm, 3min) to obtain a cellular pellet.