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Figure 4 | Retrovirology

Figure 4

From: Localization, quantification and interaction with host factors of endogenous HTLV-1 HBZ protein in infected cells and ATL

Figure 4

Quantification of endogenous HBZ expression in HTLV-1 chronically infected and in ATL tumor cells. a Three serial dilutions of purified GST-tagged HBZ protein (HBZ.GST) in nanograms (ng) were prepared in lysis buffer, run in SDS-PAGE gel, blotted in nitrocellulose filter, and filter reacted with the 4D4-F3 anti-HBZ mAb. Similarly, 40 μl of 1 ml cell lysates from C5MJ, ATL-2s, PH961 patient ATL tumor cells [PH961(a) and PH96(b)], two distinct cell lysates of equivalent number of cells, 293T cells transfected with HBZ-myc (293T.HBZ) and Jurkat cells, were run in the same gel and processed as above. Cell lysates were from the same cells analyzed by confocal microscopy and shown in Figure 1. It must be underlined that in order to obtain comparable western blots, C5MJ, ATL-2s and PH961 patient cell lysates were prepared from 10 times more cells as compared to 293T.HBZ cell lysates; this corresponded to 330.000 cells for the first three cell lysates vs 33.000 cells for 293T.HBZ cell lysate. Jurkat cell lysate was from 35.000 cells. b The expression of endogenous α-tubulin, used as internal control of sample loading. In this case, lysates from equal number of cells of the various samples were loaded in the gel. c The calculated amounts of HBZ in picograms/cell and in number of molecules/cell for the distinct cell sample are listed.

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