Figure 5 | Scientific Reports

Figure 5

From: Selective cytotoxicity of a novel immunotoxin based on pulchellin A chain for cells expressing HIV envelope

Figure 5

Binding, internalization and intracellular localization of fluorescent ITs. (A) Env-transfected 293 T cells were incubated with cold PBS in the presence or absence of 0.02% sodium azide. 45 min later, Alexa488-ITs were separately added to each set of slides in the presence of sCD4-IgG2. The slides were counterstained with BFA-bodipy, incubated an additional 60 min under the same conditions. Five min later, cells were washed 3X with cold PBS in the presence or absence of azide. The cells were fixed in 2% paraformaldehyde or one drop SlowFade Gold Mountant DAPI. Cells were observed with a 62X oil-immersion objective. Z-stack images collected at 1 µm sections. The bottom of cells represents the closest plane to the slide. ITs are green, ER and Golgi are red. Some samples have nuclei stained with Hoechst Dye (Blue). Colocalization of red and green dyes appears yellow. The white bar indicates 10 µm. (B,C) A comparison between Env-transfected 293 T cells (B) and 293 T cells (C) incubated with the same description in panel (A) but in the absence of sodium azide, in order to show the specific targeting the transfected cells. BFA-bodipy was used to demonstrate accumulation of ITs on the ER and Golgi.