Figure 3. Endogenous PAI colocalizes... | Rockefeller University Press

Figure 3.

$Figure 3. Endogenous PAI colocalizes with tPA on the cell surface after stimulation. Cultured bovine chromaffin cells were incubated for 10 s in buffer with (A and B) or without (C–E) 56 mM K+ at 34°C. The solution was replaced with buffer containing 5.6 mM K+, and the cells were immediately placed on ice. Cells were then incubated with antibodies to tPA (A and C) and PAI (B and D) for 60 min on ice, and then processed and imaged by confocal microscopy. (A and B) Arrowheads indicate instances of colocalization of secreted tPA and PAI. When the fraction of puncta with colocalized tPA and PAI was calculated for n = 12 cells, it ranged from 67 to 90%, with a mean of 77.2 ± 2.2%. n = 333 total puncta. (C–E) Unstimulated cells, which were processed for tPA and PAI and visualized as in A and B, have little or no secreted tPA or PAI on the plasma membrane. Images that are to be compared directly (e.g., A and C; B and D) were acquired at the same microscope settings and adjusted to the same brightness and contrast when making the figures. Bars, 2 µm.$

Endogenous PAI colocalizes with tPA on the cell surface after stimulation. Cultured bovine chromaffin cells were incubated for 10 s in buffer with (A and B) or without (C–E) 56 mM K⁺ at 34°C. The solution was replaced with buffer containing 5.6 mM K⁺, and the cells were immediately placed on ice. Cells were then incubated with antibodies to tPA (A and C) and PAI (B and D) for 60 min on ice, and then processed and imaged by confocal microscopy. (A and B) Arrowheads indicate instances of colocalization of secreted tPA and PAI. When the fraction of puncta with colocalized tPA and PAI was calculated for n = 12 cells, it ranged from 67 to 90%, with a mean of 77.2 ± 2.2%. n = 333 total puncta. (C–E) Unstimulated cells, which were processed for tPA and PAI and visualized as in A and B, have little or no secreted tPA or PAI on the plasma membrane. Images that are to be compared directly (e.g., A and C; B and D) were acquired at the same microscope settings and adjusted to the same brightness and contrast when making the figures. Bars, 2 µm.