Plan apo 100 na 1.4 objective

Fixed samples were viewed with either a Nikon Eclipse 90i fluorescence microscope equipped with an oil-immersion Nikon plan Apo 100×/NA 1.4 objective, a Nikon plan Fluor 20×/NA 0.5 objective, or a Nikon plan Fluor 40×/NA 0.75 objective and a Hamamatsu GRCA-ER camera (Hamamatsu Photonics) or a Zeiss AxioImager M2 fluorescence microscope equipped with an oil-immersion Zeiss plan Apo 100×/NA 1.4 objective and a Hamamatsu ORCA-R2 camera. Optical z-sections with 0.2 µm spacing were acquired using Volocity 6.3.1 software acquisition module (Perkin Elmer).

Cells were imaged on a Zeiss Axioimager M2 fluorescence microscope with a z motor for acquisition of slices along the z-axis. An oil-immersion Zeiss plan Apo 100×/NA 1.4 objective and a Hamamatsu ORCA-R2 camera were used to acquire images with optical z-slices of 0.2 µm. Images were acquired, registry corrected, deconvolved, and brightness/contrast adjusted using Volocity imaging software version 6.3 or 6.3.1 (Perkin Elmer, Waltham, MA). Fluorescent foci in the PV were identified and counted manually after identifying the boundaries of the PV based on the change in fluorescence patterns of the fluorescently tagged proteins. Statistical differences were determined using unpaired, two-tailed T tests with the Welch’s correction for unequal SDs with Prism (Graphpad, San Diego, CA). PV were ranked as positive based on one or more distinct fluorescent foci detected within the bounds of the PV or negative if no foci could be observed inside the PV.