Axiocam monochrome digital camera

Differential interference contrast and fluorescent images were captured with an Axioimager A1 (ZEISS) equipped with epifluorescence (Filter Set 13 for GFP [excitation BP 470/20, beam splitter FT 495, emission BP 503–530] and Filter Set 20 for Cherry [excitation BP 546/12, beam splitter FT 560, emission BP 575–640]) and an AxioCam monochrome digital camera (ZEISS). Images were processed and viewed using Axiovision Rel. 4.7 software (ZEISS). A 100× Plan-Neofluar objective (NA1.30) was used with Immersol 518F oil (ZEISS). Confocal images were captured by an LSM 5 Pascal (ZEISS) inverted confocal microscope with 488-nm (emission filter BP 503–530) and 543-nm (emission filter BP 560–615) lasers, and images were processed and viewed using LSM Image Browser software (ZEISS). All images were taken at 20°C.
To quantify the fluorescence intensity of LMP-1::sfGFP, 15 images were taken of each cell type with equal exposure time and quantified using ImageJ 1.42q (National Institutes of Health).

Cell corpses were identified by their raised button–like morphology using Nomarski optics. The number of somatic cell corpses in the head region of living embryos was quantified either at different embryonic stages for time course analysis or at the twofold stage as indicated in the legends of Figs. 1, S1, and S2. At least 15 animals were quantified at each stage in each strain. To examine cell corpse duration, embryos at the precomma stage were mounted on agar pads. Images in 30 z series (1.0 µm/section) were captured every 2 min for 2–4 h using an Axioimager M1 microscope equipped with an AxioCam monochrome digital camera (Carl Zeiss Inc.). Images were processed and viewed using Axiovision Rel 4.7 software.
To examine phagosome maturation, differential interference contrast (DIC) and fluorescence images were captured using an Axioimager A1 microscope (Carl Zeiss Inc.). The total number of cell corpses and the number of cell corpses that were labeled by different phagosomal markers were scored using DIC and fluorescent microscopy. The percentage of cell corpses labeled by phagosomal markers was calculated as (number of labeled cell corpses/total number of corpses) × 100.

Embryos mounted on 2% agar pads were anesthetized in 3 µl M9 buffer (1 liter contains 3 g KH₂PO₄, 6 g Na₂HPO₄, 5 g NaCl, and 1 mM MgSO₄) containing 33 mM sodium azide. The numbers of button-like cell corpses and pit-like vacuoles in the head region of living embryos at various developmental stages (comma, 1.5-fold, 2-fold, 2.5-fold, 3-fold, and 4-fold) were scored using DIC optics. 15 embryos at each developmental stage were scored for each strain. To examine embryonic cell corpse duration, embryos at the two-cell stage were mounted on 2% agar pads in egg salt buffer (118 mM NaCl and 48 mM KCl) at 20°C. Images in 30 Z-sections (1.0 µm/section) were captured every minute for 400 min using an Axioimager M2 microscope (ZEISS) equipped with an AxioCam monochrome digital camera (ZEISS). Images were processed and viewed using ZEN 2 pro software (ZEISS).