Orca er cooled

For immunofluorescence, frozen sections from biopsies of patients with common nevi, dysplastic nevi and primary malignant melanomas were fixed with acetone and blocked in 10% normal goat serum (Vector Laboratories) for 30 min. The tissue sections were incubated with mouse anti-human CD11c (clone Bly6, BD PharMingen) and rabbit anti-human SOCS2 (clone EPR2588(2), LifeSpan Biosciences), overnight at 4°C. The tissues were then amplified with goat anti-mouse Alexa Fluor 488 (Invitrogen, Eugene, OR, USA) and goat anti-rabbit Rhodamine Red X (Invitrogen, Eugene, OR, USA), for 30 min the next day. Images were acquired using the appropriate filters of a Zeiss Axioplan 2 widefield fluorescence microscope with a Plan Neofluar 20 9 0.7 numerical aperture lens and a Hamamatsu Orca ER cooled charge-coupled device camera, controlled by METAVUE software (MDS Analytical Technologies, Downington, PA, USA). Images in each figure are presented both as single color stains (green and red) located above the merged image, so that localization of two markers on similar or different cells can be appreciated. Cells that co-express the two markers in a similar location are yellow in color. A white line denotes the junction between epidermis and the dermis.

For fluorescence microscopy, worms at the L3–L4 stage were paralyzed with levamisole on an agar pad. All images were collected on a PerkinElmer Ultraview multispectral spinning disk confocal microscope equipped with a Zeiss 1.4 numerical aperture oil immersion ×63 objective lens and a Prior piezoelectric objective focusing device. Images were acquired with a Hamamatsu ORCA ER cooled CCD camera controlled with Volocity software. Post-acquisition image manipulations as well as Pearson correlation coefficient calculations were performed using Fiji software.^{39 (link)} Images shown in Figure 3C represent projections of acquired z-stacks.