Dmlb light microscope

To examine the morphology of the hippocampus, hippocampal tissue sections were stained with Hematoxylin and Eosin (H&E), and visualized with a Leica DMLB light microscope under a 10X objective. The images shown were acquired with a Leica DC 300 camera (Leica Microsystems AG).

Shoot apices and young leaves were fixed in formalin–acetic acid–alcohol (FAA) solution for 24 h [62 ] and buffered neutral formalin in 0.1 M sodium phosphate buffer (pH 7.0) for 48 h [63 ]. For the micromorphological study, mature nectaries were isolated, dehydrated in a graded ethanol series, dried by the critical point method, mounted on aluminium stub, and sputter-coated with gold, with subsequent observation in a Zeiss Sigma VP scanning electron microscope (Carl Zeiss, Oberkochen, Germany).
For anatomical analyses, shoot apices and young leaves were isolated, dehydrated through a tertiary butyl alcohol series [62 ], embedded in Paraplast^® (Leica Microsystems Inc., Heidelberg, Germany), and serial-sectioned at a 12 µm thickness on a Leica RM2145 rotary microtome. Longitudinal and transverse sections were stained with astra blue and safranin O [64 ] and the slides were mounted with resin Permount (Fisher Scientific, Pittsburgh, PA, USA). Observations and photographs were performed using a Leica DMLB light microscope.

Cells were fixed in 1% paraformaldehyde (v/v) for 25 minutes, and then washed in PBS. Ceramide labelling was conducted using an antibody from Alexis (San Diego, CA. USA). Cells that were stained for UGT8 (antibody was obtained from Abcam, Cambridge, MA. USA) were permeabilized and then the antigenic sites were un-masked using sodium citrate buffer. Immunostaining was then done using a Histostain-plus kit (Invitrogen, Carlsbad, CA) following the manufacturer’s instructions, previously described [3 (link), 6 (link)]. Images were acquired using LEICA DM LB light microscope and Leica IM1000 software.
Tumour tissues were sliced into two halves. One half was embedded in optimal cutting temperature (OCT) media, and stored at -80 C. Cryo- 8μm sections were prepared, and used for ceramide labelling as described above. The other half was fixed in freshly prepared 1% paraformaldehyde (v/v). Histopathology was assessed using hematoxylin and eosin (H&E) staining.
Tumour cell death was evaluated using a TUNEL assay and fibrosis was revealed using Masson trichrome staining. TUNEL assay results were evaluated using Image J as was previously described [6 (link), 8 (link)], where areas of cell death were quantified relative to the total area of tumour sections.

Cysts and infective juvenile stages were examined at the USDA MNGDBL and at the University of Nebraska Nematology Laboratory. Select juvenile measurements are presented in Table 2 alongside measurements from Gerber and Maas’s (1982) redescription. Images of juveniles and adult males were taken with a Leica DMLB light microscope with differential interference contrast optics and a Leica DC300 video camera. All juveniles examined at the University of Nebraska were provided an identification number which links specimen images, measurements, and placement on phylogenetic trees. Cysts were prepared for scanning electron microscopy by fixation in a 4% formalin solution followed by a graded series of alcohol to 100% ethyl alcohol prior to critical point drying and coating with gold. Images were obtained on a Hitachi S-3000N scanning electron microscope located in the Morrison Microscopy Core Research Facility at the University of Nebraska.

Immunohistochemistry was viewed with a Leica DMLB light microscope (Milton Keynes, UK). Analysis across five random fields of view was performed using the Leica digital image capture program. Values are expressed as a percentage of total immunoperoxidase staining.