Leica dm4000b microscope

The EOs were subjected to anatomical examinations via light microscopy. Three F2-hybrids were fixed shortly after death by immersion in 4% formalin and the caudal peduncle was cut off. Dehydration and embedding in paraffin followed conventional techniques (Mulisch and Welsch 2010 ). The EO was sectioned serially at 4–6 µm thickness in the transverse or sagittal plane using a Leica 2035 Biocut microtome. Sections were mounted on glycerin/albumin-coated slides. Following deparaffination and rehydration, sections were stained with Azan or HE (Mulisch and Welsch 2010 ), dehydrated and coverslipped with DePeX mounting medium (Sigma-Aldrich). Sections were investigated with a Leica microscope DM4000B. Pictures were taken and processed using the Leica DFC 480 camera together with the Leica IM software version 4.0.

For immunofluorescence microscopy, each sample was sectioned using a Leica vibratome (VT1000S, Leica, Wetzlar, Germany) to yield 60–80-µm transversal cuts. The sections were washed three times in PBS using a 24-well plate and blocked with MP/PBS for 1 h. The sections were then incubated with a primary monoclonal antibody for 1 h. The samples were probed with three different monoclonal antibodies: (a) xylan CRCC M140 antibody [35 (link)] diluted 1:10 in MP/PBS; (b) arabinoxylan LM11 antibody [36 (link)] diluted 1:10 in MP/PBS; and (c) MLG antibody [44 (link)] diluted 1:1500 in MP/PBS. After antibody binding, the samples were washed three times with PBS and incubated with secondary antibodies (Alexa Fluor 514—Sigma/Aldrich anti-mouse for CRCC M140 and MLG and Alexa Fluor 568—Sigma/Aldrich anti-rat for LM11), both diluted 1:10,000 in MP/PBS. The sections were visualized with a Leica microscope (DM 4000 B) under YFP filter for Alexa Fluor 514 and TX2 filter for Alexa Fluor 568. Sections treated only with secondary antibody were used as negative controls. Images were processed with ImageJ software (version 10.2).

The TMA with 125 NPC patients were purchased from Superbiotek (Shanghai, China). IHC determined the expression of NSUN2 in NPC and normal tissues as previously described (16 (link)). After deparaffinizing and blocking endogenous peroxidase, the slides were incubated with antibody against NSUN2 (1:800 dilution, 20854-1-AP, Proteintech, Wuhan, China). Arrays were then washed with PBS and incubated for 30 min with the EnVision™+ Dual Link System-HRP (Dako, Carpinteria, CA). After rinsing thrice with PBS for 3 min each time, the slides were incubated with DAB reagent (Dako, Carpinteria, CA) for 3–5 min and evaluated under a light microscope. The TMA were then counter-stained with hematoxylin and observed under a Leica microscope (DM4000b, Wetzlar, Germany). The results were evaluated independently by two blinded pathologists according to the following scoring criteria: 0, negative; 1, weak; and 2–3, strong. The degree of staining was scored as the proportion of the positive staining area relative to the whole cancerous area: 0, <5%; 1, 5–25%; 2, 26–50%; 3, 51–75%; and 4, >75%.

The histology of mouse testicular tissue fragments (S2 File, Panel A) was described and categorized according to the table in S2 File—Panel B and plotted in the graph (S2 File, Panel C).
Images from domestic cat testis sections of 6 animals and one hundred and fifty organ culture samples (n = 6 x 5 conditions x 5 time points) were taken with a Leica camera (DFC480) coupled to a Leica Microscope (DM4000B) at 200x magnification. After random selection of approximately 50% of the images taken from each sample, Image J software was used to quantify all PGP9.5 positive cells present in the image (corresponding to germ cell number, as all germ cells present in the tissue were labeled by PGP9.5) and the area (measured in arbitrary units) covered by seminiferous tubules, necrotic and interstitial tissue using a grid with 1.5 square inches. The area covered by the different types of tissue was represented as percentage of the total area measured (% of seminiferous tubules, % of necrotic tissue and % of interstitial tissue and the number of germ cells were normalized in terms of the area covered by seminiferous tubules (number of germ cells divided by area of seminiferous tubules).

The EOs were subjected to anatomical examinations via light microscopy. For the preparation of the histological slides, the fish were euthanized by an overdose of ethylenglycolmonophenylether and the caudal peduncle was cut off. The tissue was fixed by immersion in 4% formalin in the case of small fish or in 10% formalin in larger fish. Fixed tissue of larger fish was decalcified with 7.5% EDTA (pH 7.4). Dehydration and embedding in paraffin followed established protocols (Mulisch and Welsch 2010 ). The EO of each specimen was sectioned serially at 4–6 µm either in the transverse or the sagittal plane using a Leica 2035 Biocut microtome. Sections were mounted on glycerin/albumin-coated slides. Following deparaffinization and rehydration, sections were stained with Azan or HE (Mulisch and Welsch 2010 ), dehydrated and coverslipped with DePeX mounting medium (Sigma-Aldrich). Sections were investigated with a Leica microscope DM4000B. Pictures were taken and processed by using the Leica DFC 480 camera together with the Leica IM software version 4.0.