Adult zebrafish were treated with 750 nM neocuproine in a beaker for 24 hr and then kept in individual tanks filled with fish water after drug washout. Prior to imaging fish were treated with 1 mg/ml (−)-epinephrine (+)-bitartrate salt (Acros Oragnics) for 2 min and then anesthetized with 0.17 mg/ml tricaine. Fish were then placed on their sides in a plastic Petri dish and imaged in the same locations over time using anatomical and cellular landmarks. Fish were viewed with a Leica DM550B microscope and images were captured with a Leica DFC365FX camera. Representative examples of direct differentiation (Animation 1 ) and symmetric division (Animation 2 ) fates are provided. The fish were then placed in fish water for recovery.
Dm550b microscope
Manufactured by Leica
The Leica DM550B is a microscope designed for laboratory applications. It features a binocular viewing head and is equipped with objectives for magnification and observation. The DM550B provides the basic functionality required for microscopic analysis and examination of samples.
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Lab products found in correlation
4 protocols using dm550b microscope
1
In Vivo Zebrafish Cardiomyocyte Imaging
2
Histological Analysis of Colon Sections
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Histology and immunohistochemical staining were performed on 5-µm thick formalin-fixed, paraffin-embedded colon sections. Colon sections stained with hematoxylin and eosin (H&E), Periodic acid–Schiff (PAS) stain, and Gram stain. Briefly, sections were deparaffinized, followed by antigen retrieval using the antigen retrieval solution (Agilent Technologies, Santa Clara, CA, U.S.A.). Endogenous peroxidase activity was quenched with 3% H2O2 in PBS. Sections were stained using specific primary antibody (Ki-67, Cell Signaling, Danvers, MA, U.S.A.) and VECTASTAIN ABC Kit and diaminobenzidine (Vector Laboratories, Burlingame, CA, U.S.A.). Counterstaining was performed with hematoxylin and stained sections were visualized by Leica DM550B microscope. Ki-67 immunostaining was quantified by counting Ki-67-positive cells in at least 20 crypts per mouse.
3
Pollen Viability and Germination Assay
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Pollen from all strains that produced anthers was tested for viability with a modified Alexander’s stain [52 (link)] and for the ability to form pollen tubes by gently spreading anthers on E solidified with agar. For staining, pollen from 5–10 anthers was stained, and at least 169 grains per anther were scored as viable or dead. Pollen tube formation was observed at 20× using a Leica DM550B microscope 1 h later. Fertility is described as the percent of pollen grains with germination tubes. At least 97 grains from 4–7 anthers were observed.
4
Immunohistochemical Analysis of Colonic Tissue
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Colons were excised and cleaned with phosphate buffered saline followed by fixation in neutral buffered formalin (Thermo Fisher, Waltham, MA, USA). The fixed colon tissues were embedded in paraffin and 5 μm thick sections were sliced and placed on glass microscope slides. Immunohistochemical staining was performed as described previously.8 (link) Briefly, sections were deparaffinized, followed by antigen retrieval using the antigen retrieval solution (Agilent Technologies, Santa Clara, CA, USA). Endogenous peroxidase activity was quenched with 3% H2O2 in phosphate buffered saline for 10 min. Sections were stained using specific primary antibodies and Vectastain ABC kit and diaminobenzidine (Vector Laboratories, Burlingame, CA, USA). Counterstaining was performed with hematoxylin and stained sections were visualized by Leica DM550B microscope. Alternatively, in some experiments, fluorescent dye-labeled secondary antibody was used and sections were visualized by LSM 510 (Zeiss) confocal microscopy.
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