Optiphot 2 microscope

Manufactured by Nikon

Sourced in Japan, United States

The Optiphot-2 is a microscope designed for laboratory use. It features a binocular observation head and a multi-position nosepiece that can accommodate a variety of objective lenses. The microscope is capable of providing magnification capabilities for various applications in a research or educational setting.

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Lab products found in correlation

Dibutyl phthalate, by Merck Group (2 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (2 mentions) Lactophenol cotton blue, by Merck Group (2 mentions) Zoe fluorescent cell imager, by Bio-Rad (2 mentions) Eclipse ni fluorescence microscope, by Nikon (2 mentions) Image pro plus 6, by Media Cybernetics (2 mentions) Coolsnap pro camera, by Media Cybernetics (2 mentions) Entellan, by Merck Group (2 mentions) Alexa fluor 488 phalloidin, by Thermo Fisher Scientific (1 mentions) Poly l lysine (pll), by Merck Group (1 mentions) Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions) Ihc reagents, by Biocare Medical (1 mentions) Nis elements software, by Nikon (1 mentions) Nis elements 2, by Nikon (1 mentions) Calcein, by Merck Group (1 mentions) Cfw 1308c, by Techcomp Instruments (1 mentions) Sp8 confocal microscope, by Leica camera (1 mentions) Eclipse e800, by Nikon (1 mentions) Stereo investigator software, by MicroBrightField (1 mentions) Dp70 camera, by Olympus (1 mentions)

66 protocols using optiphot 2 microscope

Standardized Morphometric Analysis Pipeline

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All morphometric measures were performed using Stereo Investigator software (version 9.14.5 32-bit, MicroBrightField, Inc., Williston, VT, USA) coupled to a Nikon Optiphot-2 microscope, as described previously (Kelley, et al., 2013 ). To control for inter-individual variability, a single investigator preformed all analyses. To ensure low intra-individual variability, five mice chosen at random were measured non-consecutively three times during each triad (beginning, middle, end) of experiment. No significant difference was found across these time points. All analyses were conducted blinded to genotype and treatment.
Photomicrographs were taken on a Nikon Optiphot-2 microscope (Tokyo, Japan) connected to Stereo Investigator software (MicroBrightField, Inc.) (Figs. 1, 2, 3, 6). Background correction was used at the time of image capture to establish evenness of illumination across the field, and scale bars were added within the Stereo Investigator software. Panels were compiled in PowerPoint (version 14.0.6129.5000, 32-bit, Microsoft, Redmond, WA, USA), and in Figs. 1, 2, and 3 each micrograph was equally corrected for brightness and contrast. No retouching or further manipulations were performed.

Kelley C.M., Powers B.E., Velazquez R., Ash J.A., Ginsberg S.D., Strupp B.J, & Mufson E.J. (2014). Maternal choline supplementation differentially alters the basal forebrain cholinergic system of young-adult Ts65Dn and disomic mice. The Journal of comparative neurology, 522(6), 1390-1410.

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Bone Histomorphometry Analysis in Mice

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The WT and cMpl−/− mice were administered an intraperitioneal injection of the fluorochrome calcein (30mg/kg) 13 and 3 days prior to sacrifice to label actively forming bone surfaces. Static and dynamic histomorphometric analysis of trabecular bone was performed on femurs as previously described (Feher et al.,2010 (link); Warden et al.,2008 (link)). Histological measurements were made with a semiautomatic analysis system (Bioquant OSTEO 7.20.10, Bioquant Image Analysis Co.) attached to a microscope with an ultraviolet light source (Nikon Optiphot 2 microscope, Nikon). Measurements were done on one stained (static) and one unstained (dynamic) section for each animal.

Meijome T.E., Baughman J.T., Hooker R.A., Cheng Y.H., Ciovacco W.A., Balamohan S.M., Srinivasan T.L., Chitteti B.R., Eleniste P.P., Horowitz M.C., Srour E.F., Bruzzaniti A., Fuchs R.K, & Kacena M.A. (2015). C-Mpl is expressed on osteoblasts and osteoclasts and is important in regulating skeletal homeostasis. Journal of cellular biochemistry, 117(4), 959-969.

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Visualizing Immune Synapse Formation

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HEK cells transfected with hCD48 or the corresponding empty vector were first labelled with CellTracker Blue CMAC. After washing, labelled HEK cells were mixed for 10 min at 37°C and a final volume of 500 μL with 1x10⁵ YT cells at a ratio of 1:1 to allow conjugate formation. The suspension was placed on coverslips coated with poly-L-lysine (Sigma-Aldrich) for 10 min at 37°C, washed with PBS, fixed with 4% formaldehyde for 10 min, and permeabilized with 0.05% Triton X-100 for another 10 min. Subsequently, samples were blocked with 20% rabbit serum and 6% fetal bovine serum in PBS and incubated with Alexa Fluor 488 Phalloidin (Invitrogen) and the primary anti-perforin mAb, followed by a secondary antibody goat anti-mouse IgG (H+L)-Alexa Fluor 555. Coverslips were mounted in ProLong Gold antifade reagent (Invitrogen). Fluorescence images were acquired using a Nikon Optiphot-2 microscope (Nikon Corp.). Synapse stages were defined as: 0, conjugates lacking actin polymerization and perforin polarization; 1, conjugates with polymerized actin, but missing perforin polarization; 2, conjugates with actin polymerization and perforin clustered at the immune synapse. When indicated, NK cells were pre-incubated with 10 μg/mL of A43-Fc or CTL-Fc fusion proteins, at 37°C during 30 min prior to being mixed with the target cells.

Martínez-Vicente P., Farré D., Sánchez C., Alcamí A., Engel P, & Angulo A. (2019). Subversion of natural killer cell responses by a cytomegalovirus-encoded soluble CD48 decoy receptor. PLoS Pathogens, 15(4), e1007658.

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Immunohistochemical Staining of Aortic Valves

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For staining of aortic valves, IHC reagents were obtained from Biocare Medical (Pacheco, CA, USA). Isotype rabbit IgG was used as negative control. In brief, 10-μm sections were deparaffinized in Tissue Clear and rehydrated in ethanol. For antigen retrieval, slides were subjected to high-pressure boiling in DIVADecloaker heat retrieval solution (pH 6.0) or TE buffer (pH 9.0). After blocking with Background Sniper, primary antibodies diluted in Da Vinci Green solution were applied and incubated at room temperature for 1 h. A double-stain probe-polymer detection kit (Mach 2) containing both alkaline phosphatase and horseradish peroxidase was applied, with subsequent detection using Warp Red (for PRG4) and Vina Green (for BMP2 and RUNX2). All slides were counterstained with Hematoxylin, dehydrated, and mounted in Pertex (Histolab, Gothenburg, Sweden). Images were taken using a Nikon OPTIPHOT-2 microscope equipped with digital camera and NIS-Elements software (Nikon Instruments Inc., Melville, NY, USA). The following primary antibody was used in the study: anti-PRG4 (HPA028523; Sigma, St. Louis, MO, USA).

Artiach G., Carracedo M., Seime T., Plunde O., Laguna-Fernandez A., Matic L., Franco-Cereceda A, & Bäck M. (2020). Proteoglycan 4 is Increased in Human Calcified Aortic Valves and Enhances Valvular Interstitial Cell Calcification. Cells, 9(3), 684.

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Epifluorescence Microscopy of Click-labeled Cells

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After the click chemistry reaction, samples were filtered onto black polycarbonate 0.2 μm filters (Whatman, UK), stained with SYBR Green, and rinsed with sterile 1x PBS. Samples were counted in a Nikon Optiphot-2 microscope (Nikon Instruments, Japan) with Hg lamp C-SHGI (Nikon Instruments, Japan). Images were obtained with NIS-Elements 2.20 (Nikon Instruments). The fluorescence filters used were B-2A (Ex450-490) for AF488 azide and SYBR Green, and G2-A (Ex510-560) for LDS 751.

Lindivat M., Larsen A., Hess-Erga O.K., Bratbak G, & Hoell I.A. (2020). Bioorthogonal Non-canonical Amino Acid Tagging Combined With Flow Cytometry for Determination of Activity in Aquatic Microorganisms. Frontiers in Microbiology, 11, 1929.

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Quantifying Epidermal Thickness and p-STAT3

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Formaldehyde-fixed, paraffin-embedded ear skin samples were stained with H&E using standard procedures. Images were acquired using a Nikon Optiphot 2 microscope (Nikon, Tokyo, Japan). Epidermal thickness was measured with a computer-assisted image analysis software. For immunohistochemical analysis, skin sections were incubated with primary antibodies against murine p-STAT3, followed by the appropriate secondary antibodies. Rinsed sections were counterstained with hematoxylin.

Yamada D., Vu S., Wu X., Shi Z., Morris D., Bloomstein J.D., Huynh M., Zheng J, & Hwang S.T. (2022). Gain-of-function of TRPM4 predisposes mice to psoriasiform dermatitis. Frontiers in Immunology, 13, 1025499.

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Verifying Transformants via PCR

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The obtained transformants were verified by three passes on their respective plates containing the antibiotics. DNA extraction was then performed by grinding frozen mycelia and adding phenolchloroform [23 ]. Next, 100 ng of the extracted DNA was used as a template to verify the transformants using PCR to detect the presence of the corresponding resistance and fluorescence cassettes. The nourseothricin-resistance cassette was amplified using the oligonucleotides PNAT-XbaI and 5NAT-BglII, the eGFP cassette was amplified using the oligonucleotides 5CST-EGFP and 3CST-EGFP, the phosphinothricin-resistance cassette was amplified using the oligonucleotides PtrpC-XbaI (5′-ATTCTAGAATCGACAGAAGATGATATTGAAGGA-3′) and Bar-down (5′-TCAGATCTCGGTGACGGGC-3′), and the mCherry cassette was amplified using the oligonucleotides Pgpdashort (5′-GCGGAGAGACGGACGGAC-3′) and cherry-down (5′-TTACTTGTACAGCTCGTCCATGCC-3′). Protein expression of the fluorescent reporter protein was confirmed by visual inspection using a Nikon OPTIPHOT-2 microscope (Nikon, Tokyo, Japan) equipped with epifluorescent illumination.

Padilla-Guerrero I.E, & Bidochka M.J. (2017). Agrobacterium-Mediated Co-transformation of Multiple Genes in Metarhizium robertsii. Mycobiology, 45(2), 84-89.

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Femoral Bone Histomorphometry Analysis

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Static and dynamic histomorphometric analyses of cortical and trabecular bone were performed on the right femur (mid-shaft and distal regions) as previously described[20 (link),21 (link)]. Briefly, bones were embedded, undecalcified in 99% methyl-methacrylate with 3% dibutyl phthalate (Sigma-Aldrich, St. Louis, Mo, USA). Transverse thick sections (~40 microns) were taken from the mid-shaft using a diamond bladed wire saw, and frontal plane thin sections (6 microns) were obtained from the distal femur using a microtome. Histological measurements were made with a semiautomatic analysis system (Bioquant OSTEO 7.20.10, Bioquant Image Analysis Co.) attached to a microscope with an ultraviolet light source (Nikon Optiphot 2 microscope, Nikon). For dynamic measures of bone formation the following data was derived: mineralizing surface (MS/BS, %), mineral apposition rate (MAR, μm/day), and bone formation rate (BFR/BS; μm³/mm² year). For static measures of the distal femur osteoid surface/bone surface (OS/BS, %), number of osteoblast (N.Ob/T.Ar), and number of osteoclasts (N.Oc/T.Ar) were obtained from von Kossa/McNeal’s stained tissue. Measurements were completed on one von Kossa/McNeal’s stained (static) and one unstained (dynamic) section for each animal. The reader was blinded to the animal genotype.

Hooker R.A., Chitteti B.R., Egan P.H., Cheng Y.H., Himes E.R., Meijome T., Srour E.F., Fuchs R.K, & Kacena M.A. (2015). Activated leukocyte cell adhesion molecule (ALCAM or CD166) modulates bone phenotype and hematopoiesis. Journal of Musculoskeletal & Neuronal Interactions, 15(1), 83-94.

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Dynamic Bone Histomorphometry in Mice

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The C57BL/6, GATA-1^low/low, OPG^−/−, and GATA-1^low/low X OPG^−/− mice were administered an intraperitioneal injection of the fluorochrome calcein (30mg/kg) 13 and 3 days prior to sacrifice to label actively forming bone surfaces. At sacrifice the right femur was removed and stored in 10% neutral buffered formalin (NBF) for 48 hours, and then transferred to 70% ethanol (EtOH) for storage prior to processing. Static and dynamic histomorphometric analysis of trabecular bone was completed on the distal femur as previously described (Feher, et al., 2010 (link), Warden, et al., 2008 (link)). Tartrate resistant acid phosphatase staining was used to identify osteoclasts in the distal femur as detailed previously (Feher, et al., 2010 (link), Warden, et al., 2008 (link)). Histological measurements were made with a semiautomatic analysis system (Bioquant OSTEO 7.20.10, Bioquant Image Analysis Co.) attached to a microscope with an ultraviolet light source (Nikon Optiphot 2 microscope, Nikon). Measurements were done on one von Kossa/McNeal’s stained (static) and one unstained (dynamic) section for each animal. All analyses were performed blinded to treatment group.

Meijome T.E., Hooker R.A., Cheng Y.H., Walker W., Horowitz M.C., Fuchs R.K, & Kacena M.A. (2015). GATA-1 Deficiency Rescues Trabecular but not Cortical Bone in OPG Deficient Mice. Journal of cellular physiology, 230(4), 783-790.

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Bone Histomorphometry Analysis of WT and Lnk-/- Mice

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Actively forming bone surfaces of WT and Lnk^−/− mice were labeled by intraperitoneal (IP) injections of the fluorochrome calcein (30mg/kg; Sigma) at 13 and 3 days prior to sacrifice. At sacrifice, the left femur was removed and stored in 10% neutral buffered formalin for 24-hours, followed by storage in 70% alcohol. The right femur was removed, wrapped in saline soaked gauze and stored at −80C for biomechanical analysis. Femoral trabecular (distal region) and cortical (mid-shaft region) bone was analyzed by static and dynamic histomorphometry as previously described [Feher et al., 2010 (link); Warden et al., 2008 (link)]. Mid-sagittal sections of 8 μm were cut by a Reichert-Jung 2050 microtome (Magee Scientific, Inc) and stained with McNeal’s tetrachrome for static histomorphometry. Mid-sagittal sections of 8 μm were cut and left unstained for dynamic histomorphometry and wall thickness analysis. A semiautomatic analysis system (Bioquant OSTEO 7.20.10, Bioquant Image Analysis Co.) attached to a microscope with an ultraviolet light source (Nikon Optiphot 2 microscope, Nikon) was used to make histological measurements on one stained (static) and unstained (dynamic) section for each animal.

Olivos DJ I.I.I., Alvarez M., Cheng Y.H., Hooker R.A., Ciovacco W.A., Bethel M., McGough H., Yim C., Chitteti B.R., Eleniste P.P., Horowitz M.C., Srour E.F., Bruzzaniti A., Fuchs R.K, & Kacena M.A. (2017). Lnk Deficiency leads to TPO-Mediated Osteoclastogenesis and Increased Bone Mass Phenotype. Journal of cellular biochemistry, 118(8), 2231-2240.

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