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Ix71 inverted research microscope

Manufactured by Olympus
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Sourced in United States, Japan

The IX71 is an inverted research microscope designed for a variety of laboratory applications. It features an upright optical path and a stage that is positioned above the objective lenses. The IX71 is capable of various imaging techniques, including bright-field, phase contrast, and fluorescence microscopy.

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Lab products found in correlation

Dp30bw, by Olympus (2 mentions) Crystal violet powder, by Merck Group (1 mentions) Uplanfln objective, by Olympus (1 mentions) Analysis research acquisition software, by Olympus (1 mentions) Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488 conjugated antibody, by Thermo Fisher Scientific (1 mentions) Bz 9000e, by Keyence (1 mentions) Nepa21, by Nepa Gene (1 mentions) Sp5 confocal microscope, by Leica (1 mentions) Stx stage top incubator, by Tokai Hit (1 mentions) Paraformaldehyde (pfa), by Merck Group (1 mentions) Alexa fluor 488 donkey anti mouse igg h l, by Thermo Fisher Scientific (1 mentions) Dapi nucleic acid stain, by Thermo Fisher Scientific (1 mentions) Rhodamine red x goat anti rabbit igg h l, by Thermo Fisher Scientific (1 mentions) Periodic acid solution, by Merck Group (1 mentions) Schiff s solution, by Merck Group (1 mentions) Dp72 camera, by Olympus (1 mentions)

Close spelling variants of this product

Inverted microscope (2417 citations) IX71 microscope (1097 citations) IX71 inverted microscope (813 citations) Inverted research microscope (2 citations)

17 protocols using ix71 inverted research microscope

1

Cell Wound Healing Assay with Taraxerol Acetate

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This assay was performed as previously described (17 (link)). Briefly, U87 cells (1×105 cells/ml) were seeded into a 6-well plate and incubated at 37°C for 24 h until a 95% confluent monolayer of cells was attained. Following 12 h of starvation, a 100-ml pipette tip was used to create a straight cell-free wound. Each well was washed three times with PBS to remove any cell debris, and then subjected to taraxerol acetate (0, 10, 50 and 150 µM) in DMEM. Subsequent to 48 h of incubation at 27°C, the cells were fixed and stained with 5% ethanol containing 0.3% crystal violet powder (Sigma-Aldrich) for 30 min, and images of randomly selected fields were captured under an IX71 inverted research microscope (Olympus Corporation). The number of cells that migrated into the scratched area were counted and the lengths of wound were determined by Image J software, version 1.46 (http://imagej.nih.gov/ij/).
HONG J.F., SONG Y.F., LIU Z., ZHENG Z.C., CHEN H.J, & WANG S.S. (2016). Anticancer activity of taraxerol acetate in human glioblastoma cells and a mouse xenograft model via induction of autophagy and apoptotic cell death, cell cycle arrest and inhibition of cell migration. Molecular Medicine Reports, 13(6), 4541-4548.
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2

Detailed Immunostaining Protocol for Neuronal Markers

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Immunostaining was performed as described previously24 (link) using frozen sections or entire cells fixed in 24-well plates and 3.5-cm film-bottom dishes (Matsunami Glass). The stained cells were viewed with an IX71 inverted research microscope (Olympus) and a DeltaVision ELITE fluorescence microscope (CORNES Technologies). Frozen sections were prepared as previously described24 (link). Hematoxylin and eosin staining of 3-μm-thick specimens was performed as previously described24 (link). Growth cone immunostaining was performed by attaching OVs to 3.5-cm film-bottom dishes on D28, 1 day after administration of chemical agents, and conducting double-staining with phalloidin and GAP43 antibody. The primary antibodies and their respective dilutions used in this study are listed in Table 1, and negative controls for these antibodies are indicated in Fig. S1.

Antibody list. The antibodies are used for immunohistochemistry.

Gene ProductSupplierProduct NumberDilution
1BRN3BSanta Cruzsc-60261:50
2ATOH7MilliporeAB56941:50
3ISLET1Abcamab206701:50
4SNCGGeneTexGTX1104831:50
5TUJ1SigmaT-50761:200
6TAUCell Signaling Technology#4019S1:100
7NFLCell Signaling Technology#2837S1:100
8GAP43AbcamAb758101:100
Yokoi T., Tanaka T., Matsuzaka E., Tamalu F., Watanabe S.I., Nishina S, & Azuma N. (2017). Effects of neuroactive agents on axonal growth and pathfinding of retinal ganglion cells generated from human stem cells. Scientific Reports, 7, 16757.
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3

Natural Transformation in P. aeruginosa Biofilms

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In total, 10 cm lengths of Tygon laboratory tubing (2 mm ID) were inoculated with 1/100 dilution of an overnight culture (OD600 ≈ 6) of P. aeruginosa in CAMHB and allowed to attach for 2 h under static conditions after which continuous flow was commenced at a rate of 80 µl min−1 at room temperature. Influent media was CAMHB containing either no added DNA or DNA added at a final concentration of 1 µg ml−1 for pUCPSK plasmid DNA or 0.1 mg ml−1 for gDNA or eDNA. At harvest, the attached biofilm was removed by sonication and biofilm-associated bacteria collected by centrifugation. Transformants were selected by plating onto LB agar with appropriate antibiotic selection and incubated for 24 h at 37 °C. The transformation frequency was calculated from the number of transformants/total number of harvested c.f.u. To visualize natural transformation by PAO1 continuous-flow biofilms, an IBIDI μ-slide I (with flow kit) was inoculated and cultured as described for the Tygon tubing biofilms. Biofilms were imaged using an Olympus IX71 inverted research microscope with a ×100 10.4 numerical aperture UPlanFLN objective, FViewII monochromatic camera and AnalySIS Research acquisition software (Olympus Australia, Notting Hill, VIC, Australia) fitted with an environmental chamber (Solent Scientific, Segensworth, UK) for fluorescent imaging.
Nolan L.M., Turnbull L., Katrib M., Osvath S.R., Losa D., Lazenby J.J, & Whitchurch C.B. (2020). Pseudomonas aeruginosa is capable of natural transformation in biofilms. Microbiology, 166(10), 995-1003.
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4

Immunostaining of Cell and Tissue Samples

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Immunostaining was performed using frozen sections or whole cells fixed on the dish. Whole cells specimens were fixed with 4% paraformaldehyde (pH 7.0) for 20 min at room temperature. After two rinses with PBS, specimens were incubated with 0.1% Triton X-100 for 15 min at room temperature and then washed three times with PBS for 5 min each. Specimens of frozen sections or whole cells were then incubated with 3% bovine serum albumin (BSA) for 30 min at room temperature followed by primary antibody incubation for 16 h at 4°C. The primary antibodies used in this study and their dilutions are listed in Supplementary Table 2. Secondary antibody reactions were carried out by incubation with the corresponding species-specific Alexa Fluor-488-conjugated antibodies (1:500, Invitrogen) for 1 h at room temperature in the dark. After four washes with PBS for 5 min each, specimens were mounted with ProLong Gold Antifade Reagent with DAPI (Invitrogen) and viewed with an IX71 inverted research microscope (Olympus) or BZ-9000E (KEYENCE).
Tanaka T., Yokoi T., Tamalu F., Watanabe S.I., Nishina S, & Azuma N. (2015). Generation of retinal ganglion cells with functional axons from human induced pluripotent stem cells. Scientific Reports, 5, 8344.
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5

Microfluidic Assay for Leukocyte Adhesion

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The assembly of the multichannel microfluidic devices used in this study and the coating of coverslips with recombinant human P-selectin-Fc, ICAM-1-Fc, and IL-8 has been described previously.3 (link),12 (link),19 Briefly, coverslips were coated with P-selectin-Fc (2 μg/ml), ICAM1-Fc (10 μg/ml), and IL-8 (5 μg/ml) for 2 h and then blocked for 1 h with casein (1%) at room temperature (RT). After coating, coverslips were sealed to polydimethylsiloxane chips by magnetic clamps to create flow chamber channels ~29 μm high and ~300 μm across. By modulating the pressure between the inlet well and the outlet reservoir, a wall shear stress of 2 or 6 dyn/cm2 was applied. HL-60 cells (5 × 106 cells/ml) were perfused in the microfluidic device over a substrate of recombinant human P-selectin-Fc, recombinant human ICAM-1-Fc with or without IL-8. The rolling and arrest of cells were recorded by an IX71 inverted research microscope (Olympus America Inc, Cypress, CA, USA) with a 10× NA 0.3 air objective. The number of rolling and arrested cells were counted in 5 channels per group. The rolling duration and rolling distance were acquired from the images by analyzing 15 cells started rolling to arrest.
Sun H., Fan Z., Gingras A.R., Lopez-Ramirez M.A., Ginsberg M.H, & Ley K. (2019). Frontline Science: A flexible kink in the transmembrane domain impairs β2 integrin extension and cell arrest from rolling. Journal of leukocyte biology, 107(2), 175-183.
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6

Quantifying Axonal Density from RGCs

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The density of the Western blots was quantified using ImageJ software (Ver 1.53, NIH, Bethesda, MA, USA), and a relative score was calculated compared to the control under normal oxygen conditions, obtained from three independent experiments.
The immunostained specimens were imaged using an IX71 inverted research microscope (Olympus, Tokyo, Japan), and the images were processed using ImageJ. To quantify the number of axons from the RGCs, the images were sharpened and converted into 8-bit binary images using ImageJ. The relative axon counts were statistically analyzed for each condition.
Yoshida T., Yokoi T., Tanaka T., Matsuzaka E., Saida Y., Nishina S., Takada S., Shimizu S, & Azuma N. (2024). Modeling of Retina and Optic Nerve Ischemia–Reperfusion Injury through Hypoxia–Reoxygenation in Human Induced Pluripotent Stem Cell-Derived Retinal Ganglion Cells. Cells, 13(2), 130.
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7

Transfection and Time-Lapse Analysis of Axonal Transport

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The NTRK1 expression vector (RG213091; ORIGENE), the pPAmCherry-Mito Vector (TaKaRa Bio), and the pcDNA™6.2/C-EmGFP Vector (Invitrogen) were electroporated into cultured cells. For electroporation, NEPA21 (NEPAGENE) with platinum electrodes was used. The cultured colony was injected with fast green-dyed DNA solution using a sharp glass pipette, placed between the electrodes, and electroporated with voltage pulses (poring pulse: voltage 100 V, width 2.5 ms, interval 50 ms, two times; transfer pulse: voltage 20 V, width 50 ms, interval 50 ms, five times). The cultured cells were then allowed to develop in humidified incubators. Observations were made with an IX71 inverted research microscope (Olympus).
The time series of anterograde axonal flow was conducted by injection of Alexa Fluo-555 conjugated cholera toxin subunit B (Life Technologies) into the RGCR. Time-lapse analysis was performed immediately after the injection of cholera toxin B with a DeltaVision ELITE (CORNES Technologies).
Tanaka T., Yokoi T., Tamalu F., Watanabe S.I., Nishina S, & Azuma N. (2015). Generation of retinal ganglion cells with functional axons from human induced pluripotent stem cells. Scientific Reports, 5, 8344.
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8

Multimodal Retinal Imaging and Registration

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Epiluorescent images of the retina mounted on the MEA were taken with an Olympus IX71 inverted research microscope (10x magnification). Images were taken after visual stimulation to avoid photoreceptor bleaching that would undermine visual responses, and before optogenetic stimulation to minimize tdTomato bleaching by the intense microLED light. After immunostaining, confocal images of the retina were taken on the Leica SP5 Confocal microscope (60X magnification, z-stack spacing 0.34 μm). Images were stitched together in FIJI using the plugins for either pairwise or grid collection stitching35 (link). MEA epifluorescent images and immunostained confocal images were then manually registered by matching the unique pattern of RGC somas’ locations. Matching was done by adjusting position, rotation and scale of one of the images.
Pisano F., Zampaglione E., McAlinden N., Roebber J., Dawson M.D., Mathieson K, & Sher A. (2017). Large scale matching of function to the genetic identity of retinal ganglion cells. Scientific Reports, 7, 15395.
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9

Quantifying Neutrophil Rolling and Arrest

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To study the rolling and arrest of neutrophils, isolated human neutrophils (5 × 106 cells ml−1) were perfused in the microfluidic device over a substrate of recombinant human P-selectin-Fc and recombinant human ICAM-1-Fc with or without IL-8 under a shear stress of 6 dyn cm−2. Neutrophils were incubated with Nexinhib20 (10 μM) or vehicle (DMSO) for one hour at RT before being perfused into the microfluidic devices. Time-lapse images (one frame per second) were taken by an IX71 inverted research microscope (Olympus America) with a 40× NA 0.9 air objective during the perfusion to quantify rolling velocity. The quantification was done using the “Manual tracking” plugin in FIJI-ImageJ v2.0. Cell tracks (Fig. 1A) and rolling velocity were obtained (Fig. 1B,C). After perfusion with neutrophils for 10 minutes, the microfluidic device was washed with RPMI-1640 without phenol red plus 2% HSA for 5 minutes. Then, the arrested neutrophils were counted in nine fields-of-view per group (Fig. 1D).
Liu W., Cronin C.G., Cao Z., Wang C., Ruan J., Pulikkot S., Hall A., Sun H., Groisman A., Chen Y., Vella A.T., Hu L., Liang B.T, & Fan Z. (2022). Nexinhib20 inhibits neutrophil adhesion and β2 integrin activation by antagonizing Rac-1-GTP interaction. Journal of immunology (Baltimore, Md. : 1950), 209(8), 1574-1585.
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10

Time-lapse Analysis of Axonal Growth

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A protocol similar to the one described for the local and sustained agent release assay was used to observe time-lapse analysis of axonal growth. A piece of hydrogel containing SLIT1 (5 μg/ml) diluted in Matrigel was placed in front of attached OVs on D29. Time-lapse observation was performed for 18 h in a STX Stage Top Incubator (TOKAI HIT) and imaged using an IX71 inverted research microscope (Olympus).
Yokoi T., Tanaka T., Matsuzaka E., Tamalu F., Watanabe S.I., Nishina S, & Azuma N. (2017). Effects of neuroactive agents on axonal growth and pathfinding of retinal ganglion cells generated from human stem cells. Scientific Reports, 7, 16757.
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