Lumplfln 40xw

Manufactured by Olympus

Sourced in Japan

The LUMPLFLN 40XW is a long working distance, water immersion objective lens designed for Olympus microscopes. It has a numerical aperture of 0.8 and a working distance of 3.3 mm. The lens is suitable for a variety of microscopy techniques, including fluorescence and live-cell imaging.

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Lab products found in correlation

Fm4 64, by Thermo Fisher Scientific (3 mentions) Bx51wi, by Olympus (2 mentions) Matlab, by MathWorks (2 mentions) Xlfluor 2x 340, by Olympus (1 mentions) Ixonem, by Oxford Instruments (1 mentions) Xlumplfln20xw, by Olympus (1 mentions) Ba510 550, by Olympus (1 mentions) Ba575if, by Olympus (1 mentions) Nipkow confocal scanner csu21, by Yokogawa (1 mentions) Cmos camera, by Hamamatsu Photonics (1 mentions) Igor pro, by Wavemetrics (1 mentions) Resonant galvanometer, by Thorlabs (1 mentions) H10770pa 40, by Hamamatsu Photonics (1 mentions) Ff670 sdi01, by IDEX Corporation (1 mentions) Ff01 720 sp, by IDEX Corporation (1 mentions) Ff562 di03, by IDEX Corporation (1 mentions) Pxie 6341, by National Instruments (1 mentions) Texas red, by Thermo Fisher Scientific (1 mentions) Sd9 stimulator, by Natus (1 mentions) Polystyrene cell culture dish, by Corning (1 mentions)

17 protocols using lumplfln 40xw

In Vivo Nanoimaging of Cardiomyocytes

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The details of the microscopic system for in vivo nanoimaging have been described in our previous studies [5 (link), 6 (link)]. In brief, an upright microscope (BX-51WI, Olympus Co., Tokyo, Japan) combined with a Nipkow confocal scanner (CSU21, Yokogawa Electric Co., Tokyo, Japan) and an electron multiplying CCD (EMCCD) camera (iXonEM+, Andor Technology Ltd, Belfast, Northern Ireland) were used at a 512 × 512 (or 512 × 170) pixel resolution at an exposure time of 28 (or 9.8) ms. A water immersion lens, either 60× (LUMPLFLN 60XW, N/A 1.00, Olympus Co.), 40× (LUMPLFLN 40XW, N/A 0.80, Olympus Co.), or 20× (XLUMPLFLN 20XW, N/A 1.00, Olympus Co.), and also a 2× lens (XLFluor 2X/340, N/A 0.14, Olympus Co.) were used to visualize the LV surface.
AcGFP-expressing myocytes were excited by a 488 nm laser light (HPU50211-PFS, Furukawa Electric Co., Tokyo, Japan), and the resultant fluorescence signals (emission filter: BA510–550, Olympus Co., Tokyo, Japan) were detected. In the experiments with CellMask, the heart was excited at 532 nm (MiniGreen FCIM-100; Snake Creek Lasers, Friendsville, PA, USA), and the resultant fluorescence signals (emission filter: BA575IF, Olympus Co.) were detected. When excited at 532 nm, the wavelength range for the detection of the fluorescence of CellMask was >575 nm.

Kobirumaki-Shimozawa F., Shimozawa T., Oyama K., Kushida Y., Terui T., Ishiwata S, & Fukuda N. (2018). Optimization of Fluorescent Labeling for In Vivo Nanoimaging of Sarcomeres in the Mouse Heart. BioMed Research International, 2018, 4349170.

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Fluorescence Imaging of sDarken Sensors

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Prior to imaging cells were washed with PBS.Imaging was performed at an upright LNscope from Luigs&Neumann using a 40x water immersion objective (LUMPLFLN40xW, Olympus).
Videos were recorded with a CMOS camera (Hamamatsu) at a framerate of 1 Hz for 2 min.
Neurotransmitter and drugs were directly applied to the bath with a gravity driven perfusion system.
Videos were analyzed in ImageJ {Schneider:2012dw. ROIs were selected around the cell membrane. Mean gray (fluorescence) values for each timepoint were measured. Obtained values were corrected by subtracting background fluorescence. Data were normalized to the first frame by dividing all mean values by the according basal fluorescence (F_base)of the cell. To obtain ΔF/F values, we subtracted baseline fluorescence prior (F₀) to ligand application with fluorescence after application of the ligand (F).

△ F / F = \frac{F - F_{0}}{F_{0}}

For titration curves, data were fitted in Igor Pro(WaveMetrics) with a Hill function.
For a better comparison to existing positive going sensors, we report the typical decrease in fluorescence of sDarken as positive values for K_d plots and agonist specificity.

Kubitschke M., Müller M., Wallhorn L., Pulin M., Mittag M., Pollok S., Ziebarth T., Bremshey S., Gerdey J., Claussen K.C., Renken K., Groß J., Gneiße P., Meyer N., Wiegert J.S., Reiner A., Fuhrmann M, & Masseck O.A. (2022). Next generation genetically encoded fluorescent sensors for serotonin. Nature Communications, 13, 7525.

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Live Cell Imaging of NRK Cells

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A total of 3 × 10⁴ NRK cells (TSPAN4-GFP/Mito-DsRed) used in this study (Fig. 2) were seeded on human fibronectin (200X diluted with 1X PBS) pretreated SiO₂ coated mirror and returned to a CO₂ incubator for 24 h. On the next day, the cell-seeded mirror was directly transferred to the custom-designed sample chamber and submerged in DMEM supplemented with 10% FBS and 1% Pen/Strep antibiotics. Then, live cells labeled with GFP and DsRed were recorded at a 2-Hz volume rate with LUMPLFLN40XW (Olympus) at 37 °C.

Xiong B., Zhu T., Xiang Y., Li X., Yu J., Jiang Z., Niu Y., Jiang D., Zhang X., Fang L., Wu J, & Dai Q. (2021). Mirror-enhanced scanning light-field microscopy for long-term high-speed 3D imaging with isotropic resolution. Light, Science & Applications, 10, 227.

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Two-Photon Calcium Imaging Setup

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The 2-photon calcium imaging setup was identical to a previously published design^{15 (link)}. Two-photon illumination was achieved with a Ti:Sapphire laser (Chameleon Vision II, Coherent) operating at 920nm. Fluorescence was acquired using a 40× 0.8 NA objective (LUMPLFLN40X/W, Olympus) and GaAsP PMTs (H10770PA-40, Hamamatsu) after passing through a dichroic (FF670-SDi01, Semrock), an IR filter (FF01–720sp, Semrock), reflected by a second dichroic (FF562-Di03, Semrock) and passing through a final bandpass filter (FF01–520/60, Semrock). The PMT output signal was amplified (Variable High Speed Current Amplifier; #59–179, Edmund Optics) and digitized (PXIe-7961R FlexRIO, National Instrument). The microscope was controlled by ScanImage (Vidrio Technologies) software using additional analog output units (PXIe-6341, National Instruments) for the laser power control and the scanners control. Double-distilled water was used as the immersion medium for the objective. Average beam power measured at the front of the objective was 60–160 mW. The region between the objective and imaging site was shielded from external sources of light using a black rubber tube. Horizontal scans of the laser were achieved using a resonant galvanometer (Thorlabs). Typical fields of view measured approximately 500×500 μm, and data was acquired at 30Hz.

Nieh E.H., Schottdorf M., Freeman N.W., Low R.J., Lewallen S., Koay S.A., Pinto L., Gauthier J.L., Brody C.D, & Tank D.W. (2021). Geometry of abstract learned knowledge in the hippocampus. Nature, 595(7865), 80-84.

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Retrograde Neuronal Labeling via Electroporation

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Texas Red (100 mg ml^–1; dextran, Texas Red, 3,000 MW, lysine fixable) (ThermoFisher Scientific) in patch-clamp intracellular saline (see above) lacking ATP, GTP, biocytin and Alexa-568–hydrazide-Na was backfilled into a patch pipette. The pipette was positioned near the cell body (without any collagenase application) and two to five pulses of 10 V (2 ms duration) were applied using an SD9 stimulator (Grass Instruments). All fills and anatomy were carried out with flies on the wheel under the two-photon microscope (as in calcium imaging, except using a ×40/0.80 NA objective (LUMPLFLN 40XW, Olympus) and a 590–650 nm bandpass filter (Chroma) to filter emitted light before entering a second GaAsP detector (Hamamatsu).

Vijayan V., Wang F., Wang K., Chakravorty A., Adachi A., Akhlaghpour H., Dickson B.J, & Maimon G. (2023). A rise-to-threshold process for a relative-value decision. Nature, 619(7970), 563-571.

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In-vivo Osteocyte Calcium Imaging

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Multiphoton imaging (MPM) was performed at the dorsal mid-shaft region of the MT3 in vivo, sampling osteocytes in a plane located ~ 20μm below the periosteal surface. Osteocyte Ca²⁺ imaging was performed using a 40x magnification water immersion objective (Olympus LUMPLFLN 40XW, NA = 0.8; working distance = 3.3 mm) focused at the mid-diaphysis. Excitation was at 920nm wavelength and a 490–560nm bandpass filter was used for detection. Time series images were acquired at a rate of 6 frames per second. The sampling ROIs at the magnification used were 250μm² located immediately on either side of the mid-diaphysis. Ca²⁺ intensity measurements were performed by post-processing time series images using ImageJ (NIH). Individual osteocytes were delineated and mean pixel intensity values were collected in each frame before and during loading. The intensities for each cell of interest were normalized to the mean baseline intensity for that cell over a 30s period prior to the start of cyclic loading. Responding osteocytes were defined as those cells showing a >25% increase in normalized fluorescence intensity during loading. Figure 2 shows an example of the fluorescence intensity changes typically seen in cells that respond to mechanical loading. In addition to counting the number of cells, we also analyzed the fold increase in mean intensity during loading compared to non-loaded baseline.

Lewis K.J., Cabahug Zuckerman P., Boorman-Padgett J.F., Basta-Pljakic J., Louie J., Stephen S., Spray D.C., Thi M.M., Seref-Ferlengez Z., Majeska R.J., Weinbaum S, & Schaffler M.B. (2021). Estrogen Depletion on In vivo Osteocyte Calcium Signaling Responses to Mechanical Loading. Bone, 152, 116072.

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Cerebral Wall Imaging Protocol

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Cross-sectional or en face cultures of cerebral walls were prepared as described previously [6 (link), 18 (link), 52 (link)]. Briefly, cerebral walls were microsurgically processed and embedded in a polystyrene cell-culture dish (Corning) with Atelocell IAC-30 collagen gel (Koken) at a concentration of 0.3 mg/mL. Confocal time-lapse images were obtained on a BX51W1 microscope (Olympus) equipped with a CSU-X1 laser scanning confocal unit (Yokogawa) with a 40× objective lens (LUMPLFLN40XW, Olympus) or 100× objective lens (LUMPLFL100XW, Olympus) and an iXon+ EMCCD camera (Andor), in an on-stage culture chamber (Tokai Hit) filled with 45% N₂, 40% O₂, and 5% CO₂. For confocal imaging of all VZ cells in the subapical region (Fig 2B), processed cerebral hemispheric walls were stained with FM4-64 (Thermo Fisher Scientific) at a concentration of 5 μg/mL, as described previously [27 (link), 52 (link)]. The areal occupancy of either somata (Fig 2B, defined such that the diameter of each sectional area was >3 μm) or processes was examined as described previously [45 (link)].

Shinoda T., Nagasaka A., Inoue Y., Higuchi R., Minami Y., Kato K., Suzuki M., Kondo T., Kawaue T., Saito K., Ueno N., Fukazawa Y., Nagayama M., Miura T., Adachi T, & Miyata T. (2018). Elasticity-based boosting of neuroepithelial nucleokinesis via indirect energy transfer from mother to daughter. PLoS Biology, 16(4), e2004426.

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Multicolor Fluorescence Microscopy Setup

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The optical setup consists of a commercial Olympus upright microscope (BX43), a laser arm for fluorescence illumination, and an optical collection arm. The objective lens used in the experiment included a 20×/0.5 NA (Olympus, Cat. #UMPLFLN20XW) water-immersion objective, a 40×/0.8NA (Olympus, Cat. #LUMPLFLN40XW) water-immersion objective, and a 60×/1.1 NA (Olympus, Cat. #UMPLFLN20XW) water-immersion objective. The light source used for fluorescence excitation was a switchable continuous multimode fiber laser (λ = 488 nm and λ = 561 nm, Oxxius, Cat. #L4Cc; λ = 640 nm, 89North, Cat. #LDI-7). In the following optical path, the system included a relay lens pair with an optical magnification of 1.15×; thus, the final equivalent magnification was 23×, 46×, and 69× for the objectives. A two-dimensional piezo (Coremorrow, Cat. #P33.T2S) was inserted at the pupil plane of the 4 F system, and the microlens array (pitch size of 97.5 µm with a focal length of 1.95 mm) was placed at the image plane of the 4 F system. After that, with a second 1:1 optical relay lens pair, an sCMOS camera (PCO Panda4.2, 2048 × 2048 pixels) was placed at the conjugated focal plane of the MLA (Fig. S6). The excitation light path was coupled to an upright microscope with a four-color filter set (405/488/561/640 nm, Chroma, 89901v2).

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Imaging of Fibronectin-Adhered L929 Cells

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A total of 3 × 10⁴ L929 cells used in this study (Fig. 1 and Fig. S1) were seeded on human fibronectin (200X diluted with 1X PBS) pretreated SiO₂ coated mirrors and returned to a CO₂ incubator for 24 h. Then, the cells were fixed with 4% paraformaldehyde, washed with PBS three times, stained with WGA-647 (Thermal Fisher, 500X diluted with 0.05% Tween 20 in 1X PBS) for 2 h at room temperature, and protected from light. Afterward, the cells were thoroughly washed with 1X PBS three times and then transferred to a custom-designed sample chamber filled with 1X PBS, and imaged with LUMPLFLN40XW (Olympus) at room temperature.

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Real-Time Confocal Imaging of Radiation-Induced Cellular Damage

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Images were obtained using a confocal laser scanning microscope (LSM; FV-1000, Olympus Co., Tokyo) equipped with three lasers at 473, 559 and 635 nm, and captured using a × 40 objective lens (LUMPLFLN 40XW, Olympus Co., Tokyo) for a scan area of 317.2 × 317.2 μm² (2,048 × 2,048 pixels) with a scan speed of 20 µm/s and at 45% laser light transmissivity. A CO₂ incubation chamber (INUG2F-UK, Tokai-Hit, Shizuoka) with a gas mixer (GM-5000, Tokai-Hit, Shizuoka) was installed on a motorized X-Y stage (MPT-AS04FV, SIGMAKOKI, Tokyo) of the LSM. For the FNTD read-out [10 ], a dichroic mirror split at 670 nm was used and scanned at 10 μm below the surface. For cell imaging with Alexaflour^®488 or 53BP1-GFP, a 505-nm dichroic mirror was used. The image resolution and size were set to be the same as the cell images. For all the image acquisitions, the FNTDs were first scanned with a 635-nm laser to image ion traversals and then scanned with a blue laser for cell imaging to avoid ‘recording’ of fluorescence in the FNTD crystal with the focused blue laser light. The image acquisition started 20 min after irradiation, and live images were captured for up to 6 h.

Kodaira S., Konishi T., Kobayashi A., Maeda T., Ahmad T.A., Yang G., Akselrod M.S., Furusawa Y, & Uchihori Y. (2014). Co-visualization of DNA damage and ion traversals in live mammalian cells using a fluorescent nuclear track detector. Journal of Radiation Research, 56(2), 360-365.

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