Water immersion objective

Manufactured by Zeiss

Sourced in Germany, Austria

The 20× water immersion objective is a high-magnification lens designed for use with microscopes. It provides a magnification of 20x, making it suitable for detailed observation and analysis of specimens immersed in water or other aqueous media. The objective is optimized for use with water-based samples, ensuring accurate and high-quality imaging.

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Lab products found in correlation

Axio observer z1, by Zeiss (5 mentions) Labview, by National Instruments (3 mentions) Axiovert 35, by Zeiss (2 mentions) Obis 532 ls laser, by Coherent Inc (2 mentions) Lsm 880 nlo confocal microscope, by Zeiss (2 mentions) Stereo discovery 20 microscope, by Zeiss (2 mentions) Alizarin red, by Merck Group (2 mentions) Lsm 510, by Zeiss (2 mentions) Ultraview vox, by PerkinElmer (2 mentions) Lsm 780, by Zeiss (2 mentions) Axioimager m2 microscope, by Visitron (2 mentions) Visiview, by Visitron (2 mentions) Ixon dv887esc bv, by Oxford Instruments (1 mentions) Axio observer 7, by Zeiss (1 mentions) Ti sapphire femtosecond laser, by Spectra-Physics (1 mentions) Lsm 7 mp, by Zeiss (1 mentions) 492 sp25 nm filter, by IDEX Corporation (1 mentions) Nm filter, by IDEX Corporation (1 mentions) Calcein blue, by Merck Group (1 mentions) Tricaine, by Merck Group (1 mentions)

25 protocols using water immersion objective

Single-Molecule Fluorescence Microscopy

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Experiments were carried out on a fluorescence microscope (AxioObserver Z1, Carl Zeiss, Thornwood, NY, United States), equipped with a 63x water immersion objective (Zeiss; N.A. = 0.95) and prism-based TIRF illumination. The light source was an OBIS 640LX laser from Coherent Inc., (Santa Clara, CA, United States). Fluorescence was observed through a 665 nm long pass filter (LP665; Semrock, Rochester, NY, United States) by an electron multiplying CCD (DV887ESC-BV; Andor Technologies, Belfast, United Kingdom). The EMCCD was cooled to −70^°C, and the gain was typically set to an electron gain factor of 200. The prism-quartz interface was lubricated with glycerol to allow easy translocation of the sample cell on the microscope stage. The beam was totally internally reflected at an angle of 72 from the surface normal, resulting in an evanescent wave that decays exponentially with a characteristic penetration depth of ∼100 nm. Which means that vesicle fluorescence is not visible until they approach the membrane. An elliptical area of 250 x 65 μm was illuminated. The laser intensity, shutter, and camera were controlled by a homemade program written in LabVIEW (National Instruments, Austin, TX, United States).

Coffman R.E., Kraichely K.N., Kreutzberger A.J., Kiessling V., Tamm L.K, & Woodbury D.J. (2022). Drunken lipid membranes, not drunken SNARE proteins, promote fusion in a model of neurotransmitter release. Frontiers in Molecular Neuroscience, 15, 1022756.

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Supported Bilayer Fusion Assay

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Dense core granule to planar supported bilayer fusion assay experiments were performed on a Zeiss Axio Observer 7 fluorescence microscope (Carl Zeiss), with a 63x water immersion objective (Zeiss, N.A. 0.95) and a prism-based TIRF illumination. Laser light at 514 nm from an argon ion laser (Innova 90C, Coherent), controlled through an acousto-optic modulator (Isomet), and at 640 nm from a diode laser (Cube 640, Coherent) were used as excitation sources. The characteristic penetration depths were between 90 and 130 nm. An OptoSplit (Andor Technology) was used to separate two spectral bands (540 nm – 610 nm, and 655 nm – 725 nm). Fluorescence signals were recorded by an EMCCD (iXon DV887ESC-BV, Andor Technology).

Bendahmane M., Chapman-Morales A., Kreutzberger A.J., Schenk N.A., Mohan R., Bakshi S., Philippe J., Zhang S., Kiessling V., Tamm L.K., Giovannucci D.R., Jenkins P.M, & Anantharam A. (2020). Synaptotagmin-7 enhances calcium-sensing of chromaffin cell granules and slows discharge of granule cargos. Journal of neurochemistry, 154(6), 598-617.

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Single Vesicle Docking and Fusion Imaging

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Experiments examining single-vesicle docking and fusion events were performed on a Zeiss Axiovert 35 fluorescence microscope (Carl Zeiss, Thornwood, NY), equipped with a 63x water immersion objective (Zeiss; N.A. = 0.95) and a prism-based TIRF illumination. The light source was an OBIS 532 LS laser from Coherent Inc. (Santa Clara CA). Fluorescence was observed through a 610 nm band pass filter (D610/60; Chroma, Battleboro, VT) by an electron multiplying CCD (DU-860E; Andor Technologies). The prism-quartz interface was lubricated with glycerol to allow easy translocation of the sample cell on the microscope stage. The beam was totally internally reflected at an angle of 72 o from the surface normal, resulting in an evanescent wave that decays exponentially with a characteristic penetration depth of ~100 nm. An elliptical area of 250 x 65 µm was illuminated. The laser intensity, shutter, and camera were controlled by a homemade program written in LabVIEW (National Instruments, Austin, TX).

Nyenhuis S.B., Karandikar N., Kiessling V., Kreutzberger A.J., Thapa A., Liang B., Tamm L.K., & Cafiso D.S. (2020). Conserved Arginine Residues in Synaptotagmin 1 Regulate Fusion Pore Expansion Through Membrane Contact.

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Two-Photon Imaging of Zebrafish Facial Lobe

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Images were acquired using a two-photon microscope and 20× water immersion objective (Zeiss). A femtosecond Ti:Sapphire laser (Spectra Physics) was used as an excitation source at 920 nm. Images were acquired using Zeiss software at 7.5 Hz from a single optical plane at a time. We collected data from up to three different optical planes (with 10 μm separation) of the facial lobe in each juvenile zebrafish, covering a large portion of the facial lobe. For ensuring the reliability and robustness of our conclusions we used large sample size of juvenile zebrafish in all our imaging experiments. Here we used n = 10 fish for taste categories, n = 14 fish and n = 11 fish for taste concentrations, n = 7 fish for taste mixtures and n = 10 fish for taste mixture morphing experiments.

Vendrell-Llopis N, & Yaksi E. (2015). Evolutionary conserved brainstem circuits encode category, concentration and mixtures of taste. Scientific Reports, 5, 17825.

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Two-Photon and Confocal Microscopy of AGuIX-RhoB

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AGuIX^®-RhoB (10 mM) were added to the apical phase of the cells during the indicated time. Two-photon microscopy was performed as described in Sancey et al. [39 (link)], using a LSM 7 MP (Zeiss, Germany) equipped with a 20 × water-immersion objective (NA 1.0; Zeiss) and ZEN 2010 software for detection of the NPs. Laser excitation was done at 800 nm with a Ti:Sapphire laser (Chameleon vision II; Coherent, UK). Fluorescence emissions were detected simultaneously by three non-descanned photomultiplier tubes with a 492/SP25 nm filter (Semrock, US) for blue autofluorescence and Hoechst emission, a 542/50 nm filter (Semrock, US) for green autofluorescence emission, and a 617/73 nm filter (Semrock, US) for AGuIX^®-RhoB fluorescence emission. Autofluorescence and second harmonic generation of biological structures could also be collected in the 3 channels due to the presence of collagen, lectin and elastin as example. Confocal microscopy was performed using an LSM 510 (Zeiss Germany) equipped with a 40 × oil-immersion objective (NA 1.2; Zeiss). Laser excitations/emissions were 760 nm biphotonic/400–450 nm for Hoechst, 488 nm/500–550 nm for FITC/GFP, 543 nm/550–600 nm for Rhodamine-B, 633 nm/650–705 nm for Cy-5, respectively.

Sancey L., Sabido O., He Z., Rossetti F., Guignandon A., Bin V., Coll J.L., Cottier M., Lux F., Tillement O., Constant S., Mas C, & Boudard D. (2020). Multiparametric investigation of non functionalized-AGuIX nanoparticles in 3D human airway epithelium models demonstrates preferential targeting of tumor cells. Journal of Nanobiotechnology, 18, 129.

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Zebrafish Skeletal Staining and Histology

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Zebrafish in vivo skeletal staining was incubated with 0.05% Alizarin red (Sigma, A5533) for 1 hour and then washed with system water three times. Alizarin red and Alcian blue whole skeleton staining was performed as described 12 (link). For histological analysis, samples were fixed in 4% paraformaldehyde, decalcified in 15% EDTA and embedded in paraffin as described 32 (link). Sections (5-μm thick) were stained with Safranin O/Fast Green and hematoxylin and eosin (H & E). Picric-sirius red staining was used to detect collagen fiber density and organization as described 33 (link). Zebrafish embryos were imaged with a SteREO Discovery 20 microscope (Carl Zeiss) or LSM880NLO confocal microscope with a 20× water immersion objective (Carl Zeiss). The ceratohyal length measurements, the relative area of osterix labeled osteoblasts measurements and the mineralization intensity measurement were conducted using Image J.

Sun X., Zhang R., Chen H., Du X., Chen S., Huang J., Liu M., Xu M., Luo F., Jin M., Su N., Qi H., Yang J., Tan Q., Zhang D., Ni Z., Liang S., Zhang B., Chen D., Zhang X., Luo L., Chen L, & Xie Y. (2020). Fgfr3 mutation disrupts chondrogenesis and bone ossification in zebrafish model mimicking CATSHL syndrome partially via enhanced Wnt/β-catenin signaling. Theranostics, 10(16), 7111-7130.

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Multicolor Zebrafish Skeletal and Organelle Staining

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Zebrafish in vivo skeletal staining was incubated with 0.2% Calcein (Sigma, C0875) solution (pH 7.0) for 10 min or 0.2% Calcein blue (Sigma, M1255) solution (pH 7.0) for 1 h or 0.05% Alizarin red (Sigma, A5533) for 1 h and then washed with system water three times. Cell internal membranes labeled with vital dye BODIPY TR Methyl Ester (C34556, Invitrogen) as described by the manufacturer, the embryos were stained with 100 μM MED for 1 h, then wash three times with egg water. BODIPY TR Ceramide (D7540, Invitrogen) prominent labeling of the Golgi apparatus with 50 μM for 2 h. LysoTracker Green DND-26 (L7526, Invitrogen) was used to in vivo label acidic notochord vacuoles after 5–6 dpf, 50 μM stained for 1–2 h depending on acidification degree of notochord vacuoles. Zebrafish embryos were anesthetized in tricaine (Sigma) and mounted in 1% LMP agarose. Calcein staining embryos were imaged with a SteREO Discovery 20 microscope (Carl Zeiss). Other live staining embryos were imaged with a LSM880NLO confocal microscope with a ×20 water immersion objective (Carl Zeiss). Images collected every 10 min for time series live cell imaging of notochord development from 20 to 30 hpf.

Sun X., Zhou Y., Zhang R., Wang Z., Xu M., Zhang D., Huang J., Luo F., Li F., Ni Z., Zhou S., Chen H., Chen S., Chen L., Du X., Chen B., Huang H., Liu P., Yin L., Qiu J., Chen D., Deng C., Xie Y., Luo L, & Chen L. (2020). Dstyk mutation leads to congenital scoliosis-like vertebral malformations in zebrafish via dysregulated mTORC1/TFEB pathway. Nature Communications, 11, 479.

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Two-Photon Imaging of Neural Activity

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Neural activity measurements were collected with two-photon laser scanning systems. For the experiment described in Fig 3, data were collected for one to three planes per fish using successive recordings of a single plane at 3.4 Hz (LSM 7 MP upright with 20× water-immersion objective, Zeiss). For the remaining experiments, volumetric images were recorded at 3.3 Hz (upright with 16× water-immersion objective, Scientifica) for six planes simultaneously, which were evenly spanning the ventral side of the OB within a range of 10–110 μm deep. In both cases, a mode-locked Ti:Sapphire laser tuned to 920 nm (for GCaMP6s recordings) or 840 nm (for Rhod-2 AM recordings) was used to excite the fluorophores.

Kermen F., Lal P., Faturos N.G, & Yaksi E. (2020). Interhemispheric connections between olfactory bulbs improve odor detection. PLoS Biology, 18(4), e3000701.

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Intravital Microscopy of Intestinal Microcirculation

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As previously described, rhodamine 6G (100 μL, 20 mg/mL, Sigma-Aldrich) was used for fluorescence in vivo leukocyte labelling 5 min prior to IVM. Next, mice underwent relaparotomy and a proximal intestinal segment was fixed for in vivo analysis of interactions between platelets, leukocytes and vascular endothelial cells in postischemic intestinal postcapillary venules (pcvs). Animal’s body temperature was maintained at 37 °C with a heating pad and exposed intestine was kept wet by periodic administration of warm saline. A multicolor intravital fluorescence microscope (BX51WI, Olympus Life Science Europa GmbH, Hamburg, Germany) equipped with a 20× water-immersion objective (Zeiss, Oberkochen, Germany) and a wavelength switching illumination system (Lambda DG-4, Novato, Canada) was applied. Digital visualization and offline recording of individual vessels and fluorescently labelled cells were performed using an emission splitting system in combination with a microscope camera (Cascade II 512, Photometrics, Tucson, AZ, USA). In all animals, pcvs (15–30 µm of diameters) in three randomly selected and non-overlapping microscopic fields were scanned and recorded for 90 s with one picture every 50 ms.

Chen Z., Mohr A., Heitplatz B., Hansen U., Pascher A., Brockmann J.G, & Becker F. (2020). Sulforaphane Elicits Protective Effects in Intestinal Ischemia Reperfusion Injury. International Journal of Molecular Sciences, 21(15), 5189.

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Intravital Imaging of synNotch-CAR T Cells

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Intravital two-photon images were acquired with Zeiss LSM 780 NLO equipped with a Ti:Sapphire laser (MaiTai HP, Spectra Physics) tuned to 760 nm (for excitation of tagBFP⁺ synNotch-CAR T cells and mCherry⁺ tumor) and 900 nm (for excitation of GFP⁺ synNotch-CAR T cells), respectively, and focused through a Zeiss 20× water immersion objective (numerical aperture of 1.0). Before imaging, mice were anesthetized with isoflurane and the headplate was fixed into the head posts of a custom-made moving stage (Thorlabs, UC Berkeley Physics Machine Shop). Anesthesia was maintained at 1% isoflurane through a nose cone, and body temperature was kept stable via a temperature-controlled heating pad. Images of 598 µm × 598 µm areas of in vivo tumors were acquired at 512 pixel × 512 pixel resolution for standard images and 1024 pixel × 1024 pixel resolution for higher-resolution images. Volume images were acquired over a 30- to 200-µm Z range in 5- or 10-µm steps. Time-lapse datasets were acquired either in single planes over time periods up to 45 min or in combined time + Z series over a 598 × 598 area (X × Y) with variable Z ranges (Z = 5 to 100 µm) with 1- to 5-µm steps. Movies were processed in Zen software for three-dimensional reconstructions.

Choe J.H., Watchmaker P.B., Simic M.S., Gilbert R.D., Li A.W., Krasnow N.A., Downey K.M., Yu W., Carrera D.A., Celli A., Cho J., Briones J.D., Duecker J.M., Goretsky Y.E., Dannenfelser R., Cardarelli L., Troyanskaya O., Sidhu S.S., Roybal K.T., Okada H, & Lim W.A. (2021). SynNotch-CAR T cells overcome challenges of specificity, heterogeneity, and persistence in treating glioblastoma. Science translational medicine, 13(591), eabe7378.

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