Lsm 510 laser scanning microscope

Manufactured by Zeiss

Sourced in Germany, United States, Japan

The LSM 510 is a laser scanning microscope manufactured by Zeiss. It is designed to provide high-resolution images of microscopic samples by scanning them with a focused laser beam and detecting the resulting fluorescence or reflectance. The microscope features advanced optics and detectors to capture detailed information about the structure and composition of the sample.

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Lab products found in correlation

Axio observer z1, by Zeiss (6 mentions) Prolong gold antifade mountant, by Thermo Fisher Scientific (5 mentions) Vectashield, by Vector Laboratories (4 mentions) Triton x 100, by Merck Group (3 mentions) Alexa fluor 488, by Thermo Fisher Scientific (3 mentions) Hoechst 33432, by Nacalai Tesque (2 mentions) Orca charge coupled device, by Hamamatsu Photonics (2 mentions) Axiovert 200m microscope, by Zeiss (2 mentions) Eight well chamber slide, by Thermo Fisher Scientific (2 mentions) Hoechst 33342, by Thermo Fisher Scientific (2 mentions) Appropriate secondary antibody, by Thermo Fisher Scientific (2 mentions) Axioplan 2 microscope, by Zeiss (2 mentions) Biolistic pds 1000 he particle delivery system, by Bio-Rad (2 mentions) Cy5 dutp, by Enzo Life Sciences (2 mentions) Plan neofluar, by Zeiss (2 mentions) Mowiol 4 88, by Merck Group (1 mentions) Axio zoom v16 fluorescence stereomicroscope, by Zeiss (1 mentions) Axiovert 200m fluorescence microscope, by Zeiss (1 mentions) Alexa fluor 555 conjugate, by Thermo Fisher Scientific (1 mentions) W32464, by Thermo Fisher Scientific (1 mentions)

85 protocols using lsm 510 laser scanning microscope

Imaging Vascular Development in Zebrafish

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Live and whole-mount hybridized embryos were photographed on agarose-coated dishes using either an epifluorescence Leica MZ16 F stereomicroscope (1X Plan Apo objective, NA0.141) (Leica, Wetzlar, Germany) equipped with digital camera or an Axio Zoom.V16 fluorescence stereomicroscope (Zeiss). Phalloidin-stained embryos were acquired on an Axiovert 200M fluorescence microscope (Zeiss) equipped with ApoTome.2 to enhance resolution. Evaluation of ISV defects was carried out on developing vessels in the region of the trunk above the prolongation of the yolk, as indicated in Figure 4A.
Confocal analysis of filopodia was performed using a LSM510 laser scanning microscope (Zeiss). To this purpose, 26–28 hpf embryos were fixed overnight with a PBS-based solution containing 1% PFA, 0.1% glutaraldehyde and 3% sucrose and mounted on glass slides with Mowiol 4.88 (Sigma). For consistency reasons and in order to minimize stage-related discrepancies, filopodia evaluation was carried out on the ISV pair in which the first vessel had already reached the roof of the trunk and the adjacent vessel was still growing up.

Tobia C., Chiodelli P., Barbieri A., Buraschi S., Ferrari E., Mitola S., Borsani G., Guerra J, & Presta M. (2019). Atypical Chemokine Receptor 3 Generates Guidance Cues for CXCL12-Mediated Endothelial Cell Migration. Frontiers in Immunology, 10, 1092.

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Lectin-Based Visualization of hESC and iPSC

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hESCs and iPSCs cultured on matrigel-coated coverslips for 4–5 days were incubated with wheat germ agglutinin (Alexa Fluor 555 Conjugate) (W32464; Invitrogen) and ECL (Fluorescein labeled Erythrina Cristagalli Lectin (ECL, ECA)) (FL-1141; Vector) at 1:200, for 10 minutes at room temperature. The cells were then washed three times with PBS and fixed for 20 minutes with 4% paraformaldehyde. They were washed three times with PBS and counterstained with DAPI. Imaging was performed on a Zeiss LSM510 laser scanning microscope.

Rapti K., Stillitano F., Karakikes I., Nonnenmacher M., Weber T., Hulot J.S, & Hajjar R.J. (2015). Effectiveness of gene delivery systems for pluripotent and differentiated cells. Molecular Therapy. Methods & Clinical Development, 2, 14067-.

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Microfluidic Bead and Cell Rolling

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The beads experiments were performed with a Nikon Eclipse TE 300 inverted microscope, at a 20× objective and with a MQ003MG-CM B&W camera (Ximea). The camera was controlled with the XimeaCop software acquiring at 200 frames per second (FPS). The cell experiments were performed on a LSM510 laser scanning microscope (Zeiss) equipped with 20× dry objective. The cell movies were acquired at a frame rate of 1.27 FPS. Fluigent Flow-EZ pressure controller ranging from 0 mbar to 1000 mbar was used to apply pressure on the glass vials. 100 μm inner diameter polytetrafluoroethylene (PTFE, IDEX 1571) tubing was used to connect the glass vials to the microfluidic chip through a custom-made vial cap. The bead and cell movies were analysed using FIJJ software and manual particle tracking plugins.
The chip was primed with deionized water at high pressure (200 mbar) to remove bubbles. A first set of particles (beads per cells) were loaded in the chip at 70 mbar to be trapped. Once all trapping sites were filled the applied pressure was decreased to 20 mbar for the rolling behaviour of beads and 2 mbar for cell rolling. After each use the PDMS slab was removed from the chip and channels were cleaned with bleach (14%) and sulfuric acid (H₂SO₄ 96%, TECHNIC). The cleaned chips were then bonded with PDMS and used again for further experiments.

Duchamp M., Dahoun T., Vaillier C., Arnaud M., Bobisse S., Coukos G., Harari A, & Renaud P. (2019). Microfluidic device performing on flow study of serial cell–cell interactions of two cell populations. RSC Advances, 9(70), 41066-41073.

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Immunocytochemical Analysis of NF-κB Activation

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Immunocytochemistry for confocal microscopy was performed on NFκB. NHEKs were seeded in each well of the slide chamber at a density of 1×10⁴ cells per well. The cells were treated with HDM (10 mg/ml) or C48/80 (10 µg/ml) for 6 hours. After being washed with phosphate buffered saline (PBS), cells were fixed in 4% paraformaldehyde in PBS for 10 minutes, and then permeabilized with PBS containing 0.5% Triton X-100 (PBS-T) for 15 minutes. The cells were incubated with PBS-T containing 1% bull serum albumin for 1 hour and stained with the anti-NFκB (1:150; Abcam, Cambridge, MA, USA) overnight at 4℃, followed by incubation with Alexa Fluor 488 goat anti-rabbit IgG (1:500; Invitrogen) for 1 hour. Images were visualized using confocal microscopy with a 20X objective on an LSM 510 laser scanning microscope (Carl Zeiss) and analyzed using the LSM 5 browser imaging software.

Choi D.I., Park J.H., Choi J.Y., Piao M., Suh M.S., Lee J.B., Yun S.J, & Lee S.C. (2020). Keratinocytes-Derived Reactive Oxygen Species Play an Active Role to Induce Type 2 Inflammation of the Skin: A Pathogenic Role of Reactive Oxygen Species at the Early Phase of Atopic Dermatitis. Annals of Dermatology, 33(1), 26-36.

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Visualizing Mitochondrial Protein SLC25A11

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Cells were fixed with 4% (w/v) paraformaldehyde and permeabilized with 0.5% Triton X-100. The cells were then stained with anti-SLC25A11 polyclonal antibody, Mitotracker (M22425, Invitrogen Life Technologies, Carlsbad, CA, USA) and Alexa Fluor 488-conjugated anti-rabbit antibody (A11008, Invitrogen Life Technologies, Carlsbad, CA, USA). Live cell imaging was taken by LSM510 Laser Scanning Microscope and Axio Observer Z1 (Carl Zeiss, Oberkochen, Germany). The relative intensity was normalized by the arithmetic mean intensity (from Zen software 2.6 blue edition).

Lee J.S., Lee H., Lee S., Kang J.H., Lee S.H., Kim S.G., Cho E.S., Kim N.H., Yook J.I, & Kim S.Y. (2019). Loss of SLC25A11 causes suppression of NSCLC and melanoma tumor formation. EBioMedicine, 40, 184-197.

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Immunofluorescence Assay for TWIST1 in MCF-7 Cells

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For immunofluorescence assays, MCF-7 cells were seeded onto glass coverslips in 6-well plates and grown for 24 h. Then, media were replaced by fresh ones with 10% FBS and containing either doxorubicin alone (1 μM) and/or melatonin (1 nM), or vehicle (ethanol at a final concentration lower than 0.0001%). After 12 h, cells were washed with PBS, fixed in 4% formaldehyde solution for 30 min and then permeabilized with 0.2% Triton X-100/PBS for 15 min (twice). Coverslips were incubated with anti-TWIST1 primary antibodies overnight at 4 °C, followed by 1 h incubation with fluorescence-tagged secondary antibody at room temperature, and then stained with DAPI for 5 min. Finally, coverslips were observed, and cell images were captured at 40× magnification with an LSM-510 laser scanning microscope (Carl Zeiss Inc., Sliedrecht, The Netherlands) fluorescence microscope.

Menéndez-Menéndez J., Hermida-Prado F., Granda-Díaz R., González A., García-Pedrero J.M., Del-Río-Ibisate N., González-González A., Cos S., Alonso-González C, & Martínez-Campa C. (2019). Deciphering the Molecular Basis of Melatonin Protective Effects on Breast Cells Treated with Doxorubicin: TWIST1 a Transcription Factor Involved in EMT and Metastasis, a Novel Target of Melatonin. Cancers, 11(7), 1011.

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Analysis of Mitochondrial Membrane Potential

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Mitochondrial membrane potential was analyzed by measuring tetramethylrodamine ester (TMRE) (87,917, Sigma, St. Louis, MO, USA) followed by an established method [19 ]. Cells were plated 4 well chambered coverglass (155382, Thermo Fisher Scientific) in 0.5 ml culture media. After 24 h, cells were transfected with NT siRNA or SLC25A11 siRNA (40 nM) for 48 h at 37 C^o. 100 nM of TMRE and 5 μg/ml of Hoechst 33342 were added to the culture medium for 15 min at 37 C^o. Negative control was treated with 50 μM trifluoromethoxy carbonylcyanide phenylhydrazone (FCCP, C2920, Sigma, St. Louis, MO, USA) in serum free media for 15 min at 37 C^o before TMRE stanning. 4 well chambered cover glass was placed on LSM510 Laser Scanning Microscope in the presence of the TMRE and Hoechst 33342. Live cell imaging was taken by LSM510 Laser Scanning Microscope and Axio Observer Z1 (Carl Zeiss, Oberkochen, Germany). The relative intensity of TMRE was normalized by the arithmetic mean intensity (from Zen software 2.6 blue edition).

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Immunofluorescence Staining of Cardiomyocytes

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Immunofluorescence staining was carried out with antimyosin heavy chain antibody (Leica Biosystems, Wetzlar, Germany) and Alexa555-conjuagted antimouse IgG antibody (Invitrogen) as previously described [65 (link)]. A total of 50 myocardial cells were randomly selected, and the surface area of these cells was measured with ImageJ v4.16 software. Detection of nuclear acetylation was done by immunofluorescence. Staining for acetylated histone was performed using the immunofluorescence method previously described [66 (link)], with some modifications. In brief, cardiomyocytes were fixed with 3.7% formaldehyde for 15 min, blocked in 1% BSA and 0.5% NP-40/TBS-Ca for 1 h, and then incubated overnight with anti-acetyl-histone-H3 (K9) antibody or anti-acetyl-histone-H3 (K122) antibody with anti-myosin heavy chain antibody. Further, the cells were incubated with antirabbit Cy3 antibody and antimouse DyLight-649 antibody for 2 h and then stained with Hoechst 33432 (Nacalai Tesque). Immunofluorescence was observed using a LSM 510 Laser Scanning Microscope (Carl Zeiss, Oberkochen, Germany).

Funamoto M., Sunagawa Y., Katanasaka Y., Shimizu K., Miyazaki Y., Sari N., Shimizu S., Mori K., Wada H., Hasegawa K, & Morimoto T. (2021). Histone Acetylation Domains Are Differentially Induced during Development of Heart Failure in Dahl Salt-Sensitive Rats. International Journal of Molecular Sciences, 22(4), 1771.

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Immunofluorescence Staining Protocol

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The blocking step was performed in blocking buffer (donkey serum 1:100 in 0.1% PBS-Tween) at room temperature for 1 hour, followed by incubation with the primary antibody at 4°C overnight. After washing, the samples were incubated with secondary antibody (1:1,000 in blocking buffer) at room temperature for 1 hour. Samples were washed thrice with 0.1% PBS-Tween for 30 minutes, stained with 4′,6-diamidino-2-phenylindole (DAPI) for 10 minutes, and were washed with PBS for 30 minutes. Samples were mounted on glass slides with Vectashield (Vector laboratories, Newark, CA, USA) solution and analyzed with an LSM 510 laser scanning microscope (Carl Zeiss Micro Imaging GmbH, Göttingen, Germany). The primary antibodies and titers used in this work were as follows: HAS1 (either goat, 1:200, Santa Cruz Biotechnology #SC-23145, Dallas, TX, USA; or rabbit, 1:200, Abcam #ab198846, Cambridge, UK), HAS2 (rabbit, 1:200, Santa Cruz Biotechnology #SC-66916), HAS3 (goat, 1:200, Santa Cruz Biotechnology #SC-34204), ICAM2 (rat, 1:200, BioLegend #105602, San Diego, CA, USA), Sox2 (rat, 1:200, eBioscience #14-9811-82, San Diego, CA, USA). All secondary antibodies were applied according to the origin of the primary antibody (Jackson ImmunoResearch, Jackson Grove, PA, USA). DAPI (Invitrogen) was used at 1:10,000 titer.

Umugire A., Lee S., Lee C.J., Choi Y., Kim T, & Cho H.H. (2022). Hyaluronan Synthase 1: A Novel Candidate Gene Associated With Late-Onset Non-syndromic Hereditary Hearing Loss. Clinical and Experimental Otorhinolaryngology, 15(3), 220-229.

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Immunofluorescence Confocal Microscopy Protocol

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For immunofluorescence confocal microscopy, cells were plated onto coverslips in 12 well or 24 well plates. Cells were transfected with plasmids as indicated in figures. Cells were incubated in full media or EBSS for 2 h and fixed in 4 % paraformaldehyde for 10 min followed by permeabilization with 0.1 % saponin in 3 % BSA. Cells were then blocked in 3 % BSA and then stained with primary antibodies followed by washings with PBS and then incubation with appropriate secondary antibodies for 1 h at room temperature. Coverslips were mounted using ProLong Gold Antifade Mountant (Invitrogen) and analyzed by confocal microscopy using the Zeiss LSM510 Laser Scanning Microscope.

Jia J., Bissa B., Brecht L., Allers L., Choi S.W., Gu Y., Zbinden M., Burge M.R., Timmins G., Hallows K., Behrends C, & Deretic V. (2020). AMPK, a key regulator of metabolism and autophagy, is activated by lysosomal damage via a novel galectin-directed ubiquitin signal transduction system. Molecular cell, 77(5), 951-969.e9.

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