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Zen 2012 sp5

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The ZEN 2012 SP5 software is a comprehensive imaging and analysis platform developed by Zeiss. It provides a user-friendly interface for controlling and configuring Zeiss microscopes, as well as tools for image acquisition, processing, and analysis.

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Lab products found in correlation

Lsm 780 confocal microscope, by Zeiss (3 mentions) Tunel assay kit, by Promega (2 mentions) Rat anti mouse ki67 clone sola15 antibody, by Thermo Fisher Scientific (2 mentions) Alexa fluor 488 anti rat secondary antibody, by Thermo Fisher Scientific (2 mentions) Lsm 780, by Zeiss (2 mentions) Lsm 710, by Zeiss (2 mentions) 34 channel quasar detector, by Zeiss (2 mentions) Lsm 780 microscope, by Zeiss (2 mentions) Gfp trap agarose beads, by Proteintech (1 mentions) Ld c apochromat 1.1 w, by Zeiss (1 mentions) C apochromat na 1.2 korr fcs m27, by Zeiss (1 mentions) Fluorescence mounting medium, by Agilent Technologies (1 mentions) Lsm 880 airyscan, by Zeiss (1 mentions) Elyra ps 1 confocal microscope, by Zeiss (1 mentions) Cy 3 affinipure donkey anti mouse igg h l, by Jackson ImmunoResearch (1 mentions) Ms 295 p1, by Thermo Fisher Scientific (1 mentions) Lipofectamine 3000, by GeneCopoeia (1 mentions) Lsm 780 confocal imaging system, by Zeiss (1 mentions) Fluo 4 am, by Thermo Fisher Scientific (1 mentions)

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ZEN 2012 software (475 citations)

13 protocols using zen 2012 sp5

1

Quantifying Apoptosis and Proliferation in Tumor Sections

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To determine extent of apoptosis induced by different formulations, a TUNEL assay kit (Promega) was used. Briefly, paraffin-embedded sections were deparaffinized with xylene and rehydrated using an ethanol gradient. Sections were incubated with the TUNEL enzyme mixture for 1 h following the manufacturer’s instructions and nuclei were stained using Prolong Gold Antifade Mountant with DAPI. The sections were imaged using an LSM710 confocal microscope (data collection: Zen 2012 SP5, Zeiss; data processing: Fiji-ImageJ version 1.52p).
Expression of proliferation marker ki67 was monitored using immunohistochemistry. Rehydrated tumor sections were immersed in sodium citrate buffer (pH 6.0) and heated at 95 °C for 20 min to retrieve antigen. Sections were blocked using goat serum (10% in Tris Buffered Saline), incubated with rat anti-mouse Ki67 (Clone SolA15) antibody (eBiosciences, catalog # 14-5698-82, 1:100 in blocking buffer) for 1 h followed by a 30-min incubation with Alexa-Fluor 488 anti-rat secondary antibody (Invitrogen catalog # A-11006, 1:500 in Tris Buffered Saline). Nuclei were stained using Prolong Gold Antifade Mountant with DAPI. The sections were imaged using an LSM710 confocal microscope (data collection: Zen 2012 SP5, Zeiss) and the nuclei were pseudo-colored to red during image processing using Fiji-ImageJ version 1.52p.
Majumder P., Singh A., Wang Z., Dutta K., Pahwa R., Liang C., Andrews C., Patel N.L., Shi J., de Val N., Walsh S.T., Jeon A.B., Karim B., Hoang C.D, & Schneider J.P. (2021). Surface-fill Hydrogel Attenuates the Oncogenic Signature of Complex Anatomical Surface Cancer in a Single Application. Nature nanotechnology, 16(11), 1251-1259.
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2

Quantifying Apoptosis and Proliferation in Tumor Sections

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To determine extent of apoptosis induced by different formulations, a TUNEL assay kit (Promega) was used. Briefly, paraffin-embedded sections were deparaffinized with xylene and rehydrated using an ethanol gradient. Sections were incubated with the TUNEL enzyme mixture for 1 h following the manufacturer’s instructions and nuclei were stained using Prolong Gold Antifade Mountant with DAPI. The sections were imaged using an LSM710 confocal microscope (data collection: Zen 2012 SP5, Zeiss; data processing: Fiji-ImageJ version 1.52p).
Expression of proliferation marker ki67 was monitored using immunohistochemistry. Rehydrated tumor sections were immersed in sodium citrate buffer (pH 6.0) and heated at 95 °C for 20 min to retrieve antigen. Sections were blocked using goat serum (10% in Tris Buffered Saline), incubated with rat anti-mouse Ki67 (Clone SolA15) antibody (eBiosciences, catalog # 14-5698-82, 1:100 in blocking buffer) for 1 h followed by a 30-min incubation with Alexa-Fluor 488 anti-rat secondary antibody (Invitrogen catalog # A-11006, 1:500 in Tris Buffered Saline). Nuclei were stained using Prolong Gold Antifade Mountant with DAPI. The sections were imaged using an LSM710 confocal microscope (data collection: Zen 2012 SP5, Zeiss) and the nuclei were pseudo-colored to red during image processing using Fiji-ImageJ version 1.52p.
Majumder P., Singh A., Wang Z., Dutta K., Pahwa R., Liang C., Andrews C., Patel N.L., Shi J., de Val N., Walsh S.T., Jeon A.B., Karim B., Hoang C.D, & Schneider J.P. (2021). Surface-fill Hydrogel Attenuates the Oncogenic Signature of Complex Anatomical Surface Cancer in a Single Application. Nature nanotechnology, 16(11), 1251-1259.
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3

Live Cell Fluorescence Microscopy Imaging

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Fluorescence microscopy using live cells was performed using an inverted confocal laser scanning microscope (LSM780; Carl Zeiss, Germany). For 3D and time-lapse images, HeLa cells expressing RFP-B23 were excited at a wavelength of 561 nm, and the emission signal band was detected at 570–630 nm. The interval for time-lapse imaging was 3 min, and z-stack images were taken at 0.5 µm intervals using a 5× magnification objective lens (C-Apochromat, 63×/1.2NA). All live cell measurements were performed at 37 °C in 5% CO2 culture conditions. Fluorescence images were processed using software (Zen 2012 SP5; Carl Zeiss, Germany) installed on an LSM780 confocal microscope system for 3D reconstruction or intensity profile analyses.
Kim T.K., Lee B.W., Fujii F., Kim J.K, & Pack C.G. (2019). Physicochemical Properties of Nucleoli in Live Cells Analyzed by Label-Free Optical Diffraction Tomography. Cells, 8(7), 699.
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4

Quantifying Faa1 Membrane Recruitment

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For Fig. 2 A, Fig. 3 D, and Fig. S1, A and B, wild type or mutant versions of Faa1-EGFP were recruited to GFP-Trap Agarose beads (gta, Chromotek). Assays were performed under equilibrium conditions with 1 mM of the prey liposomes in buffer (150 mM NaCl, 50 mM Hepes/Tris pH 7.5) labeled with 0.5% lissamine rhodamine-DHPE (L-1392; Invitrogen). Beads were imaged at room temperature with a Zeiss LSM700 confocal microscope (RRID:SCR_017377) and a Plan-Apochromat 20×/0.8 WD 0.55 mm objective. The imaging software was Zeiss ZEN 2012 SP5 (RRID:SCR_013672) running on Windows 10 (64-bit). For quantification, lines were drawn across each bead in Fiji (Version 2.9.0/1.53t, RRID:SCR_002285), and the maximal values across the lines were taken. Values were averaged for each sample within each replicate and then among replicates. The final values were then normalized to max. Beads from three independent experiments were quantified for the rhodamine signal.
For testing the recruitment of Faa1 to Atg9 proteoliposomes, Atg9-EGFP proteoliposomes were immobilized on GFP-Trap Agarose beads (gta, Chromotek). The assay was performed under equilibrium conditions with a prey concentration of 1 µM Faa1-mCherry. Imaging and analysis were carried out as stated above for the membrane–protein interaction assay.
Baumann V., Achleitner S., Tulli S., Schuschnig M., Klune L, & Martens S. (2024). Faa1 membrane binding drives positive feedback in autophagosome biogenesis via fatty acid activation. The Journal of Cell Biology, 223(7), e202309057.
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5

Quantification of Filipin and Lamp1 Distribution

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The weighted colocalization coefficients were obtained using the Zeiss Zen image acquisition software (ZEN 2012 SP5 [64 bit]; Release Version14.0.0.0) after the intensity thresholds were set based on single probe controls. Imaris Cell 9.8 (Oxford Instruments) was used to analyze filipin staining (2D fluorescence images) and the Lamp1 3D distribution. For filipin quantification of number of puncta per cell, the nuclei and cells were defined based on the inverted filipin signal and the smoothed filipin image, respectively, and the filipin puncta segmentation was done on the original filipin image. This was necessary for accurate cell separation. For the Lamp1 3D distribution in cells, the nuclei and vesicles were segmented based on their respective images, whereas the cells were defined based on the Lamp1 images following histogram equalization. Lamp1 distribution was quantified by the mean vesicle distance from the nucleus calculated from the shortest distance of each vesicle to the cell nucleus.
Cherry P., Lu L., Shim S.Y., Ebacher V., Tahir W., Schatzl H.M., Hannaoui S, & Gilch S. (2023). Loss of small GTPase Rab7 activation in prion infection negatively affects a feedback loop regulating neuronal cholesterol metabolism. The Journal of Biological Chemistry, 299(2), 102883.
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6

Imaging Embryo Development with Confocal Microscopy

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Embryos at the ten-somite stage were mounted in 0.8% low-melting agarose and imaged at 25 °C using a laser scanning confocal microscope (LSM 710, Carl Zeiss) running the software Zen 2012 sp5 (Carl Zeiss). Confocal images of the region of interest in ubiquitous or mosaic membrane-labelled embryos were taken either at 2.5 s intervals using a ×40 water immersion objective (LD C-Apochromat 1.1 W, Carl Zeiss) or through a 3D timelapse with 1.0 µm optical sections every 16 s using a ×25 water immersion objective (LD C-Apochromat 1.1 W, Carl Zeiss). Imaging of ferrofluid droplets in the embryo was done as previously described28 (link). To visualize myosin and actin dynamics at the cell–cell contact over time, confocal images of Tg(actb2:MA-mCherry2)hm29 × Tg(actb2:myl12.1-EGFP) and Tg(actb2:MA-mCherry2)hm29 × Tg(actb1:GFP-Has UTRN)e116) double transgenic embryos, respectively, were taken in the region of interest at 2 s intervals using a ×40 water immersion objective. Ferrofluid droplets were labelled using a custom-synthesized fluorinated rhodamine dye58 (link), which was diluted in the ferrofluid oil at a final concentration of 37 μM.
Mongera A., Pochitaloff M., Gustafson H.J., Stooke-Vaughan G.A., Rowghanian P., Kim S, & Campàs O. (2022). Mechanics of the cellular microenvironment as probed by cells in vivo during zebrafish presomitic mesoderm differentiation. Nature Materials, 22(1), 135-143.
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7

Laser-Induced DNA Damage Assay

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The total irradiation laser power of a fiber-coupled 355 nm Coherent laser was adjusted to 12.8 mW of the 80 mW laser via a 40X C-Apochromat NA 1.2 Korr FCS M27 water immersion objective on a Zeiss LSM780 confocal microscope. Micro-irradiation was performed across the cell nucleus in a single pixel strip (height 208 nm) with either 3 s (high energy) or 400 ms (low energy) scanning times. With these two energy levels, two different DNA damages were created; detectable oxidatively-induced base lesions (e.g., 8-oxoG) or SSBs, respectively. DSBs were created with greater than 2-fold higher energy irradiation, by increasing the irradiation power from 12.8 mW to 32 mW with 3 s irradiation time across the nucleus with the same objective and microscope as above. Spot irradiation was performed by docking 12.8 mW of the 355 nm laser light for 65 ms via a 40X C-Apochromat NA 1.2 Korr FCS M27 objective a 1 μm2 area of the nucleus using the FCS hardware of a Zeiss 780 confocal microscope with Zeiss Zen 2012 SP5 software.
Janoshazi A.K., Horton J.K., Zhao M.L., Prasad R., Scappini E.L., Tucker C.J, & Wilson S.H. (2019). Shining Light on the Response to Repair Intermediates in DNA of Living Cells. DNA repair, 85, 102749.
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8

Radiation-Induced DNA Damage Analysis

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siRNA‐transfected cells were grown on sterilized coverslips and treated with 10 Gy RT and then incubated at 37°C for the indicated times. Cells were fixed with 4% paraformaldehyde for 15 min, washed with PBS and then permeabilized with 0.5% Triton X‐100 diluted in PBS at room temperature for 15 min. After being washed with PBS, the cells were blocked with a blocking buffer (5% normal goat serum) at room temperature for 1 h and incubated with a gamma‐H2A histone family member x (γH2AX) antibody (1:200) diluted in blocking buffer at 4°C overnight. After being washed with PBS, cells were incubated with Alexa 555‐conjugated anti‐rabbit IgG (1:1000) at room temperature for 1 h in the dark. After being washed, the cells were incubated with Hoechst 33342 stain at room temperature for 10 min. The glass slides were washed with PBS and mounted with Dako fluorescence mounting medium. Fluorescence microscopy of the live cells was performed using an inverted confocal laser scanning microscope (LSM880 Airyscan; Carl Zeiss). Fluorescence images were processed using Zen 2012 SP5 software (Carl Zeiss of the LSM880 Airyscan confocal microscopy system for 3D reconstruction or intensity profile analyses). Foci were counted using ImageJ software.
Park S.S., Kwon M.R., Ju E.J., Shin S.H., Park J., Ko E.J., Son G.W., Lee H.W., Kim Y.J., Moon G.J., Park Y., Song S.Y., Jeong S, & Choi E.K. (2023). Targeting phosphomevalonate kinase enhances radiosensitivity via ubiquitination of the replication protein A1 in lung cancer cells. Cancer Science, 114(9), 3583-3594.
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9

Inverted Confocal Microscopy Imaging

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An inverted confocal LSM780 microscope (Zeiss, Jena, Germany) was used in all experiments with either a 10× (NA = 0.3), 40× (NA = 1.4, water immersive), or a 100× (NA = 1.46, oil immersive) objective. Excitation at 405, 488, 561, and 633 nm was used to visualize Hoechst 33342, Alexa Fluor 488, Alexa Fluor 555/594, allophycocyanin and Alexa 700 fluorescence, respectively. Emission was measured in CLSM λ-mode using a 34-channel QUASAR detector (Zeiss) set to a 405–695 nm range. For quantitative analysis, images were captured as 2 × 2 tile grids at the same regions of each specimen using the 40 × objective, with an individual xyz tile size of 354 × 354 × 30 μm. Higher magnification images were acquired in z-stacks at the region of interest using the 100× objective. Spectral unmixing was performed using ZEN 2012 SP5 software (Zeiss). Finally, the images were processed using Adobe Photoshop CS version 5 (Adobe Systems, Mountain View, CA).
Bogorodskiy A.O., Bolkhovitina E.L., Gensch T., Troyanova N.I., Mishin A.V., Okhrimenko I.S., Braun A., Spies E., Gordeliy V.I., Sapozhnikov A.M., Borshchevskiy V.I, & Shevchenko M.A. (2020). Murine Intraepithelial Dendritic Cells Interact With Phagocytic Cells During Aspergillus fumigatus-Induced Inflammation. Frontiers in Immunology, 11, 298.
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10

Immunofluorescence Staining for Lamin A/C and Troponin T

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The cells were fixed with 4% paraformaldehyde for 20 min at room temperature. Then they were washed three times with PBS 1x for 5 min. Next, the cells were permeabilized with 0.3% Triton diluted in 1x PBS for 15 min. Subsequently, blocking was done using a 5% BSA solution diluted in PBS 1x for 60 min. We then placed the primary antibody in question in PBS-BSA 3% at 1:100 (LMNA, cat. MA3-1,000, ThermoFisher scientific) for Lamin A/C; and 1:200 (Troponin T, cat. MS-295-P1, ThermoFisher scientific) for Troponin T, cardiac isoform, dilutions, and incubated overnight at 4°C. The following day, we washed 3x with 1x PBS for 5 min. After the washes, we placed the secondary antibody Cy™3 AffiniPure Donkey Anti-Mouse IgG (H + L) (cat. 715150, Jackson ImmunoResearch) diluted in PBS-BSA 3% at a 1:400 ratio for 2 h at room temperature. After incubation, we washed 3x with 1x PBS for 5 min. Finally, we placed DAPI (cat. D9542, Sigma-Aldrich) for 5 min and washed 3x times with PBS 1x for 5 min. The coverslips were sealed using 13 µL of Fluoroumont (ThermoFisher scientific). Images were taken with the ×100 objective on the Elyra PS.1 confocal microscope (Carl Zeiss), ZEN 2012 SP5 software (Carl Zeiss), at the Advanced Microscopy Unit (UMA) of the National Center for Structural Biology and Bioimaging (UFRJ, Rio de Janeiro).
Monnerat G., Kasai-Brunswick T.H., Asensi K.D., Silva dos Santos D., Barbosa R.A., Cristina Paccola Mesquita F., Calvancanti Albuquerque J.P., Raphaela P.F., Wendt C., Miranda K., Domont G.B., Nogueira F.C., Bastos Carvalho A, & Campos de Carvalho A.C. (2022). Modelling premature cardiac aging with induced pluripotent stem cells from a hutchinson-gilford Progeria Syndrome patient. Frontiers in Physiology, 13, 1007418.
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