Alex fluor 594

Manufactured by Jackson ImmunoResearch

Alex Fluor 594 is a fluorescent dye that can be used for various biological applications. It has an excitation maximum at 590 nm and an emission maximum at 617 nm, making it suitable for detection and visualization in fluorescence-based techniques.

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Lab products found in correlation

Lsm 700 confocal microscope, by Zeiss (2 mentions) Mouse on mouse fluorescein kit, by Vector Laboratories (1 mentions) Anti gfap, by Thermo Fisher Scientific (1 mentions) Anti neun, by Merck Group (1 mentions) Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (1 mentions) Donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions) Nb100 2288, by Novus Biologicals (1 mentions) Tcs sp5 confocal microscope, by Leica (1 mentions) Alexa fluor 488, by Jackson ImmunoResearch (1 mentions) Anti lamin a c, by Merck Group (1 mentions) Anti flag, by Merck Group (1 mentions) Ab179475, by Abcam (1 mentions) Ix83 inverted microscope, by Hamamatsu Photonics (1 mentions) F3648, by Merck Group (1 mentions) Orca flash 4.0 camera, by Hamamatsu Photonics (1 mentions)

4 protocols using alex fluor 594

Immunohistochemical Analysis of Mouse Brain

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Anesthetized mice were perfused transcardially with heparinized PBS followed by 10% NBF. Three-mm pieces were coronally using a cutting matrix and immersed overnight in 10% NBF and then embedded in paraffin. Ten μm FFPE brain sections were deparaffinized and rehydrated. Antigen retrieval was then performed using 10 mM sodium citrate buffer (pH 6.0). Brain sections were stained with anti-NeuN (Clone A60; Millipore, Darmstadt, Germany) or anti-APC (Clone CC-1; Abcam, Cambridge, UK) for 30 min at room temperature using the Mouse-On-Mouse Fluorescein Kit (Vector, Burlingame, CA), then stained overnight at 4°C with rabbit anti-VP1 [graciously provided by R. Garcea (University of Colorado, Boulder, CO)] followed by donkey anti-rabbit IgG conjugated to Alex Fluor 594 (Jackson ImmunoResearch, West Grove, PA). Astrocyte staining was performed using directly conjugated anti-GFAP (Clone GA5; eBioscience, San Diego, CA). Tissue sections were then mounted with ProLong Gold Anti-Fade Reagent with DAPI (Life Technologies, Carlsbad, CA). Images were acquired using a Leica DM4000 B LED microscope (Leica-Camera, Wetzlar, Germany).

S.h.w.e.t.a.n.k., Frost E.L., Mockus T.E., Ren H.M., Toprak M., Lauver M.D., Netherby-Winslow C.S., Jin G., Cosby J.M., Evavold B.D, & Lukacher A.E. (2019). PD-1 Dynamically Regulates Inflammation and Development of Brain-Resident Memory CD8 T Cells During Persistent Viral Encephalitis. Frontiers in Immunology, 10, 783.

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Immunohistochemical Analysis of γ2 Subunit in Sim1-Cre Mice

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For immunohistochemistry studies, we used the previously descried protocol^{20 (link)}. Brain sections were prepared from control or Sim1-Cre::γ2^flox/flox::γ2-TB^flox/flox male adult mice (8–10 weeks) after cardiac perfusion at ZT0, 6, 12, and 18. For validation of AAV-GFP and AAV-Cre-GFP mediated γ2 subunit deletion and replacement, brains were harvested at the end of studies with cardiac perfusion, except mentioned otherwise in the text. Primary antibody against γ2 (1:100, NB300-190, Novus Biologicals), BMAL1 (1:200, NB100-2288, Novus Biologicals), c-Fos (1:1000, #2250, Cell Signaling Technology), and cleaved caspase-3 (1:300, #9664, Cell Signaling) were used. The sections were visualized with Alexa Fluor 488® or Alex Fluor 594® (1:200, Jackson immunoResearch Laboratories, Inc.). The signal was captured and imaged with a TCS SP5 confocal microscope (Leica, Nussloch, Germany).
For c-Fos counting, five sections that contain the PVH at matched anteroposterior levels were chosen from each mouse. Within the PVH boundary, all c-Fos-positive cells with bright and clear oval nuclei profiles were counted under the fluorescent microscope with the 200× objective. The number from all sections were averaged and analyzed in all animals.

Kim E.R., Xu Y., Cassidy R.M., Lu Y., Yang Y., Tian J., Li D.P., Van Drunen R., Ribas-Latre A., Cai Z.L., Xue M., Arenkiel B.R., Eckel-Mahan K., Xu Y, & Tong Q. (2020). Paraventricular hypothalamus mediates diurnal rhythm of metabolism. Nature Communications, 11, 3794.

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Immunofluorescence Assay for Lamin A/C

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Cells grown on coverslips were fixed 24 h after transfection with 3% paraformaldehyde, permeabilized in PBS/0.5% Triton X-100, and incubated in PBS/0.1% Triton X-100/2% BSA for 25 min. Cells were incubated for 30 min each with primary and secondary antibodies in PBS/0.1% Triton X-100/1% BSA. Antibodies were anti-Flag (1:200; Sigma), anti-lamin A/C [50 (link)] (1:400), and anti-rabbit Alex Fluor® 594 (1:200; Jackson ImmunoResearch). DNA was stained with Hoechst 33258. Coverslips were mounted with Mowiol and examined on a LSM 700 confocal microscope (Zeiss) at the Imaging Facility of the Functional and Adaptive Biology Unit of University Paris Diderot/CNRS.

Paulsen J., Sekelja M., Oldenburg A.R., Barateau A., Briand N., Delbarre E., Shah A., Sørensen A.L., Vigouroux C., Buendia B, & Collas P. (2017). Chrom3D: three-dimensional genome modeling from Hi-C and nuclear lamin-genome contacts. Genome Biology, 18, 21.

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Visualizing Integrin Dynamics in HT-1080 Cells

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Non-transfected or GFP-VANGL2 transfected HT-1080 cells were plated on vitronectin or fibronectin treated coverslip dishes. Cells were left untreated or treated with MnCl₂ and fixed using 4% paraformaldehyde/PBS at room temperature or ice-cold methanol at −20°C. Washed cells were blocked using PBS, 5% normal donkey serum, and 0.1% Triton X-100 for 1 h at room temperature. Cells were incubated with LAMP1 (1:100, D2D11 XP; Cell Signaling Technology), fibronectin (1:100, F3648, Millipore Sigma), paxillin (1:100, AHO0492, Thermo Fisher), or integrin αv (1:100, ab179475, Abcam) primary antibody in block for 1 h at room temperature, washed twice with PBS, and incubated an additional 1 h in the dark with Donkey anti-Rabbit or anti-mouse Alex Fluor 594 (1:500, 711–585-152/715–585-150; Jackson ImmunoResearch Laboratories). Cells were washed three times with PBS. NucBlue Fixed Cell DAPI Stain (Thermo Fisher Scientific) was used to label nuclei. Imaging was performed using a 40x oil objective (NA = 1.3) or 20x dry objective (NA = 0.75) and an Olympus IX83 inverted microscope equipped with a Hamamatsu Orca Flash 4.0 camera. For some images, maximum intensity projections were generated from z-stack slices (0.5 μm).

Jessen T.N, & Jessen J.R. (2018). VANGL2 protein stability is regulated by integrin αv and the extracellular matrix. Experimental cell research, 374(1), 128-139.

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