Triton x 100

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Triton X-100 is a non-ionic surfactant commonly used in various laboratory applications. It functions as a detergent and solubilizing agent, facilitating the solubilization and extraction of proteins and other biomolecules from biological samples.

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Lab products found in correlation

Bovine serum albumin (bsa), by Merck Group (1314 mentions) Paraformaldehyde (pfa), by Merck Group (885 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (727 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (663 mentions) Hoechst 33342, by Thermo Fisher Scientific (386 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (338 mentions) Dimethyl sulfoxide (dmso), by Merck Group (304 mentions) Tween 20, by Merck Group (287 mentions) Alexa fluor 488, by Thermo Fisher Scientific (269 mentions) Propidium iodide, by Merck Group (234 mentions) Sodium chloride (nacl), by Merck Group (207 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (194 mentions) Paraformaldehyde (pfa), by Electron Microscopy Sciences (180 mentions) Sodium dodecyl sulfate (sds), by Merck Group (177 mentions) Phosphate buffered saline (pbs), by Merck Group (175 mentions) Lsm 710, by Zeiss (158 mentions) Formaldehyde, by Merck Group (151 mentions) Goat serum, by Merck Group (150 mentions) Paraformaldehyde (pfa), by Thermo Fisher Scientific (148 mentions) Bovine serum albumin (bsa), by Thermo Fisher Scientific (146 mentions)

11 070 protocols using triton x 100

Immunolabeling GFAP in Cell Cultures

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For immunolabeling with GFAP, the
cultures were fixed in 4% paraformaldehyde for 15 min. After washing
with PBS, the cells were permeabilized with 0.5% Triton X-100 (Sigma-Aldrich,
USA) in PBS for 5 min and for 10 min. It was followed by blocking
with 4% bovine serum albumin + 1% normal goat serum in 0.25% Triton
X-100 (Sigma-Aldrich, USA) for 1 h at room temperature and, subsequently,
in a mixture of the primary anti-GFAP rabbit antibody (G9269 Sigma-Aldrich,
USA) overnight at 4 °C. After washing in PBST 0.25% Triton X-100,
goat anti-rabbit conjugated with Texas red secondary antibodies applied
for 2 h at room temperature followed by DAPI staining for 5 min followed
by three washing with 0.1% Triton X-100.
Images were captured
using a citation 5 (Biotek, Agilent Technoloies, USA) fluorescence
microscope, and quantification was performed as reported earlier.^{57 (link),58 (link)}

Mahaling B., Pandala N., Wang H.C, & Lavik E.B. (2022). Azithromycin Protects Retinal Glia Against Oxidative Stress-Induced Morphological Changes, Inflammation, and Cell Death. ACS Bio & Med Chem Au, 2(5), 499-508.

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Immunohistochemistry of Inner Ear Tissues

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Utricles were dissected in ice-cold HBSS and fixed in 4% formaldehyde for 1 hr at room temperature. Whole inner ears were fixed for 18 hr at 4˚C, treated with 0.88 M sucrose for 18 hr at 4˚C, embedded in Tissue-Tek O.C.T. (Sakura), and frozen in liquid-nitrogen vapor. Wholemounted sensory epithelia or 10 μm frozen sections were then blocked with 3% normal donkey serum (Sigma-Aldrich) in 500 mM NaCl, 0.3% Triton X-100 (Sigma-Aldrich), and 20 mM tris(hydroxymethyl)aminomethane (Bio-Rad) at pH 7.5. The primary antisera—goat anti-Sox2 (Santa Cruz), rabbit anti-Myo7A (Proteus Bioscience), rabbit anti-GFP (Torrey Pines Biolabs), mouse anti-Yap (Santa Cruz), and rabbit anti-Yap (Cell Signaling)—were reconstituted in blocking solution and applied overnight at 4˚C. For labeling with E-cadherin antibodies from clone DECMA-1 (Millipore) no Triton X-100 was used in the blocking solution.
Samples were washed with phosphate-buffered saline solution supplemented with 0.1% Tween 20 (Sigma-Aldrich), after which Alexa Fluor-labeled secondary antisera (Life Technologies) were applied in the same solution for 1 hr at room temperature.
Phalloidin conjugated to Alexa 633 was used to label filamentous actin and nuclei were stained with 3 μM DAPI.

Gnedeva K., Jacobo A., Salvi J.D., Petelski A.A, & Hudspeth A.J. (2017). Elastic force restricts growth of the murine utricle. eLife, 6, e25681.

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Immunohistochemical Labeling of Recorded Neurons

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After patch-clamp recordings, the recording electrode was carefully withdrawn, and the slices were fixed overnight at 4 ºC in 4% paraformaldehyde in phosphate buffer solution (PBS). After thorough rinsing with PBS and permeabilization with 0.3% Triton X-100 (Sigma-Aldrich), the slices were incubated overnight with streptavidin-CY3 (1:50, 438,315, Invitrogen, USA) or streptavidin-CY5 (1:50, 438,315, Invitrogen, USA). Slices were then rinsed with PBS and mounted onto Fisher SuperFrost slides for visualization in an Olympus FV1000 confocal microscope. Low magnification (10×) was used to localize the recorded neurons, confirming that they were within the boundaries of the VTA. Slices that contained a labeled neuron were transferred back to a PBS solution and after additional permeabilization with 0.3% Triton X-100, they were incubated overnight with a mouse anti-TH antibody (1:100, MAB318, Millipore, Burlington, MA). Sections were rinsed with PBS and then incubated with secondary Donkey anti-Mouse Alexa Fluor 405 (1:100, ab175658, abcam, Cambridge, MA) for 1 hr at 30°C. Sections were rinsed with PBS and then mounted onto Fisher SuperFrost slides and visualized using higher magnification (40×) on an Olympus FV1000 confocal microscope.

Miranda-Barrientos J., Chambers I., Mongia S., Liu B., Wang H.L., Mateo-Semidey G.E., Margolis E.B., Zhang S, & Morales M. (2021). Ventral tegmental area GABA, glutamate, and glutamate-GABA neurons are heterogeneous in their electrophysiological and pharmacological properties. The European journal of neuroscience.

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Immunofluorescence Analysis of Neuronal Markers

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Cells were fixed with 4% paraformaldehyde for 15 min at room temperature (RT). Then, the cells were permeabilized with 0.5% Triton X-100 (Sigma, USA) for 5 min and blocked with 10% goat serum (Millipore, USA) diluted in 0.5% Triton X-100 and PBS for 30 min at RT. Primary antibodies were diluted into blocking liquid and incubated overnight at 4 °C. Then, cells were transferred into secondary antibodies and incubated for 1 h at RT in dark. Primary and secondary antibodies were listed in Table S5. The nucleus was stained with DAPI (Beyotime, China) for 5 min. Coverslips were mounted with a fluorescence quencher. Images were captured with IX73 Olympus inverted microscope (Olympus, Japan) or confocal microscope imager LSM710 or LSM800 (Zeiss, Germany), and image analyses were performed with software ZEN (Zeiss, Germany). Neuronal markers of TUJ1, MAP2, GABA, GFAP, SYP1, and PSD95, were measured by the fluorescence quantitative analysis using ImageJ software. 5–8 images were analyzed from each image.

He L., Wang S., Peng L., Zhao H., Li S., Han X., Habimana J.D., Chen Z., Wang C., Peng Y., Peng H., Xie Y., Lei L., Deng Q., Wan L., Wan N., Yuan H., Gong Y., Zou G., Li Z., Tang B, & Jiang H. (2021). CRISPR/Cas9 mediated gene correction ameliorates abnormal phenotypes in spinocerebellar ataxia type 3 patient-derived induced pluripotent stem cells. Translational Psychiatry, 11, 479.

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Whole Mount Tissue Clearing and Staining

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Whole mount aortic arch, mediastinal adipose tissue, and thymus were fixed in Cytofix (BD Biosciences) 1:3 diluted in PBS overnight. The tissue block was washed in PBS for a day and blocked with 1% mouse serum (Milipore Sigma), 1% BSA (Milipore SIGMA), 0.3% TritonX-100 in PBS for 24h at room temperature while agitating. Subsequently, primary fluorescently conjugated antibodies were added 1:100 for 72h at 37°C in the dark while agitating. Hoechst (Thermo Scientific) was added at a concentration of 1:1000 for the final 2h. After staining, the whole mount tissue block was washed in PBS with 0.2% Triton X-100 and 0.5% 1-Thioglycerol (Milipore Sigma) overnight at room temperature. The buffer was replaced twice. C_e3D clearing solution (6 (link)) was freshly prepared as follows:
1ml contained 400μl (40%) N-methylacetamide, 1μl of Triton X-100 (0.1%), 5μl 1-Thioglycerol (0.5%) (all obtained from Milipore Sigma). For optimal solvation of all reagents, the buffer was shaken at 37°C for several hours. The whole mount tissue was removed from the washing buffer, carefully dried, and transferred into the clearing solution overnight at room temperature in the dark. The following day, the cleared whole mount tissue was mounted between two 1.5 borosilicate glass coverslips in clearing medium.

Winkels H., Ghosheh Y., Kobiyama K., Kiosses W.B., Orecchioni M., Ehinger E., Suryawanshi V., Herrera-De La Mata S., Marchovecchio P., Riffelmacher T., Thiault N., Kronenberg M., Wolf D., Seumois G., Vijayanand P, & Ley K. (2021). Thymus-derived CD4+CD8+ cells reside in mediastinal adipose tissue and the aortic arch. Journal of immunology (Baltimore, Md. : 1950), 207(11), 2720-2732.

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Immunofluorescence Staining Protocol

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Slides were washed with PBS solution, fixed with 4% paraformaldehyde (Sigma-Aldrich, Saint Louis, Missouri, USA) for 10 min at room temperature, and permeabilized with 0.2% Triton X-100 (Sigma-Aldrich, Saint Louis, Missouri, USA) in PBS for 10 min. Then slides were blocked with 3% BSA (Sigma-Aldrich, Saint Louis, Missouri, USA) in PBS with 0.1% Triton X-100 for 10 min, followed by incubation with indicated primary antibodies (Table S3) at 4°C overnight. Then slides were washed three times with 0.1% Triton X-100 in PBS, each time for 10 min and incubated with fluorescence-labeled secondary antibodies in dark at room temperature for 1 hour. Nuclei were visualized by DAPI (Molecular Probes, Eugene, Oregon, USA) staining. Coverslips were mounted onto slides by VECTASHIELD HardSet Mounting Medium (Vector Laboratories Inc., Burlingame, CA, USA). Labeled sections were imaged using Nikon A1R-A1 confocal microscopy system.

Wong Y.Q., Xu H., Wu Q., Liu X., Lufei C., Xu X.Q, & Fu X.Y. (2018). STAT3-Inducible Mouse ESCs: A Model to Study the Role of STAT3 in ESC Maintenance and Lineage Differentiation. Stem Cells International, 2018, 8632950.

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RNA Isolation from Cell Fractions

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The protocol for RNA isolation from different cell fractions was adapted from Rio et al.⁸¹ Briefly, 0.6 × 10⁶ cells (70% confluency) were trypsinized and collected by centrifugation. Cell pellets were washed twice with cold PBS, after which the cell pellet was resuspended in 0.5 mL of ice-cold cell disruption buffer (20 mM Tris-HCl [pH 7.5], 1.5 mM MgCl₂, 10 mM KCl, and 10 mM dithiothreitol). After 10 min in cell disruption buffer on ice, the cells were transferred to a 1-mL glass Dounce Tissue Grinder (Wheaton) and homogenized using 15 strokes with a tight pestle. The crude cell lysate was transferred to a fresh tube and 5 μL of 10% Triton X-100 (Sigma-Aldrich) was added to a final concentration of 0.1% Triton X-100 and mixed by inversion five times. Cell nuclei were pelleted immediately by centrifuging the homogenate at 1,500 × g at 4°C for 5 min. The supernatant containing the cytoplasmic fractions was collected. RNA was isolated from both fractions using the Aurum Total RNA Mini Kit (Bio-Rad) according to the manufacturer’s protocol. All RNA preparations were quantified using a NanoVue spectrometer (GE Healthcare Europe). Approximately 500 ng of RNA of the cytoplasmic fractions and equal volumes of the corresponding nuclear fractions were used as a template for subsequent cDNA synthesis and RT-qPCR.

van Agtmaal E.L., André L.M., Willemse M., Cumming S.A., van Kessel I.D., van den Broek W.J., Gourdon G., Furling D., Mouly V., Monckton D.G., Wansink D.G, & Wieringa B. (2017). CRISPR/Cas9-Induced (CTG⋅CAG)n Repeat Instability in the Myotonic Dystrophy Type 1 Locus: Implications for Therapeutic Genome Editing. Molecular Therapy, 25(1), 24-43.

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Immunofluorescence Staining of MCF-7 Cells

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MCF-7 cells cultured in a 6-well plate were stimulated with EA extract (30 μg/mL) for 72 h. At the end of the incubation period, the cells were washed two times with ice-cold PBS and fixed with 4% paraformaldehyde in PBS for 30 min. Fixed cells were washed again with PBS, permeabilized with 2% Triton X-100 (Sigma) for 30 min, and blocked with 30% FCS in 0.01% Triton X-100 for 1 h. The final step consisted in staining the cells with DAPI (4’,6’-diamidino–2-phenylindole) in a dark chamber for 15 min. Cell nuclei were analyzed at a magnification of 40× with a fluorescence Olympus IX73 microscope equipped with an integrated DP74 camera (Olympus, Tokyo, Japan).

Danciu C., Muntean D., Alexa E., Farcas C., Oprean C., Zupko I., Bor A., Minda D., Proks M., Buda V., Hancianu M., Cioanca O., Soica C., Popescu S, & Dehelean C.A. (2018). Phytochemical Characterization and Evaluation of the Antimicrobial, Antiproliferative and Pro-Apoptotic Potential of Ephedra alata Decne. Hydroalcoholic Extract against the MCF-7 Breast Cancer Cell Line. Molecules, 24(1), 13.

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Immunostaining of PVR Membrane Samples

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PVR membrane specimens were fixed in 4% paraformaldehyde solution and washed with PBS. Then, the samples were permeabilized with 0.1% Triton‐X‐100 (Sigma‐Aldrich) and blocked with 2.5% BSA in PBS. Subsequently, all samples were immunostained with the indicated primary antibodies overnight at 4°C and the indicated secondary antibodies for 2 hours at room temperature. Thereafter, samples were counterstained with DAPI (Sigma‐Aldrich) and examined with an Olympus FluoView 1000 confocal microscope (Olympus).
For cellular immunofluorescence staining, cells were fixed in 4% paraformaldehyde solution for 15 minutes, permeabilized with 0.1% Triton‐X‐100 (Sigma‐Aldrich) for 15 minutes and then blocked with 2.5% BSA for 1 hour. In the following, cells were incubated with the indicated primary antibodies overnight at 4°C and the indicated secondary antibodies for 1 hour at room temperature. After being washed, cells were observed with an Olympus FluoView 1000 confocal microscope (Olympus).

Miao Q., Xu Y., Yin H., Zhang H, & Ye J. (2020). KRT8 phosphorylation regulates the epithelial‐mesenchymal transition in retinal pigment epithelial cells through autophagy modulation. Journal of Cellular and Molecular Medicine, 24(5), 3217-3228.

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Immunofluorescence Analysis of Myoblast Development

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Immunofluorescence was performed in myoblast cell cultures at days 1, 3 and 7 of development in coverslips in 6-well plates. The myoblasts were washed with PBS and then fixed in 4% paraformaldehyde for 15 minutes. The cells were permeabilized with 0.1% Triton X-100 (Sigma-Aldrich, USA) for 10 minutes and incubated in blocking solution (1% glycine, 3% bovine serum albumin (BSA), 8% fetal bovine serum and 0.3% Triton X-100 - Sigma-Aldrich, USA) for 1 hour, to prevent nonspecific binding. The myoblasts were incubated overnight at 4 °C with a rabbit anti-desmin primary antibody (Sigma-Aldrich, USA) diluted in blocking solution (1:20). The cells were washed and then incubated for 2 hours at 4 °C with an anti-rabbit FITC secondary antibody (sc-2090 - Santa Cruz, USA) diluted in blocking solution (1:400). The myoblasts nuclei were counterstained with DAPI present in Vectashield® mounting medium (Vector Laboratories Inc., USA) and the images were acquired under a fluorescence microscope (Olympus, Japan).

Duran B.O., Góes G.A., Zanella B.T., Freire P.P., Valente J.S., Salomão R.A., Fernandes A., Mareco E.A., Carvalho R.F, & Dal-Pai-Silva M. (2019). Ascorbic acid stimulates the in vitro myoblast proliferation and migration of pacu (Piaractus mesopotamicus). Scientific Reports, 9, 2229.

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