Ferric slat, gold (III) chloride (HAuCl4), other reagents related to AuNC@Fe3O4 synthesis and 3,3′,5,5′-Tetramethylbenzidine (TMB) were purchased from Sigma, Inc. (St. Louis, United States). H2O2 solution and different pH buffer solutions (pH = 2, 3, 4, 5, 6, 7, 8, and 9) were bought from Aladdin (Shanghai, China). Human umbilical vein endothelial cells (HUVEC), human HCC cell lines (HepG2 cells) and the specific culture mediums for the two cell lines were purchased from Procell (Wuhan, China). Cell Counting Kit-8 (CCK-8) was obtained from Sangon Biotech (Shanghai, China). 2′,7′-Dichlorodihydrofluorescein diacetate (DCFH-DA) was obtained from MedChemExpress (New Jersey, United States). Calcein-AM/propidium iodide (PI) kit, Annexin V-FITC apoptosis detection kit, Lyso-Tracker Red kit and Hoechst 33342 staining solution were purchased from Beyotime. Inc. (Shanghai, China).
Lyso tracker red kit
Manufactured by Beyotime
Sourced in China
The Lyso-Tracker Red kit is a fluorescent probe designed to label and track lysosomes in living cells. The kit provides a red-fluorescent dye that selectively stains acidic organelles, allowing for the visualization and study of lysosomal dynamics within the cellular environment.
Automatically generated - may contain errors
Lab products found in correlation
Annexin 5 fitc apoptosis detection kit, by Beyotime (2 mentions)
Lsm 780, by Zeiss (2 mentions)
3 3 5 5 tetramethylbenzidine (tmb), by Merck Group (1 mentions)
Gold 3 chloride haucl4, by Merck Group (1 mentions)
2 7 dichlorodihydrofluorescein diacetate dcfh da, by MedChemExpress (1 mentions)
Cell counting kit 8 cck 8, by Sangon (1 mentions)
Hoechst 33342 staining solution, by Beyotime (1 mentions)
Eclipse ni microscope, by Nikon (1 mentions)
Trichloromethane chcl3, by Beijing Chemical Works (1 mentions)
Methanol chromatographic reagent grade, by Thermo Fisher Scientific (1 mentions)
N n dimethylformamide (dmf), by Beijing Chemical Works (1 mentions)
Ros assay kit, by Beyotime (1 mentions)
Fetal bovine serum (fbs), by Corning (1 mentions)
Mitochondrial membrane potential assay kit with jc 1, by Beyotime (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Penicillin streptomycin mixture, by Solarbio (1 mentions)
Alexa fluor 594 conjugated secondary antibody, by Abcam (1 mentions)
Paraformaldehyde (pfa), by Solarbio (1 mentions)
Phalloidin ifluor 594 reagent, by Abcam (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
5 protocols using lyso tracker red kit
1
Synthesis and Cytotoxicity of AuNC@Fe3O4
2
Lysosomal Activity Quantification
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Cell lysosomal fluorescence intensity was estimated with a LysoTracker Red kit (Beyotime Biotechnology, Beijing, China). Higher fluorescence intensity represents stronger lysosomal activity. LysoTracker Red was added to each well, and the plates were incubated at 37°C for 2 h according to the manufacturer’s protocol. LysoTracker Red was then removed, and fresh medium was added. A fluorescence microscope (Nikon Eclipse NI microscope, Nikon, Inc., Japan) was used for imaging. Lysosomes showed bright fluorescence staining, and the results were analyzed by ImageJ analysis software.
3
Curcumin-Loaded MOF Nanoparticles
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Zinc nitrate hexahydrate (Zn(NO3)2·6H2O) was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. (Shanghai, China); triethylamine (TEA) and 4′,4-biphthalic acid (4,4′-BPDC) were bought from the Tianjin Guangfu Fine Chemical Research Institute (Tianjin, China); N,N-dimethylformamide (DMF) and trichloromethane (CHCl3) were obtained from the Beijing Chemical Factory (Beijing, China); methanol (chromatographic reagent grade) was obtained from Thermo Fisher Scientific (Shanghai, China); curcumin (CUR)( Figures S4–S7 ) was purchased from Shanghai Yuanye Bio-Technology Co., Ltd. (LOT:R12A10S85604, 98%); dimethyl sulfoxide (DMSO), high-glucose Dulbecco’s modified Eagle’s medium (DMEM), PBS, and a penicillin–streptomycin mixture were purchased from Solarbio (Beijing, China); fetal bovine serum (FBS) was acquired from Corning; 3-(4,5-dimethylthiazol-2-yl)-2,5-dipheny-ltetrazolium bromide (MTT) was acquired from Beijing BioDee Biotechnology Co., Ltd. (Beijing, China); 4′,6-diamidino-2-phenylindole (DAPI), an Annexin V-FITC Apoptosis Detection Kit, Lyso-Tracker Red Kit, ROS assay kit, and mitochondrial membrane potential assay kit with JC-1 were purchased from Shanghai Beyotime Biotechnology Co., Ltd. (Shanghai, China); HepG2 cells were purchased from Guangzhou Jeniobio Biotechnology (Beijing, China).
4
Endocytic Pathway Inhibitors in Cell Studies
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The inhibitors used in this study are as follows. Chlorpromazine and sucrose inhibit clathrin-mediated endocytosis; methyl-β-cyclodextrin (MβCD) and nystatin inhibit caveolin-mediated endocytosis; rottlerin and NSC23766 inhibit macropinocytosis; chloroquine and bafilomycin A1 inhibit acidification of endosomes; cytochalasin D and CK-636 inhibit actin polymerization; nocodazole and vinblastine depolymerize microtubles. All inhibitors were purchased from Selleck (USA) and Sigma-Aldrich (USA). All inhibitors, except sucrose, were dissolved in dimethyl sulfoxide (DMSO) (Sigma, USA) according to the manufacturer's instructions. Tubule-Tracker red kit and Lyso-Tracker red kit was purchased from Beyotime Biotechnology (Beijing, China). Fluorescein isothiocyanate (FITC), 4′-6-diamidino-2-phenylindole (DAPI), formaldehyde and paraformaldehyde (PFA) was purchased from Solarbio (Beijing, China). Latex beads (1 μm) were purchased from Polysciences (USA). Mouse monoclonal antibody against clathrin heavy chain and caveolin-1, rabbit polyclonal antibodies against rab5, lamp1, and cathepsin D, phalloidin-iFluor 594 Reagent and Alexa Fluor 594-conjugated secondary antibodies were purchased from Abcam (UK) and ABclonal (USA). Rat polyclonal antibodies against E. tarda have been reported previously (Zhou and Sun, 2016 (link)).
5
Autophagy Quantification in Drosophila Wing Discs
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Cells were washed three times with ice-cold PBS and were fixed for 15 min at room temperature with Immunol Staining Fix Solution (Beyotime, P0098) and then permeabilized with Immunostaining Permeabilization Buffer with Triton X-100 (Beyotime, P0096) for 20 min. Following permeabilization, nonspecific binding in the cells was blocked by Immunol Staining Blocking Buffer (Beyotime, P0102) for 1 h at room temperature. Then cells were incubated for 1 h with specific primary antibodies and after three washes with PBS, the cells were incubated for another 1h with secondary antibodies.
Lysosomal activity as a marker of autophagy in Drosophila wing imaginal discs was detected by the LysoTracker Red Kit (Beyotime, C1046). Imaginal discs dissected from third-instar larvae were collected in PBS and incubated with LysoTracker Red (1:3,000) for 15 min at 37 °C, washed with PBS three times prior to imaging. Primary antibodies included: rabbit anti-FoxO3A (1:200; Abcam, ab12162), mouse anti–Myc-tag (1:350, CST, 9B11), rabbit anti-FLAG antibody (1:200, Sigma). Secondary antibodies were anti-mouse CY3 (1:1,000) and anti-rabbit Alexa Flour 488 (1:500). All images were collected with a confocal microscope (Zeiss LSM 780).
Lysosomal activity as a marker of autophagy in Drosophila wing imaginal discs was detected by the LysoTracker Red Kit (Beyotime, C1046). Imaginal discs dissected from third-instar larvae were collected in PBS and incubated with LysoTracker Red (1:3,000) for 15 min at 37 °C, washed with PBS three times prior to imaging. Primary antibodies included: rabbit anti-FoxO3A (1:200; Abcam, ab12162), mouse anti–Myc-tag (1:350, CST, 9B11), rabbit anti-FLAG antibody (1:200, Sigma). Secondary antibodies were anti-mouse CY3 (1:1,000) and anti-rabbit Alexa Flour 488 (1:500). All images were collected with a confocal microscope (Zeiss LSM 780).
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