Cells were transiently transfected with pAcGFP1-Mito (Takara Bio) and pcDNA of mito-GFP-PPX using the Effectene reagent (Qiagen, Chatsworth, CA, USA) under serum-free conditions for 6 h in 4-chamber microscopic slides (PEZGS0416, Millipore). 48 and 72 h post-transfection, the cells were fixed with 4% paraformaldehyde followed by DAPI staining. The coverslips were mounted on slides using mounting media. Images of ten microscopic fields per well were obtained using a fluorescence microscope. Ten microscopic images under 40× magnification were obtained for each condition using a fluorescence microscope. The average percentages of GFP positive cells were calculated against the total number of DAPI stained cells in each microscopic image. The fold change in the percentage of GFP positive cells at 72 h was plotted against 42 h.
Pezgs0416
Manufactured by Merck Group
Sourced in United States
PEZGS0416 is a laboratory equipment product from Merck Group. It is a versatile device designed for use in various scientific and research applications. The core function of PEZGS0416 is to perform precise measurements and analyses within a controlled laboratory environment.
Automatically generated - may contain errors
Lab products found in correlation
Alizarin red s, by Thermo Fisher Scientific (2 mentions)
Spectramax id3, by Molecular Devices (2 mentions)
24 well plate, by NEST Biotechnology (2 mentions)
Click it tunel alexa flour 594, by Thermo Fisher Scientific (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (2 mentions)
Lsm 710, by Zeiss (2 mentions)
Effectene reagent, by Qiagen (1 mentions)
Ab9361, by Abcam (1 mentions)
Ab209484, by Abcam (1 mentions)
Ab15580, by Thermo Fisher Scientific (1 mentions)
Ab185230, by Abcam (1 mentions)
A22287, by Thermo Fisher Scientific (1 mentions)
Ab93876, by Abcam (1 mentions)
Alexa fluor 568, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Cyclic amp, by Merck Group (1 mentions)
Brain derived neurotrophic factor (bdnf), by Thermo Fisher Scientific (1 mentions)
Ascorbic acid, by Merck Group (1 mentions)
Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (1 mentions)
Carboxyfluorescein succinimidyl ester (cfse), by Merck Group (1 mentions)
21 protocols using pezgs0416
1
Mitochondrial Localization Transfection Assay
2
Dental Mesenchyme Culture and Odontogenic Differentiation
3
Immunofluorescence Staining of Bone Cells
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The decalcified samples were dehydrated in 15% sucrose/PBS solution for 2 h, 30% sucrose/PBS for 2 h, and 30% sucrose/OCT (4,583, Sakura, Torrance, CA, United States) overnight at 4°C and then embedded in OCT. Cryosections measuring 8 μm in thickness were immunofluorescence-stained following standard protocols. The primary antibodies included Runx2 (1:100, #12556, Cell Signaling Technology, Danvers, MA, United States), Osterix (1:100, ab209484, Abcam, Cambridge, United Kingdom), β-galactosidase (β-gal; 1:200, ab9361, Abcam), osteocalcin (Ocn; 1:100, ab93876, Abcam), and Sox9 (1:100, ab185230, Abcam). Alexa Fluor 488 and Alexa Fluor 568 (1:200, Invitrogen, Waltham, MA, United States) were used as secondary antibodies. DAPI (62248, Invitrogen) was used for counterstaining. ImageJ was used to analyzed the ratio of Osterix+/tdTomato + cells to Osterix + cells.
Cell samples were plated in a 4-well chamber slide (PEZGS0416, Millipore). The slides were washed with PBS, immediately fixed with 4% paraformaldehyde for 15 min, and blocked with goat serum for 30 min at room temperature. The cells were incubated with the primary antibody (Ki67, 1:200, ab15580) at 4°C overnight, then incubated with secondary antibodies at room temperature for 1 h, stained with phalloidin for 20 min (1:50, A22287, ThermoFisher Scientific), and counterstained with DAPI.
Cell samples were plated in a 4-well chamber slide (PEZGS0416, Millipore). The slides were washed with PBS, immediately fixed with 4% paraformaldehyde for 15 min, and blocked with goat serum for 30 min at room temperature. The cells were incubated with the primary antibody (Ki67, 1:200, ab15580) at 4°C overnight, then incubated with secondary antibodies at room temperature for 1 h, stained with phalloidin for 20 min (1:50, A22287, ThermoFisher Scientific), and counterstained with DAPI.
4
Neural Differentiation of NPCs
5
Dental Mesenchyme Culture and Odontogenic Differentiation
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The dental mesenchyme was obtained from 1-month old Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl mice 5 days after induction as described above. The pulp tissue was cut into pieces and incubated with growth media in 6-well plate. The culture media was left to rest for 4 days to allow tissue adhesion and cell migration. Then the culture media was changed every other day.
2 × 105 MSCs were plated in a 4-well chamber slide (PEZGS0416, Millipore) or 24-well plate (Nest, 702011) and induced in odontogenic differentiation media containing 1% FBS, β-glycerophosphate (β-GP) (5mM), ascorbic acid (50μg/ml), and dexamethasone (DEX) (10nM). 100 ng/ml mouse recombinant Igf2 was added to odontogenic induce media according different groups (showed inFigures 6F –6I ). For Dspp RNAscope in situ hybridization, cells cultured in a 4-well chamber slide were harvested 7 days after induction and the standard protocols of the manufacturer were then followed. To detect mineralized nodules, cells were induced for 21 days, fixed with 4% paraformaldehyde, and stained with 2% Alizarin red S (pH4.2) (ACROS Organics, 400480250). Alizarin red-S contents were extracted with 10% cetylpyridinium chloride in distilled water and measured at 590 nm on SpectraMax iD3 (Molecular Devices, LLC., San Jose, USA).
2 × 105 MSCs were plated in a 4-well chamber slide (PEZGS0416, Millipore) or 24-well plate (Nest, 702011) and induced in odontogenic differentiation media containing 1% FBS, β-glycerophosphate (β-GP) (5mM), ascorbic acid (50μg/ml), and dexamethasone (DEX) (10nM). 100 ng/ml mouse recombinant Igf2 was added to odontogenic induce media according different groups (showed in
6
Quantifying Apoptosis in Retinal Cells
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To determine the percentage of apoptotic cells, retinal PC were plated at 1 × 104 cells per well of a four‐chamber slide (Cat#: PEZGS0416; Millipore). The next day, medium was replaced with medium containing 10 μmol/L 1,25(OH)2D3 or solvent ethanol‐treated, and cells were incubated for 48 hours. TdT‐dUTP terminal nick‐end labeling (TUNEL) staining was performed using Click‐IT TUNEL Alexa Flour 594, as recommended by the supplier (Cat#: C10246; Life Technologies). Positive cells were counted per five high power field (×200) using a fluorescence microscope and reported as a percentage of apoptotic cells relative to total number of cells per field. DNase I treatment was used as a positive control as recommended by the supplier.
7
Cytotoxicity and Proliferation Assays
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Cells were seeded at a density of 3 × 103 cells/well in 96-well plates. After overnight incubation, fresh medium containing different doses of chemotherapeutic agents was added, and the plates were maintained for 0, 24, 48, or 72 h. The 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) reagent (cat. #G3581, Promega) was used for cell viability assays. Cell proliferation was measured using the 5-ethynyl-2′-deoxyuridine (EdU) assay. Briefly, cells were seeded at a density of 1 × 104 cells in chamber slides (cat. #PEZGS0416, Millipore) and cultured overnight. Cells were then treated with 10 μM 5-Fu for 24 h. Next, 10 μM EdU (cat. #C10339, Thermo Fisher Scientific) was added to the culture medium, which was incubated for an additional 12 or 24 h. Cells were fixed and stained following the manufacturer’s instructions.
8
Phagocytosis of Apoptotic Tumor Cells
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B16F10 cells were labeled with CFSE as previously described [50 (link)]. Briefly, detached B16F10 tumor cells were resuspended in 1 mL DMEM, stained with 1 μL of 1 μM CFSE (Invitrogen, Cat#34554), and washed twice. Tumor cells were induced to undergo apoptosis through heat-treatment with incubation at 42 °C for 2 h followed by incubation at 37 °C on a shaker at 30 rpm for 24 h. Cell death was confirmed by almost 100% trypan blue staining. The suspension was centrifuged at 1000 rpm × 5 min to remove cell debris. CFSE labeled B16F10 cells were cocultured with the indicated LECs at a 1:1 ratio in 4-well slides (Millipore, Cat#PEZGS0416; Burlington, MA, USA) for 24 h before downstream analysis.
9
Immunofluorescent Staining of FXR
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Cells were seeded in 4-well chamber slides (#PEZGS0416, Millipore, USA), washed twice with cold PBS, fixed with 500 μl 4% paraformaldehyde for 35 min, permeabilized with 200 μl 1% Triton X-100 for 10 min, and blocked with 500 μl goat serum for 30 min. Next, the cells were incubated with primary antibodies at 4 °C overnight. Subsequently, the cells were incubated with FITC-conjugated goat anti-mouse IgG antibody (#12–506, 1:200, Millipore, USA) at room temperature for 2 h. Finally, the nuclei were stained with DAPI (#C0060, 1:800, Solarbio, China) for 10 min. The primary antibody used was a rabbit anti-human FXR antibody (#187735, 1:100, Abcam, UK).
10
Quantifying Retinal Cell Apoptosis
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