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Bio one 655090

Manufactured by Greiner
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The Bio-one 655090 is a laboratory equipment designed for general use in scientific and research settings. It provides a core function of sample collection and storage.

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Lab products found in correlation

Celigo s image cytometer, by Revvity (2 mentions) Lionheart fx, by Agilent Technologies (2 mentions) Celltiter blue reagent, by Promega (1 mentions) Cisplatin, by Cayman Chemical (1 mentions) Calcusyn software, by Biosoft (1 mentions) Imagexpress micro xls widefield microscope, by Molecular Devices (1 mentions) Fluorescein in situ cell death detection kit, by Roche (1 mentions) Apo one homogeneous caspase 3 7 assay, by Promega (1 mentions) Spectramax gemini xps, by Molecular Devices (1 mentions) Ixmicro microscope, by Molecular Devices (1 mentions)

Close spelling variants of this product

Bio-one (32 citations)

9 protocols using bio one 655090

1

Immunofluorescence Analysis of TFEB and TFE3 in MHV-A59 Infected HeLa mCC1a Cells

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HeLa mCC1a cells were seeded in 96-well plates (Greiner Bio-One 655,090) at a density of 60,000 cells/cm2. Cells were incubated for 2 h with MHV-A59, washed, and treated with drugs until 16 h p.i. Subsequently, cells were fixed with 4% paraformaldehyde for 20 min, permeabilized with TX-100 1% for 30 min and blocked with 3% fish gelatin for 1 h at room temperature. The cells were incubated with anti-TFEB or anti-TFE3 1:150 over night at 4°C and with Alexa Fluor 488 secondary antibodies 1:1000 for 1 h at room temperature. To stain the membrane and nucleus, HCS CellMask Deep Red Stain (1:50,000) and Hoechst 33,342 20 mM (1:5000) were used for 15 min at room temperature, respectively. Images were acquired with Cell_Voyager (CV7000) in confocal mode using 405/488/640 nm filters, 20× objective (N.A. = 0.75), and laser-based autofocus. 16 images (20× magnification) were acquired per well and then analyzed with Columbus 2.9.1.532 software. Cells were segmented by cell nucleus (blue color) and the intensity of green fluorescence was measured to identify the cells with nuclear TFEB. The script was designed using Torin 1 (0.3 μM) as a positive control and without anti-TFEB or anti-TFE3 as a negative control.
Contreras P.S., Tapia P.J., Jeong E., Ghosh S., Altan-Bonnet N, & Puertollano R. (2023). Beta-coronaviruses exploit cellular stress responses by modulating TFEB and TFE3 activity. iScience, 26(3), 106169.
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2

Cytotoxicity Assay for Anti-Cancer Drugs

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Cells (3 × 103) were plated in each well of a 96-well plate (Greiner bio-one 655090) with increasing concentrations of cisplatin (Cayman Chemical, 13119) dissolved in DMSO. DMSO concentrations were identical for all conditions. Each condition of cisplatin concentrations was performed in triplicate. After 48 or 72 hours, plates were read using a Cell Titer Blue reagent (Promega, G8081). IC50 calculations were obtained with Calcusyn software (Biosoft). Gemcitabine (Caymen Chemical, 11690) and carboplatin (TCI, C2043) experiments were done similarly. For experiments involving exosomes, 350 million exosomes were used per reaction. For assays with inhibitors, 1 µM of MK-2206 or 5 µM of LiCl was utilized based on Meng et al.50 (link) and Coghlan et al.51 (link), respectively.
Phelps C.A., Lindsey-Boltz L., Sancar A, & Mu D. (2019). Mechanistic Study of TTF-1 Modulation of Cellular Sensitivity to Cisplatin. Scientific Reports, 9, 7990.
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3

Cell Growth Quantification via Imaging

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Depending on the experiment, between 2,500 and 10,000 cells per well
were plated in flat-bottom, optically pure 96 well plates (Greiner bio-one
655090). Cells were allowed to attach for 12–24 hours, then treatment was
applied. Plates were serially imaged during the experiment (beginning
immediately after treatment) using a Celigo S Image Cytometer (Nexcelom
Bioscience) and Celigo image analysis software was used to count cells. For each
experiment, the same image analysis algorithm was applied to all wells for all
time points.
Bader D.A., Hartig S.M., Putluri V., Foley C., Hamilton M.P., Smith E.A., Saha P.K., Panigrahi A., Walker C., Zong L., Martini-Stoica H., Chen R., Rajapakshe K., Coarfa C., Sreekumar A., Mitsiades N., Bankson J.A., Ittmann M.M., O’Malley B.W., Putluri N, & McGuire S.E. (2018). Mitochondrial pyruvate import is a metabolic vulnerability in androgen receptor-driven prostate cancer. Nature metabolism, 1(1), 70-85.
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4

Cell Growth Quantification via Imaging

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Depending on the experiment, between 2,500 and 10,000 cells per well
were plated in flat-bottom, optically pure 96 well plates (Greiner bio-one
655090). Cells were allowed to attach for 12–24 hours, then treatment was
applied. Plates were serially imaged during the experiment (beginning
immediately after treatment) using a Celigo S Image Cytometer (Nexcelom
Bioscience) and Celigo image analysis software was used to count cells. For each
experiment, the same image analysis algorithm was applied to all wells for all
time points.
Bader D.A., Hartig S.M., Putluri V., Foley C., Hamilton M.P., Smith E.A., Saha P.K., Panigrahi A., Walker C., Zong L., Martini-Stoica H., Chen R., Rajapakshe K., Coarfa C., Sreekumar A., Mitsiades N., Bankson J.A., Ittmann M.M., O’Malley B.W., Putluri N, & McGuire S.E. (2018). Mitochondrial pyruvate import is a metabolic vulnerability in androgen receptor-driven prostate cancer. Nature metabolism, 1(1), 70-85.
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5

Live-cell Imaging of Subconfluent Cells

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Cells were plated at least 24 hours prior to imaging in phenol red-free full-growth media in a 96-well plate (Greiner bio-one #655090) such that the density would remain subconfluent until the end of the imaging period. Images were acquired every 12 minutes on an ImageXpress Micro XLS widefield microscope (Molecular Devices) with a 10X 0.45NA objective; CFP exposure = 75 ms; YFP exposure = 200 ms. Cells were imaged in a humidified, 37°C chamber at 5% CO2.
Gookin S., Min M., Phadke H., Chung M., Moser J., Miller I., Carter D, & Spencer S.L. (2017). A map of protein dynamics during cell-cycle progression and cell-cycle exit. PLoS Biology, 15(9), e2003268.
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6

TUNEL Assay for Apoptosis Quantification

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Fluorescein In Situ Cell Death Detection Kit (Roche Diagnostics 11684795910) was used for the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. HLMVECs were seeded onto a sterile 96 well microplate (Greiner Bio-One 655090) and treated with CFH or vehicle control. Cells were fixed with 4% paraformaldehyde for ten minutes at room temperature, rinsed twice with PBS, stained with TUNEL reaction mixture for at least one hour at 37°C, rinsed three times with PBS, labeled with Hoechst (100 ng/mL, 35 minutes), and left in PBS for imaging. Images were captured using Lionheart FX (BioTek) at 20X magnification in DAPI (377,447) and GFP (469, 525) channels. Images were analyzed using Gen5 Image+ software version 3.10. Image preprocessing was applied using Auto Background Flattening of DAPI channel. Cellular analysis to count total number of cells included those between 5–30 μm meeting fluorescent threshold of 5,000 (DAPI); cells meeting fluorescence threshold (GFP>5,000) were counted as TUNEL+.
Tomasek T., Ware L.B., Bastarache J.A, & Meegan J.E. (2021). Cell-free hemoglobin-mediated human lung microvascular endothelial barrier dysfunction is not mediated by cell death. Biochemical and biophysical research communications, 556, 199-206.
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7

Quantitative Analysis of Cell Viability

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HLMVECs were seeded on a sterile 96 well microplate (Greiner Bio-One 655090) and treated with CFH or vehicle control. Cells were exposed to the Draq7 (Abcam ab109202) stain at 1.5 μM diluted in complete media for 10 minutes, then Hoechst (100 ng/mL, 35 minutes) Live cells were imaged using a Lionheart FX (BioTek) at 4X magnification in DAPI (377,447) and CY5 (628, 685) channels. Images were analyzed using Gen5 Image+ software version 3.10. Montage images were stitched using Linear Blend of DAPI channel. Image preprocessing was applied using Auto Background Flattening of DAPI channel. Cellular analysis to count total number of cells included those between 5–50 μm meeting fluorescent threshold of 5,000 (DAPI); cells meeting fluorescence threshold (CY5>10,000) were counted as Draq7+.
Tomasek T., Ware L.B., Bastarache J.A, & Meegan J.E. (2021). Cell-free hemoglobin-mediated human lung microvascular endothelial barrier dysfunction is not mediated by cell death. Biochemical and biophysical research communications, 556, 199-206.
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8

Caspase-3/7 Activity Assay in ALDH1high Cells

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Caspase-3/7 activities were assayed using the Apo-ONE™ Homogeneous Caspase-3/7 assay (Promega G7790) according to the manufacturer's instructions. ALDH1high cells (1 × 103/well) were seeded into black 96-well fluorescence culture plates (Greiner bio-one 655090) and incubated for 24 h, after which they were treated with TLSC702 for an additional 5 days. Equal volumes of DMEM and Apo-ONE™ caspase reagent (1:100 profluorescent substrate and lysis buffer) were then added to cells, and the mixture was incubated for 30 min. Fluorescence (excitation, 485 nm; emission, 512 nm) was measured using a fluorescence plate reader (SPECTRA max GEMINI XPS [Molecular Devices]). Background fluorescence was determined as the fluorescence from DMEM alone and deducted from all experimental values.
Tamori S., Nozaki Y., Motomura H., Nakane H., Katayama R., Onaga C., Kikuchi E., Shimada N., Suzuki Y., Noike M., Hara Y., Sato K., Sato T., Yamamoto K., Hanawa T., Imai M., Abe R., Yoshimori A., Takasawa R., Tanuma S.I, & Akimoto K. (2018). Glyoxalase 1 gene is highly expressed in basal-like human breast cancers and contributes to survival of ALDH1-positive breast cancer stem cells. Oncotarget, 9(92), 36515-36529.
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9

Time-lapse Imaging of Drug-treated Cells

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Cells were plated at least 24 hr prior to imaging in phenol red-free full-growth media in a 96-well plate (Greiner bio-one #655090) such that the density would remain sub-confluent until the end of the imaging period. For experiments that required drug addition, cells were first “pre-imaged” for ~12 hr in full growth media without any drugs. At the time of drug addition, 50% of the media in the wells was replaced with media containing a 2X drug concentration and cells were imaged for additional 48 hr in presence of the drug. Images were acquired every 12 min on an IXMicro microscope (Molecular Devices) with a 10X 0.45NA objective. Total light exposure time was kept under 300 ms for each time point. Cells were imaged in a humidified, 37°C chamber at 5% CO2. Cell tracking was done using custom MATLAB scripts, as in (Cappell et al., 2016 (link)). See Supplementary Methods for details.
Arora M., Moser J., Phadke H., Basha A.A, & Spencer S.L. (2017). Endogenous replication stress in mother cells leads to quiescence of daughter cells. Cell reports, 19(7), 1351-1364.
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