Galaxy 170r co2 incubator

Manufactured by Eppendorf

Sourced in Germany

The Galaxy® 170R CO2 Incubator is a laboratory equipment used for maintaining controlled environmental conditions to support the growth and development of cell cultures. It features a temperature and CO2 concentration regulation system to provide an optimal growth environment.

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9 protocols using galaxy 170r co2 incubator

Caco-2 and CD4+ T Cell Culture Protocol

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Caco-2 cells originally obtained from the American Type Culture Collection (ATCC, Manassas, VA) were grown in 100 x 17mm Nunclon™ Delta dishes (ThermoFisher Scientific, ca#150350) with Gibco® DMEM media (ThermoFisher Scientific, Grand Island, NY, USA, ca#11965-092) with 10% fetal bovine serum (Sigma-Aldrich, St. Louis, MO, USA, ca#F2442500ML) and 1% antibiotic-antimycotic solution (Corning Cellgro™, ThermoFisher Scientific, ca#30-004-CI) at 37 °C under 5% CO₂, in a Galaxy 170R CO₂ incubator (Eppendorf, Enfield, CT, USA).
CD4⁺ T cells enriched from human T lymphoblast CCL-119 cells, originally purchased from ATCC (>75% CD4⁺ determined by flow cytometry) were grown in T75 CELLSTAR TC Flasks (Phenix Research, Cander, NC, ca# TCG-658175) with RPMI 1640 media (ATCC, ca#30-2001) containing 10% fetal bovine serum and 1% antibiotic-antimycotic solution. After the cells were ~50% confluent, they were grown in phenol red free in RPMI with charcoalstripped fetal bovine serum from two days before treatments. Cells were treated with BPA (Sigma-Aldrich, ca#239658-50G) in DMSO, E₂ (Sigma-Aldrich, ca#E8875), VD₃ (1α, 25-dihydroxy Vitamin D3, EMD-Millipore, Burlington, MA, USA, ca#679101) in DMSO or their combination for 72 hrs. The variation of the final concentrations was indicated in the Results section. The stock solutions of BPA, E₂, and VD₃ were all 0.1 mM.

Sowers M.L., Tang H., Tian B., Goldblum R., Midoro-Horiuti T, & Zhang K. (2019). Bisphenol A Activates an Innate Viral Immune Response Pathway. Journal of proteome research, 19(2), 644-654.

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Circadian Rhythm Monitoring in U2OS Cells

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Bmal1:luc U2OS and Per2:luc U2OS cells were a gift from Dr. Andrew Liu, University of Memphis (Liu et al., 2008 (link)). U2OS cells were cultured in standard conditions. For bioluminescence recordings, U2OS cells were synchronized by changing medium to “Air Medium” (Hastings et al., 2005 (link)). Bioluminescence assays were performed at 37°C using 12-well and 96-well plates in custom-made bioluminescence recording systems (Cairn Research Ltd) composed of a charge-coupled device (CCD) camera (Andor iKon-M 934) mounted on the top of an Eppendorf Galaxy 170R CO₂ incubator. Bioluminescence data traces were analyzed using a modified version of the R script “CellulaRhythm” (Hirota et al., 2008 (link)).

Rey G., Valekunja U.K., Feeney K.A., Wulund L., Milev N.B., Stangherlin A., Ansel-Bollepalli L., Velagapudi V., O’Neill J.S, & Reddy A.B. (2016). The Pentose Phosphate Pathway Regulates the Circadian Clock. Cell Metabolism, 24(3), 462-473.

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Cytotoxicity Evaluation of GS and GS-TC in MCF-7 Cells

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The breast cancer cell line MCF-7 was used for the cytotoxicity study of pure GS and GS-TC. The cell line was grown into a mammalian culture medium with a supply of supplements (10% fetal bovine serum, penicillin 100 U/mL, streptomycin 10 U/mL) and 5% CO₂ at 37 ± 0.5 °C in a CO₂ incubator (Galaxy^® 170R CO₂ incubator, Eppendorf, Germany). GS and GS-TC were dissolved in DMSO and further diluted between 10–2000 µM with phosphate buffer. The pure GS and GS TC were added into a seeded well of the microplate (96-well plate) and further incubated for 72 h. The cell viability was determined by adding 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) into each microplate of cells. Then, the formazan crystal was fully dissolved by adding 100 µL of DMSO and absorbance was analyzed at 490 nm using a microplate reader. The % inhibition was calculated by following the formula against control.

Growth inhibition = \frac{OD control sample - OD test sample}{OD control sample} \times 100

Zafar A., Alruwaili N.K., Imam S.S., Alsaidan O.A., Alkholifi F.K., Alharbi K.S., Mostafa E.M., Alanazi A.S., Gilani S.J., Musa A., Alshehri S., Rawaf A, & Alquraini A. (2021). Formulation of Genistein-HP β Cyclodextrin-Poloxamer 188 Ternary Inclusion Complex: Solubility to Cytotoxicity Assessment. Pharmaceutics, 13(12), 1997.

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Embryo Development and Quality Evaluation

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The activated oocytes were washed three times in PBS (gibco, 21600-051-10x1L) and then cultured in potassium simplex optimization medium supplemented by 10% fetal bovine serum (Sigma, A2153-50G) at 37 o C under 5% CO2 (New Brunswick, Galaxy 170R CO2 incubator, Eppendorf) up to 5 days. The development rate of the embryo was calculated at each stage (2-cell, 4-cell, morula, blastocyst, and hatched blastocyst), by dividing the number of embryos at each stage by total activated oocytes in each treatment group. Embryo quality was calculated based on the amount of blastocyst, inner cell mass (ICM) and trophectoderm cells. Total ICM and trophectoderm cells were determined by exposed blastocyst to rabbit anti-mouse serum antibody (Sigma, M5774-2ML) for 1-hr and complement sera from guinea pig (Sigma, S1639-5ML) for 30 minutes, and followed by staining using Hoechst 34580 (Sigma, 63493-5MG) -Propidium iodide (Sigma, 81845-25MG). ICM cells located on the inside of the embryo were blue, whereas trophectoderm cells on the outside were red (Hine et al., 2008) .

Hine T.M., Nalley W.M., Uly K., Manu A., Ly J.Q., Marawali A., & Kune P. (2021). EFFECT OF OOCYTE AGE AND ACTIVATION AGENTS ON IN VITRO DEVELOPMENT OF MOUSE PARTHENOGENETIC EMBRYOS. The Journal of Animal and Plant Sciences, 31(5).

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ROS Estimation using DCFH-DA Probe

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The cell-permeable fluorogenic probe DCFH-DA (2′7′-dichloro-dihydro-fluorescein diacetate) was used to estimate ROS level [22 (link)]. In brief, A549 cells were seeded at a density of 1 × 10⁴ cells/well and CCRF-CEM cells 1 × 10⁵ cells/well in 96-well plate fluorescence. Cells were then incubated with 5 μM DCFH-DA for 45 min in a humidified 5% CO₂ atmosphere at 37 °C (New Brunswick Galaxy^® 170R CO₂ Incubator, Hamburg, Germany), washed in HBSS, and then treated with the indicated concentrations of HAL, PLEC and MRC for 1, 2, 12, 24 and 48 h. Fluorescence readings were taken at 485/20 nm excitation and 528/20 nm emission in a fluorescence plate reader (Bio-Tek Synergy HT Microplate Reader).

Sitarek P., Kowalczyk T., Synowiec E., Merecz-Sadowska A., Bangay G., Princiotto S., Śliwiński T, & Rijo P. (2022). An Evaluation of the Novel Biological Properties of Diterpenes Isolated from Plectranthus ornatus Codd. In Vitro and In Silico. Cells, 11(20), 3243.

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Cell Culture of MCF7 and FaDu Cells

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The MCF7 (human breast; mammary gland; adenocarcinoma; HTB-22™) and FaDu (human pharynx squamous cell carcinoma; HTB-43™) cell lines were obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA). The two cell models were cultured in a humidified 5% CO₂ atmosphere at 37 °C (New Brunswick Galaxy^®® 170R CO₂ Incubator). Monolayer cells were grown in Eagle’s Minimum Essential Medium (EMEM; Sigma-Aldrich) supplemented with 10% fetal bovine serum (Biowest), recommended by a manufacturer. To the cell culture medium was added penicillin (100 U/mL) and streptomycin (100 μg/mL). Then, at 80–85% confluence, the cells were detached and resuspended using TrypLE™ Express Enzyme without phenol red (Gibco™).

Sitarek P., Synowiec E., Kowalczyk T., Bangay G., Śliwiński T., Picot L., Princiotto S, & Rijo P. (2022). Anticancer Properties of Plectranthus ornatus-Derived Phytochemicals Inducing Apoptosis via Mitochondrial Pathway. International Journal of Molecular Sciences, 23(19), 11653.

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Magnetic Field Exposure System for Cell Culture

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The EMF device consists of magnetic field applicator and a dedicated precision electric signal generator (COMEF, Poland). The cylindrical in shape magnetic field applicator (length of 24.5 cm and a diameter of 9.5/14 cm (inside/outside)) is based on low inductance coreless solenoid with the length of 19.9 cm and a diameter of 10.5 cm. The number of turns per coil is 280, the resistance of the coil is 3.6 Ω and inductance is 2.5 mH. The whole coil is within an amagnetic Teflon tube to keep it clean and sterile. This system enables to produce magnetic field frequency from 0.01 Hz to 1 kHz and magnetic flux density up to 2 mT (amplitude). The magnetic field applicator is located in the cell culture incubator (New Brunswick Galaxy^® 170R CO₂ Incubator), where atmosphere composition temperature and humidity regulation were provided and continuously controlled.

Łach K., Cebulski J., Chaber R., Kocan B., Wojnarowska-Nowak R, & Banaś-Ząbczyk A. (2021). First Identification of the Effects of Low Frequency Electromagnetic Field on the Micromolecular Changes in Adipose Tissue-Derived Mesenchymal Stem Cells by Fourier Transform Infrared Spectroscopy. Journal of Medical Physics, 46(4), 253-262.

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Culturing Human Adipose-Derived Stem Cells

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Commercially available StemPRO® Human Adipose-Derived Stem Cells, lot no 1001001, Invitrogen™) were cultured in reduced serum (2%), human mesenchymal stem cell growth-supporting medium (MesenPRO RSTM Medium, Gibco™), recommended by a manufacturer. To the cell culture medium was added L-glutamine (2 mM; Gibco™) and an antibiotic and antimycotic mixed solution (100 U/ml penicillin, 0.1 mg/ml streptomycin, and 0.25 μg/ml amphotericin B; Gibco™). The cells were cultured at 37°C in a humidified atmosphere in the presence of 5% CO₂ (New Brunswick Galaxy® 170R CO2 Incubator). Then, at 80% confluence, the cells were trypsinized and resuspended using TrypLE™ Express Enzyme without phenol red (Gibco™).

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Culturing Human Umbilical Vein Endothelial Cells

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All experiments were performed on human umbilical vein endothelial cells (HUVECs). These cells were purchased from Gibco (Cascade Biologics®, catalog number C0035C) and cultured in Medium 200 (Gibco, catalog number M-200-500) supplemented with Low Serum G Growth Supplement Kit (LSGS Kit; Gibco, catalog number S003K) at 37°C and 5% CO₂ in an incubator (Galaxy® 170 R-CO₂ Incubator, New Brunswick Scientific) under a humidified atmosphere.

Kowalczyk T., Sitarek P., Skała E., Rijo P., Andrade J.M., Synowiec E., Szemraj J., Krajewska U, & Śliwiński T. (2019). An Evaluation of the DNA-Protective Effects of Extracts from Menyanthes trifoliata L. Plants Derived from In Vitro Culture Associated with Redox Balance and Other Biological Activities. Oxidative Medicine and Cellular Longevity, 2019, 9165784.

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