Andpenicillin streptomycin, by Thermo Fisher Scientific - Product details

1

Isolation and Culture of Sensory Neurons

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DRGs were dissected from adult TrpV1-Cre::tdTomato mice
(12–13 weeks) into Hank’s balanced salt solution (HBSS) (Life
Technologies). DRG were dissociated in 1 mg ml⁻¹ collagenase A plus 2.4
U ml⁻¹ dispase II (enzymes, Roche Applied Sciences) in HEPES-buffered
saline (Sigma) for 90 min at 37 °C and then triturated down to single cell level
using glass Pasteur pipettes of decreasing size. DRGs were the centrifuged over a
10% BSA gradient and plated on laminin-coated cell culture dishes (Sigma). DRGs
were cultured 24 hours in B27-supplemented neurobasal-A medium plus 50 ng/ml NGF
(Invitrogen), 2 ng/ml GDNF (Sigma), 10uM arabinocytidine (Sigma) and
penicillin/streptomycin (Life Technologies).

Wainger B.J., Buttermore E.D., Oliveira J.T., Mellin C., Lee S., Saber W.A., Wang A., Ichida J.K., Chiu I.M., Barrett L., Huebner E.A., Bilgin C., Tsujimoto N., Brenneis C., Kapur K., Rubin L.L., Eggan K, & Woolf C.J. (2014). Modeling pain in vitro using nociceptor neurons reprogrammed from fibroblasts. Nature neuroscience, 18(1), 17-24.

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Isolation and Culture of Sensory Neurons

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DRGs were dissected from adult TrpV1-Cre::tdTomato mice
(12–13 weeks) into Hank’s balanced salt solution (HBSS) (Life
Technologies). DRG were dissociated in 1 mg ml⁻¹ collagenase A plus 2.4
U ml⁻¹ dispase II (enzymes, Roche Applied Sciences) in HEPES-buffered
saline (Sigma) for 90 min at 37 °C and then triturated down to single cell level
using glass Pasteur pipettes of decreasing size. DRGs were the centrifuged over a
10% BSA gradient and plated on laminin-coated cell culture dishes (Sigma). DRGs
were cultured 24 hours in B27-supplemented neurobasal-A medium plus 50 ng/ml NGF
(Invitrogen), 2 ng/ml GDNF (Sigma), 10uM arabinocytidine (Sigma) and
penicillin/streptomycin (Life Technologies).

Wainger B.J., Buttermore E.D., Oliveira J.T., Mellin C., Lee S., Saber W.A., Wang A., Ichida J.K., Chiu I.M., Barrett L., Huebner E.A., Bilgin C., Tsujimoto N., Brenneis C., Kapur K., Rubin L.L., Eggan K, & Woolf C.J. (2014). Modeling pain in vitro using nociceptor neurons reprogrammed from fibroblasts. Nature neuroscience, 18(1), 17-24.

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3

Cell Culture Media Optimization

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All non-adherent cells were cultured in RPMI 1640 GlutaMAX, HEPES medium (Life
Technologies) supplemented with 10 % Hyclone FBS (Thermo Scientific Hyclone),
MEM non-essential amino acids solution (Life Technologies), and
penicillin/streptomycin (100 U ml⁻¹/100 µg
ml⁻¹) solution (Life Technologies). Adherent cells were cultured
in DMEM GlutaMAX medium (Life Technologies) supplemented with 10 % FetalClone
III Serum (Thermo Scientific Hyclone), MEM non-essential amino acids solution, and
100 U penicillin ml⁻¹ plus
100 µg streptomycin ml⁻¹ solution.

Kobayashi Y., Gélinas C, & Dougherty J.P. (2017). Histone deacetylase inhibitors containing a benzamide functional group and a pyridyl cap are preferentially effective human immunodeficiency virus-1 latency-reversing agents in primary resting CD4+ T cells. The Journal of General Virology, 98(4), 799-809.

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4

Colon Cancer Cell Lines and Knockdown

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Human colon cancer cell lines [HT-29 (ATTC HTB-38) and HCT-116 (ATTC
CCL-247)] as well as normal colon cell line CCD-18Co (ATCC, CRL-1459) were
obtained from the American Type Culture Collection (ATCC), Manassas, VA. Cells
were grown in DMEM medium (HyClone GE Healthcare, SH30022.01) supplemented with
10% fetal bovine serum (FBS; Thermo Scientific, 10082147) and
penicillin/streptomycin (Thermo Scientific, 15140122) at 5% CO2 and 37°C.
GLTP overexpression and depletion were confirmed by western blot analyses (Supplemental Fig. S1).
TurboGFP-GIPZ lentiviral shRNAmir constructs for GLTP, RIPK-3, MLKL and
non-targeting (scrambled) control shRNAs were supplied by the University of
Minnesota Genomics Center (http://genomics.umn.edu/rna-interference.php).

Mishra S.K., Stephenson D.J., Chalfant C.E, & Brown R.E. (2018). Upregulation of Human Glycolipid Transfer Protein (GLTP) Induces Necroptosis in Colon Carcinoma Cells. Biochimica et biophysica acta. Molecular and cell biology of lipids, 1864(2), 158-167.

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5

EGF Stimulation of A431 Cells

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A431 cells were grown in Dulbecco’s modified Eagle’s medium (DMEM)
(A.T.C.C., VA) supplemented with 10% FBS (Atlas Biologicals, CO), and
penicillin/streptomycin (ThermoFisher, MA). Trypsin/EDTA was
obtained from Gibco (ThermoFisher, MA). Unless otherwise noted, chemicals were
obtained from Sigma Chemical Co. (Sigma, MO). Recombinant EGF and c-Fos were
obtained from Active Motif (Active Motif, CA). Recombinant Ras was obtained from
Cytoskeleton, Inc. (Cytoskeleton, CO). Human EGF was obtained from Cytoskeleton,
Inc. (Cytoskeleton, CO). For EGF stimulation experiments, A431 cells were serum
restricted for 24 h with serum-free DMEM in order to synchronize the cells. The
cells were then treated with 33 ng/ml EGF for 0.5, 2, 5, 15, and 60 min
in individual 15-cm dishes (Corning, NY), followed by subsequent lysis with
BlastR lysis buffer (Cytoskeleton, CO).

Horita H., Law A., Hong S, & Middleton K. (2017). A simple toolset to identify endogenous post-translational modifications for a target protein: a snapshot of the EGFR signaling pathway. Bioscience Reports, 37(4), BSR20170919.

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6

Purification and Culture of IL-2Rα+ YT-1 NK Cells

Cited 1 time

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Unmodified YT-1⁵³ and
IL-2Rα⁺ YT-1 human NK cells⁵⁴ were cultured in RPMI complete medium
(RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM L-glutamine,
minimum non-essential amino acids, sodium pyruvate, 25 mM HEPES, and
penicillin-streptomycin [Gibco]). CTLL-2 cells purchased from ATCC were cultured
in RPMI complete with 10% T-STIM culture supplement with ConA (Corning). 24
hours prior to signaling studies, CTLL-2 cells were resuspended in RPMI lacking
T-STIM culture supplement for IL-2 starvation. Viability (>95%) of CTLL-2
cells was verified by trypan blue exclusion (counted in a hemocytometer)
immediately before performing the signaling assays. All cells were maintained at
37°C in a humidified atmosphere with 5% CO₂. The subpopulation
of YT-1 cells expressing IL-2Rα was purified via magnetic selection as
described previously^{39 (link)}.
Enrichment and persistence of IL-2Rα expression were monitored by
analysis of PE-conjugated anti-human IL-2Rα (Biolegend) antibody binding
on an Accuri C6 flow cytometer (BD Biosciences).

Silva D.A., Yu S., Ulge U., Spangler J.B., Jude K.M., Labão-Almeida C., Ali L.R., Quijano-Rubio A., Ruterbusch M., Leung I., Biary T., Crowley S.J., Marcos E., Walkey C.D., Weitzner B.D., Pardo-Avila F., Castellanos J., Carter L., Stewart L., Riddell S., Pepper M., Bernardes G.J., Dougan M., Garcia K.C, & Baker D. (2019). De novo design of potent and selective mimics of IL-2 and IL-15. Nature, 565(7738), 186-191.

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7

Diverse Cancer Cell Line Maintenance Protocol

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PL45 (pancreatic ductal adenocarcinoma (CTRL-2558)), HeLa (cervix adenocarcinoma
(CCL-2)), DU 145 (prostate carcinoma (HTB-81)) and Hep G2 (hepatocellular
carcinoma (HB-8065)) were purchased from American Type Culture Collection (ATCC)
and maintained in Dulbecco’s Modified Eagle’s Medium
(DMEM) supplemented with sodium bicarbonate (1.5 g/L). TOV-21G
(ovarian clear cell carcinoma) was purchased from ATCC (CTRL-11730) and
maintained in a 1:1 mixture of MCDB 105 medium (1.5 g/L sodium
bicarbonate) and Medium 199 (2.2 g/L sodium bicarbonate). Ramos
(Burkitt’s lymphoma (CTRL-1596)), CCRF-CEM (acute lymphoblastic
leukaemia (CRM-CCL-119)), A549 (lung carcinoma (CCL-185)) and H226 (lung
squamous cell carcinoma (CRL-5826)) were purchased from ATCC and maintained in
RPMI-1640 medium. hTERT HPNE (normal pancreatic ductal cell (CRL-4023)) was
purchased from ATCC and maintained in recommended media (1:3 mixture M3:Base
F^TM Culture Media: DMEM). All media were supplemented with
heat inactivated fetal bovine serum (10% v/v (Gibco)) and
penicillin-streptomycin (100 UI/mL (Gibco)). All cells were cultured
at 37 °C in a 5% CO₂ atmosphere. In addition
to maintain the integrity of the cell lines and prevent mutation of the cells,
the cells passaging were kept under 75.

Champanhac C., Teng I.T., Cansiz S., Zhang L., Wu X., Zhoa Z., Fu T, & Tan W. (2015). Development of a panel of DNA Aptamers with High Affinity for Pancreatic Ductal Adenocarcinoma. Scientific Reports, 5, 16788.

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8

Purification and Culture of IL-2Rα+ YT-1 NK Cells

Cited 1 time

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Unmodified YT-1⁵³ and
IL-2Rα⁺ YT-1 human NK cells⁵⁴ were cultured in RPMI complete medium
(RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM L-glutamine,
minimum non-essential amino acids, sodium pyruvate, 25 mM HEPES, and
penicillin-streptomycin [Gibco]). CTLL-2 cells purchased from ATCC were cultured
in RPMI complete with 10% T-STIM culture supplement with ConA (Corning). 24
hours prior to signaling studies, CTLL-2 cells were resuspended in RPMI lacking
T-STIM culture supplement for IL-2 starvation. Viability (>95%) of CTLL-2
cells was verified by trypan blue exclusion (counted in a hemocytometer)
immediately before performing the signaling assays. All cells were maintained at
37°C in a humidified atmosphere with 5% CO₂. The subpopulation
of YT-1 cells expressing IL-2Rα was purified via magnetic selection as
described previously^{39 (link)}.
Enrichment and persistence of IL-2Rα expression were monitored by
analysis of PE-conjugated anti-human IL-2Rα (Biolegend) antibody binding
on an Accuri C6 flow cytometer (BD Biosciences).

Silva D.A., Yu S., Ulge U., Spangler J.B., Jude K.M., Labão-Almeida C., Ali L.R., Quijano-Rubio A., Ruterbusch M., Leung I., Biary T., Crowley S.J., Marcos E., Walkey C.D., Weitzner B.D., Pardo-Avila F., Castellanos J., Carter L., Stewart L., Riddell S., Pepper M., Bernardes G.J., Dougan M., Garcia K.C, & Baker D. (2019). De novo design of potent and selective mimics of IL-2 and IL-15. Nature, 565(7738), 186-191.

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9

Primary Neuronal Cell Extraction and Culture

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Primary neuronal cells were extracted from the cortical tissue of E17-E18 rat embryos (Sprague-Dawley)29 and maintained in DMEM (GIBCO, Thermofisher Scientific, MA, USA) with 10% fetal bovine serum (FBS, Biowest, Nuaillé, France) during extraction. Afterward, chemical dissociation was carried out in FBS-free DMEM adding trypsin and incubating the mixture at 37 °C. Once the cell density was quantified in a hemocytometer, cells were resuspended in Neurobasal/FBS (GIBCO) medium supplemented with B27, GlutaMAX, and penicillin-streptomycin (GIBCO) and seeded at 3 × 105 cells per well in 12 well plates on glass coverslips. Cell culture was then maintained in an incubator at 37 °C and 5% CO2.

Gallego I., Villate-Beitia I., Soto-Sánchez C., Menéndez M., Grijalvo S., Eritja R., Martínez-Navarrete G., Humphreys L., López-Méndez T., Puras G., Fernández E, & Pedraz J.L. (2020). Brain Angiogenesis Induced by Nonviral Gene Therapy with Potential Therapeutic Benefits for Central Nervous System Diseases. Molecular pharmaceutics, 17(6).

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10

Investigating Cellular Response to WTC Dust Exposure

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Human cell line GM03798 (wild type), Epstein–Barr
virus-transformed lymphoblasts were obtained from Coriell Cell Repositories and
were grown in RPMI 1640 (Gibco) supplemented with 10% FBS and
penicillin/streptomycin (Gibco). Cell lines were regularly tested for mycoplasma
contamination. Cells were treated with the different concentrations of the
following drugs: olaparib (AZD2281; Astra Zeneca); Aphidicolin (A0781, Sigma).
WTC-PM was obtained, as previously described, from 5 locations within 0.5 miles
of Ground Zero on 9/13/01^{6 ,17 (link)}. Composition was determined by
x-ray fluorescence analysis using techniques as previously published^{17 (link)}. Cells were exposed to
200μg/ml of WTC-PM for all the experiments.

Jasra S., Giricz O., Zeig-Owens R., Pradhan K., Goldfarb D.G., Barreto-Galvez A., Silver A.J., Chen J., Sahu S., Gordon-Mitchell S., Choudhary G., Aluri S., Bhagat T.D., Shastri A., Bejan C.A., Stockton S.S., Spaulding T.P., Thiruthuvanathan V., Goto H., Gerhardt J., Haider S.H., Veerappan A., Bartenstein M., Nwankwo G., Landgren O., Weiden M.D., Lekostaj J., Bender R., Fletcher F., Greenberger L., Ebert B.L., Steidl U., Will B., Nolan A., Madireddy A., Savona M.R., Prezant D.J, & Verma A. (2022). High burden of Clonal Hematopoiesis in First Responders Exposed to the World Trade Center Disaster. Nature medicine, 28(3), 468-471.

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Andpenicillin streptomycin

Lab products found in correlation

Close spelling variants of this product

10 protocols using andpenicillin streptomycin

Isolation and Culture of Sensory Neurons

Isolation and Culture of Sensory Neurons

Cell Culture Media Optimization

Colon Cancer Cell Lines and Knockdown

EGF Stimulation of A431 Cells

Purification and Culture of IL-2Rα+ YT-1 NK Cells

Diverse Cancer Cell Line Maintenance Protocol

Purification and Culture of IL-2Rα+ YT-1 NK Cells

Primary Neuronal Cell Extraction and Culture

Investigating Cellular Response to WTC Dust Exposure

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