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Pen strep

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Pen/Strep is a sterile solution containing penicillin and streptomycin, two antibiotics commonly used in cell culture applications. It is designed to prevent bacterial contamination in cell culture media and other biological samples.

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Lab products found in correlation

Fetal bovine serum (fbs), by Merck Group (2 mentions) Hek293t, by American Type Culture Collection (2 mentions) Cytoone 24 well plate, by Starlab (2 mentions) Sodium persulfate, by Merck Group (2 mentions) Ammonium molybdate, by Merck Group (2 mentions) Ferrozine, by Merck Group (2 mentions) 2 7 dichlorofluorescin diacetate dcfda 4 6 diamidino 2 phenylindole dapi, by HiMedia (2 mentions) Congo red, by HiMedia (2 mentions) Ferric chloride, by Merck Group (2 mentions) Trichloroacetic acid, by Merck Group (2 mentions) Monosaccharide, by Merck Group (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Hydrogen peroxide, by Merck Group (2 mentions) 2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (2 mentions) Thiobarbituric acid, by Merck Group (2 mentions) Trifluoroacetic acid, by Merck Group (2 mentions) Ethylenediaminetetraacetic acid, by Merck Group (2 mentions) Sodium borohydride, by Merck Group (2 mentions) Potassium ferricyanide, by Merck Group (2 mentions) Ascorbic acid, by Merck Group (2 mentions)

Spelling variants of this product

Penicillin/Streptomycin (Pen/Strep) (2 citations)

10 protocols using pen strep

1

Fetal Bovine Serum Culture Media

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Culture media consisted of 1.5 ml of Fetal Bovine Serum (heat inactivated; Sigma F4135), 8.3 ml DMEM/F-12 50/50 with L-glutamine (Corning 15-090-CV), 10 μl of 50 mg/ml Ascorbic Acid for final 50 μm/ml concentration (Fisher #S25184). 200 μl of Pen-Strep (for total of 2 %; MP Biomedicals 1670049) was added to limit bacterial contamination.
Diaz RE J.r, & Trainor P.A. (2015). Hand/foot splitting and the ‘re-evolution’ of mesopodial skeletal elements during the evolution and radiation of chameleons. BMC Evolutionary Biology, 15, 184.
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2

Mammalian Cell Culture and Metabolite Extraction

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All mammalian cell lines (HEK293T, HeLa, A549, THP1 and MCF7) described in this paper were purchased from ATCC, and cultured in complete medium [RPMI1640 supplemented with 10% (v/v) FBS, and 1% (v/v) penicillin-streptomycin (Pen-Strep) (MP Biomedicals)] at 37 °C with 5 % (v/v) CO2 unless otherwise mentioned. The cell cultures were stained routinely with 4’,6-diamidino-2-phenylindole (DAPI) and assessed by microscopy to ensure that they were devoid of any mycoplasma contamination using established protocols33 (link). All cell lines were cultured in 15 cm tissue culture dishes (Eppendorf), and upon 80% confluence were harvested by scrapping, washed with Dulbecco’s phosphate buffer saline (DPBS) (3x times), centrifuged at 200g for 5 mins to remove excess DPBS, and stored at -80 °C till further use. For measuring the cellular levels of CoA and acetyl-CoA, these cell pellets were re-suspended in 150 μL of extraction buffer (1:1 methanol and water with 5% glacial acetic acid) by vortexing, and incubated on ice for 10 min. This was followed by the addition of 1.5 μL of 5 M ammonium formate, vigorous vortexing and a further 5 min incubation on ice. Finally, the extracts were centrifuged at 16,000g for 5 min at 4 °C, the supernatant was transferred in 1 mL glass vials, dried down under vacuum and stored at -40 °C until the LC-MS analysis34 (link).
Rajendran A., Vaidya K., Mendoza J., Bridwell-Rabb J, & Kamat S.S. (2019). Functional annotation of ABHD14B, an orphan serine hydrolase enzyme. Biochemistry, 59(2), 183-196.
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3

HIBCPP Cell Culture Protocol

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HIBCPP cells were cultured in a T175 flask (Greiner, Germany) in HIBCPP 10% medium consisting of Dulbecco’s modified eagle medium (DMEM)/Ham’s F12 1:1 medium (Life Technologies, UK) supplemented with 4 mM L-Glutamine, 5 μg/mL insulin, 10% fetal calf serum (FCS) (Gibco, USA), and 2.5% pen/strep (MP Biomedicals, Santa Ana, CA, USA). When HIBCPP cells reached confluence, they were further seeded on filter inserts (Cell inserts Sarstedt, Germany; pore diameter 5.0 μm, pore density 6.0 × 105 pores/cm2, 0.33 cm2) in standard (CytoOne© 24 well plate StarLab, Milton Keynes, UK) culture model. HIBCPP cells that reached a TEER of [50 Ω × cm2] were further switched to HIBCPP 1% medium consisting of DMEM/Ham’s F12 1:1 medium, supplemented with 4 mM L-glutamine, 5 μg/mL insulin, and 1% heat inactivated FCS. Cells were ready to use for experiments 24 h after this last medium change.
Wiatr M., Staubach S., Figueiredo R., Stump-Guthier C., Ishikawa H., Schwerk C., Schroten H., Hanisch F.G., Rudolph H, & Tenenbaum T. (2020). Echovirus-30 Infection Alters Host Proteins in Lipid Rafts at the Cerebrospinal Fluid Barrier In Vitro. Microorganisms, 8(12), 1958.
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4

HIBCPP Inverted and Normal Culture Model

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HIBCPP 10% medium consisted of Dulbecco’s modified eagle medium (DMEM)/Ham’s F12 1:1 medium (Life technologies, Birchwood, UK) supplemented with 4 mM L-Glutamine, 5 μg/mL insulin, 10% fetal calf serum (FCS; Gibco, Gaithersburg, MD, USA) and 2.5% Pen/Strep (MP Biomedicals, Irvine, CA, USA), HIBCPP 1% medium consisted of DMEM/Ham’s F12 1:1 medium, supplemented with 4 mM L-Glutamine, 5 μg/mL insulin and 1% heat-inactivated FCS. Cell inserts Sarstedt, Germany; pore diameter 5.0 μm, pore density 6.0 × 105 pores/cm2, 0.33 cm2) in the inverted (CytoOne© 12-well plate StarLab, Hamburg, Germany) or normal (CytoOne© 24-well plate StarLab, Belgium) were used in the culture model. PBS (Gibco, Thermo Fisher, Waltham, MA, USA). Primary antibody, monoclonal mouse light diagnostics™ anti-PAN Enterovirus (Merck, Darmstadt, Germany). Secondary antibody, Alexa Fluor anti-mouse 488, and phalloidin Alexa Fluor 660 (Molecular Probes, Eugene, OR, USA). Live/Dead viability assays (Thermo Fisher Scientific, Waltham, MA, USA). RNeasy™ Micro Kit Quick Start protocol (QIAGEN, Hilden, The Netherlands). MACE-Seq kit v. 2.0 (GenXPro GmbH, Germany). Qubit HS dsDNA assay (Thermo Fisher Scientific, Waltham, MA, USA). immobilization beads (Agencourt AMPure XP, Beckman Coulter, Brea, CA, USA).
Wiatr M., Figueiredo R., Stump-Guthier C., Winter P., Ishikawa H., Adams O., Schwerk C., Schroten H., Rudolph H, & Tenenbaum T. (2020). Polar Infection of Echovirus-30 Causes Differential Barrier Affection and Gene Regulation at the Blood–Cerebrospinal Fluid Barrier. International Journal of Molecular Sciences, 21(17), 6268.
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5

Macrophage Functional Assays Protocol

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RAW 264.7 murine macrophages were purchased from National Centre for Cell Science, Pune, India. 2-Deoxy-D-ribose, NaOH, ferric chloride, hydrogen peroxide, thiobarbituric acid, ferrozine, trichloroacetic acid, potassium ferricyanide, trifluoroacetic acid, DPPH, ABTS, sodium persulfate, ammonium molybdate, sodium borohydride, pyridine, ascorbic acid, ethylenediaminetetraacetic acid (EDTA), bovine serum albumin (BSA), LPS and monosaccharides were procured from Sigma chemicals Co. (St. Louis, MO, USA). A mushroom β glucan kit was used from Megazyme Institute Wicklow, Ireland. Dulbecco’s Modified Eagle Medium (DMEM), neutral red, sulfanilamide, naphthylethylenediamine dihydrochloride, phosphoric acid, Congo red, 2′,7′-dichlorofluorescin diacetate (DCFDA), 4′,6-diamidino-2-phenylindole (DAPI) were purchased from Himedia, Mumbai, India. WST was obtained from Takara Bio Inc, Japan. Fetal bovine serum (FBS) and TRIzol were purchased from Invitrogen, New Delhi, India. PenStrep, amphotericin B and cDNA preparation kit were used from MP Biomedicals, Santa Ana, CA, USA. PowerUpTM SYBR® Green Master Mix was procured from Applied Biosystems, USA.
Khatua S, & Acharya K. (2019). Alkali treated antioxidative crude polysaccharide from Russula alatoreticula potentiates murine macrophages by tunning TLR/NF-κB pathway. Scientific Reports, 9, 1713.
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6

Culturing Human Leukemic Cell Lines

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Human leukemic cell lines MOLT-4, Jurkat, Ball, HL-60, NB4, HEL, K-562, and U937 were purchased from the American Type Culture Collection (ATCC; Rockville, MD, USA) and were cultured in RPMI 1640 medium containing 10 % heat-inactivated fetal bovine serum (Gibco by Life Technologies, Carlsbad, CA, USA), 1 % pen/strep (MP Biomedicals, Solon, OH, USA) and 2 mM L-glutamine at 37 °C in 95 % air/5 % CO2 with 95 % humidity.
Huang W., Lu C., Wu Y., Ouyang S, & Chen Y. (2015). T-type calcium channel antagonists, mibefradil and NNC-55-0396 inhibit cell proliferation and induce cell apoptosis in leukemia cell lines. Journal of Experimental & Clinical Cancer Research : CR, 34(1), 54.
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7

Culturing MDA-MB-231 Cells in Normoxia and Hypoxia

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MDA-MB-231 cells were cultured in RPMI 1640 medium supplemented with 10% FBS (Sigma), 1× Glutagro (Corning), and 1× Pen/Strep (MP Biomedicals, LLC) (growth medium). Cells cultured in normoxia were maintained at 37 °C in ambient atmosphere (∼21% O2) with 5% CO2. Cells cultured in hypoxia were maintained at 37 °C, < 1% O2 in a nitrogen filled hypoxic chamber (Billups-Rothenberg, Inc.), and 5% CO2.
Costales M.G., Haga C.L., Velagapudi S.P., Childs-Disney J.L., Phinney D.G, & Disney M.D. (2017). Small Molecule Inhibition of microRNA-210 Reprograms an Oncogenic Hypoxic Circuit. Journal of the American Chemical Society, 139(9), 3446-3455.
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8

Antioxidant and Anti-inflammatory Assays

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2-Deoxy-D-ribose, ferric chloride, hydrogen peroxide, thiobarbituric acid, L-methionine, nitroblue tetrazolium (NBT), riboflavin, ferrozine, potassium ferricyanide, trichloroacetic acid, trifluoroacetic acid (TFA), 2, 2-Diphenyl-1-picrylhydrazyl (DPPH), sodium persulfate, ammonium molybdate, sodium borohydride, pyridine, toluene, dichloromethane, ascorbic acid, ethylene diaminetetraacetic acid (EDTA), butylated hydroxylanisole (BHA), gallic acid, bovine serum albumin (BSA), LPS (extraction from Escherichia coli 026:B6) and monosaccharides were procured from Sigma chemicals Co. (St. Louis, MO, USA). Toluene, DCM, and chloroform were of HPLC grade and monosaccharides were of extra pure form. A mushroom β glucan kit from Megazyme Institute Wicklow, Ireland was used. Dulbecco’s Modified Eagles Medium (DMEM), neutral red, sulfanilamide, naphthylethylenediamine dihydrochloride (NEDD), nutrient broth (NB), p-iodonitrotetrazolium chloride (INT), phosphoric acid, Congo red, 2´, 7´-Dichlorofluorescin diacetate (DCFDA), 4´, 6-Diamidino-2-Phenylindole (DAPI) were purchased from Himedia, Mumbai, India. WST and fetal bovine serum (FBS) were purchased from Takara Bio Inc, Japan and Invitrogen, Carlsbad, CA, USA respectively. PenStrep and amphotericin B were used from MP Biomedicals, Santa Ana, CA, USA.
Khatua S., Dutta A.K., Chandra S., Paloi S., Das K, & Acharya K. (2017). Introducing a novel mushroom from mycophagy community with emphasis on biomedical potency. PLoS ONE, 12(5), e0178050.
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9

Generating HIV Luciferase Reporter Cell Line

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HEK293T (CRL-11268) were obtained from ATCC and maintained in DMEM (HyClone, SH30022.FS) supplemented with 10% fetal bovine serum (FBS) (MilliporeSigma, H9268) and 1% Penicillin/Streptomycin (Pen/Strep) (MP Biomedicals, 091670049). Jurkat CD4+ T cells (TIB-152) were obtained from ATCC and maintained in RPMI-1640 (HyClone, SH30027.FS), 10% FBS, 1% Pen/Strep. Jkt-HIVLuc cells were generated in the D’Orso lab by pNL4.3-deltaEnv-deltaVpr-Luc45 (link) pseudotyped VSVG lentivirus transduction and single cell sorted by the UTSW Flow Cytometry Core into 96-well plates. Clones were expanded and screened for low basal luciferase activity and high range of inducibility upon TNFα (Sigma, T6674) dose-response stimulation.
Ramirez N.G., Lee J., Zheng Y., Li L., Dennis B., Chen D., Challa A., Planelles V., Westover K.D., Alto N.M, & D’Orso I. (2022). ADAP1 promotes latent HIV-1 reactivation by selectively tuning KRAS–ERK–AP-1 T cell signaling-transcriptional axis. Nature Communications, 13, 1109.
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10

Mammalian Cell-Based Myxoma Virus Study

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Mammalian cells used in this study include BSC-40 (ATCC CRL-2761) (in Dulbecco minimal essential medium, DMEM, Lonza), healthy human periphery CD14 + monocytes (Lonza, Walkersville, MD), THP1 (ATCC TIB-202), and THP1-Lucia (kindly provided by F. Zhu) (50) (in RPMI1640, Lonza). The complete growth medium (e.g., DMEM Lonza/BioWhittaker Catalog no 12-604Q, or RMPI1640) was supplemented with 10% FBS (Atlanta Biologicals, Minneapolis, MN), 2 mM glutamine (Corning Cellgro, Millipore Sigma, St. Louis, MO), and 100 µg per ml of Pen/Strep (Corning Cellgro, Millipore Sigma, St. Louis, MO); for RPMI1640 complete culture medium, in addition to FBS, glutamine, and Pen/Strep, 2-mercaptoethanol (MP biomedicals, Solon, OH) was supplemented to a final concentration of 0.05 µM.
The viruses used were all derived from myxoma virus (MYXV), Lausanne strain (GenBank Accession AF170726.2). The MYXV M062R deletion mutant (vMyxM062RKOgfp) has been described previously (7) . Myxoma virus stocks were prepared on BSC-40 cells and purified with sucrose step gradient through ultracentrifugation as previously described (51).
Conrad S.J., Peterson E.A., Liem J., Connor R., Nounamo B., Cannon M.J., & Liu J. (2022). Myxoma virus lacking the host range determinant M062 stimulates cGAS-dependent type 1 interferon response and unique transcriptomic changes in human macrophages.
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