Optipro serum free media

Manufactured by Thermo Fisher Scientific

Sourced in United States

The OptiPro Serum-Free Media is a cell culture medium designed for the growth and maintenance of various cell lines in a serum-free environment. It provides the necessary nutrients and growth factors to support cell proliferation and viability without the use of animal-derived serum components.

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Lab products found in correlation

Pen strep, by Thermo Fisher Scientific (2 mentions) Glutamax, by Thermo Fisher Scientific (2 mentions) Polyethylenimine (pei), by Thermo Fisher Scientific (1 mentions) Nickel nta, by Qiagen (1 mentions) Polyethylenimine (pei), by Polysciences (1 mentions) Hygromycin, by Merck Group (1 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions) Expicho s cells, by Thermo Fisher Scientific (1 mentions) Mabselect sure, by Cytiva (1 mentions) Expifectamine cho reagent, by Thermo Fisher Scientific (1 mentions) Zeba spin desalting column, by Thermo Fisher Scientific (1 mentions) 0.22 m filter, by Thermo Fisher Scientific (1 mentions) Beta propiolactone bpl, by Merck Group (1 mentions) Pen step 100x, by Corning (1 mentions) L glutamine, by Thermo Fisher Scientific (1 mentions) Fetal bovine serum (fbs), by Merck Group (1 mentions)

6 protocols using optipro serum free media

Recombinant Cytokine Receptor Purification

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cDNA corresponding to the extracellular domains of CNTFR (1–342), LIFR (1–534), and gp130 (1–619) was cloned into the pAdd2 plasmid and amplified in DH10B cells. For expression, purified plasmids were transfected into human HEK 293 cells using PEI (#23966–2, Polysciences). Briefly, PEI was dissolved in dH₂O to 1 g/L. For 500 mL transfection volume, 0.5 mg of purified DNA and 1 mL of PEI was dissolved in 10 mL of OptiPro Serum-Free Media (#12309–019, Thermo Fisher Scientific) each, then mixed immediately. After 15 min the solution was added dropwise to 500 mL of cells. The cells were incubated on a rotary shaker at 120 RPM in a humidified incubator at 37°C and 5% CO₂. Fc fusion proteins were purified using a protein A (#101142, Fisher Scientific) affinity column; proteins containing a hexahistidine tag were purified using a nickel-NTA (#30210, Qiagen) affinity column. Proteins were then further purified using size exclusion chromatography. The following extinction coefficients were used for protein quantification: CNTFR variants: 70,275 M⁻¹cm⁻¹; CNTFR-Fc variants: 206,410 M⁻¹cm⁻¹; gp130: 130,470 M⁻¹cm⁻¹; gp130-Fc: 326,800 M⁻¹cm⁻¹; LIFR: 98,610 M⁻¹cm⁻¹; and LIFR-Fc: 263,080 M⁻¹cm⁻¹.

Kim J.W., Marquez C.P., Kostyrko K., Koehne A.L., Marini K., Simpson D.R., Lee A.G., Leung S.G., Sayles L.C., Shrager J., Ferrer I., Paz-Ares L., Gephart M.H., Vicent S., Cochran J.R, & Sweet-Cordero E.A. (2019). Anti-tumor activity of an engineered decoy receptor targeting CLCF1-CNTFR signaling in lung adenocarcinoma. Nature medicine, 25(11), 1783-1795.

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Vero Cell-based Virus Titration

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Vero (African green monkey kidney epithelial) cells were plated onto 12 well plates in duplicate at a density of 5 × 10⁴ cells per well in OptiPro Serum free media (1230901, Thermo Fisher, United States), 1% Pen/Strep, 2 × Glutamax (35050061, Thermo Fisher, United States) and were cultured overnight before infection the following day. Virus preparations were diluted in 300 μL OptiPro media, starting as 1 in 8 dilution with a two-fold serial dilution down to 1 in 8192. Plates were incubated for 4 h with gentle rocking to prevent cells from drying, and then topped up to 1.2 mL media. After 6 days, wells displaying 100% GFP expression without significant cell death were selected as the dilution of virus to be used for large scale infection. Titration of the virus was also set up following the TCID₅₀ calculator. Vero cells were seeded at 1 × 10⁴ cells per well in 96 well plate. Virus was added in the dilutions 1 in 10 and diluted down 10-fold and FFU calculated by using the TCID₅₀ calculator and by observing GFP.

Collett S., Earnest L., Carrera Montoya J., Edeling M.A., Yap A., Wong C.Y., Christiansen D., Roberts J., Mumford J., Lecouturier V., Pavot V., Marco S., Loi J.K., Simmons C., Gulab S.A., Mackenzie J.M., Elbourne A., Ramsland P.A., Cameron G., Hans D., Godfrey D.I, & Torresi J. (2023). Development of virus-like particles with inbuilt immunostimulatory properties as vaccine candidates. Frontiers in Microbiology, 14, 1065609.

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Production of Recombinant Human ADAMTS13

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cDNA for human ADAMTS13 was cloned into pcDNA‐3 vector with N‐terminal Myc and multi‐His tags (ThermoFisher Scientific, Waltham, MA, USA) and delivered into human embryonic kidney cells (HEK293) using lipids as the transfection mediator cells.27 The cells were grown in Dulbecco Modified Eagle Medium (Invitrogen, Carlsbad, CA, USA) containing 500 μg/ml hygromycin (Sigma Aldrich, St. Louis, MO, USA) and 10% fetal bovine serum at 37°C. Those expressing high levels of ADAMTS13 were selected by single cell cloning using flow cytometric cell sorting. Selected cells were then incubated in Optipro serum free media (ThermoFisher Scientific, Waltham, MA, USA) for 72 hours and the conditioned media was tested for ADAMTS13 content and frozen at −80°C.

Ng C.J., McCrae K.R., Ashworth K., Sosa L.J., Betapudi V., Manco‐Johnson M.J., Liu A., Dong J., Chung D., White‐Adams T.C., López J.A, & Di Paola J. (2018). Effects of anti‐β2GPI antibodies on VWF release from human umbilical vein endothelial cells and ADAMTS13 activity. Research and Practice in Thrombosis and Haemostasis, 2(2), 380-389.

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Recombinant antibody production in ExpiCHO cells

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ExpiCHO-S cells (Thermo Fisher, A29133) were cultured in an Erlenmeyer flask with vent cap (Corning) at 37 °C with 125 rpm shaking and were transfected with 1 µg/mL of each of the heavy and light chain expression plasmids using the ExpiFectamine CHO Reagent (Thermo Fisher). Transfection mix was prepared in OptiPro Serum Free Media (Thermo Fisher), gently mixed and incubated at 25 °C for 5 min, before being added to the flask. On day 2 after transfection, ExpiCHO Enhancer (Thermo Fisher) and ExpiCHO Feed (Thermo Fisher) were added to the flask and the growth temperature was lowered to 32 °C. On day 5 after transfection, ExpiCHO Feed was added to the flask according to the manufacturer’s instructions. The culture was harvested on day 10 after transfection, by spinning down at 1400 × g for 10 min. The supernatant was passed through a 0.22 µm filter (Thermo Fisher). Antibodies were affinity-purified from the supernatant using a MabSelect SuRe (Cytiva) pre-packed column and eluted in 0.2 M sodium citrate pH 3.0, then neutralized with 1.5 M Tris-HCl pH 8.5. The antibodies were desalted using a Zeba Spin Desalting Column (Thermo Fisher) into PBS pH 7.4.

Vester S.K., Rahikainen R., Khairil Anuar I.N., Hills R.A., Tan T.K, & Howarth M. (2022). SpySwitch enables pH- or heat-responsive capture and release for plug-and-display nanoassembly. Nature Communications, 13, 3714.

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Rabies Virus Immunization and Antibody Response

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Groups of 8–10 weeks old C57BL/J6 female mice (Jackson) were immunized intramuscularly (i.m) via gastrocnemius with 100 μl (50 μl/leg) of live or inactivated RABV or RABV-ED51-mBAFF as indicated in the figures. Inactivated RABV and inactivated RABV-ED51-mBAFF were prepared as follows: virus stocks were grown in OptiPRO Serum Free Media (Gibco) [4mM L-Glutamine, 1% PS], harvested, and cell debris was removed using Corning 0.45μm filter (430516; Corning). β-Propiolactone (BPL; P5648; Sigma) was added to viral supernatants (final concentration 0.05% BPL), and incubated overnight at 4ᵒC. Treated supernatants were purified using ultracentrifugation. Viral inactivation was confirmed by viral titer [37 (link)]. Total protein concentrations were quantified by BCA Protein Assay Kit as described by the manufacturer (Pierce). Blood from immunized mice was collected via retro-orbital at 5, 7, 10 days post-immunization. RABV G-specific IgM and IgG (and subclasses) antibody levels were determined by ELISA as described previously [36 (link)–39 (link)]. VNA titers were determined by RFFIT as described previously [36 (link), 37 (link), 40 (link)].

Plummer J.R, & McGettigan J.P. (2019). Incorporating B cell activating factor (BAFF) into the membrane of rabies virus (RABV) particles improves the speed and magnitude of vaccine-induced antibody responses. PLoS Neglected Tropical Diseases, 13(11), e0007800.

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PFAS Exposure in Human Placental Cells

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PFOS, PFOA, and HFPO-DA were acquired according to Supplemental Table S1 and individual stock solutions (100,000 μg/mL) created by dissolving the PFAS in 100% methanol. Cell culture media containing PFAS treatments were prepared within 24 hours of application to cells and contained 0.1% methanol. Control media also contained 0.1% methanol. Dimethyl sulfoxide (DMSO) was not utilized for tissue culture since DMSO has been found to degrade the ether portion of HFPO-DA in solution²⁹.
The JEG3 cell line, trophoblast lineage derived from a human placental choriocarcinoma, was acquired from ATCC (HTB-36; Manassas, VA) and cultured in serum-supplemented media (SSM) and serum-free media (SFM) as detailed in Supplemental Table S2. In brief, SSM media components included Minimum Essential Media (MEM; Gibco, Grand Island, NY), Pen/Step 100x (Corning, Manassas, VA), Fetal Bovine Serum (EMD Millipore Corp., Temecula, CA). While SFM media components included OptiPRO Serum Free Media (Gibco, Grand Island, NY), Pen/Step 100x (Corning, Manassas, VA), and L-glutamine (Gibco, Grand Island, NY). Of note, the SFM chosen for this study was supplemented media that sustains cells without the addition of Fetal Bovine Serum (FBS). In addition, SFM contained 10% of the Pen/Strep that was added to SSM³⁰.

Bangma J., Szilagyi J., Blake B.E., Plazas C., Kepper S., Fenton S.E, & Fry R.C. (2020). An assessment of serum-dependent impacts on intracellular accumulation and genomic response of per- and polyfluoroalkyl substances in a placenta trophoblast model. Environmental toxicology, 35(12), 1395-1405.

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