Bovine serum

Manufactured by Merck Group

Sourced in United States, Poland, Germany, China

Bovine serum is a common laboratory reagent derived from the blood of cattle. It contains a complex mixture of proteins, growth factors, and other biomolecules that support the growth and maintenance of cell cultures in vitro. Bovine serum is commonly used as a supplement in cell culture media to provide a nutrient-rich environment for the cultivation of various cell types.

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Close spelling variants of this product

-free bovine serum albumin (BSA; Bovine Serum Albumin in dH20 Albumin from bovine serum (BSA), Insect tested fetal bovine serum Bovine Serum Albumin – A3311 Bovine serum albumin (BSA) 0.1% Serum bovine albumin Fetal bovine serum (F7524) Fatty acid-free bovine serum albumin (BSA) fraction V Adult bovine serum

80 protocols using bovine serum

Stability of 52Mn-DOTA Complex in Serum

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Two vials containing purified 52 Mn in 0.1 M HCl were dried under nitrogen flow at 130 o C. After cooling to room temperature, the 52 Mn in one vial was redissolved in isotonic HEPES buffer (10 mM HEPES, 150 mM NaCl, pH = 7.4) to a radioactivity concentration of 13 MBq/mL. The 52 Mn in the other vial was redissolved in isotonic HEPES buffer containing 25 µM DOTA, likewise to a concentration of 13 MBq/mL. Radio-TLC after 5 minutes confirmed that all 52 Mn in the DOTA-containing solution was present as Mn-DOTA. Using 100 µL aliquots of these stocks, four different 1 mL preparations of 52 Mn were produced by dilution with whole fetal bovine serum (Sigma Aldrich) and isotonic HEPES buffer: 52 Mn-DOTA in 75% bovine serum, 52 Mn in 75% bovine serum, 52 Mn-DOTA in isotonic HEPES buffer and 52 Mn in isotonic HEPES buffer.
After incubation at 37 ˚C for 2 days, the mixtures were applied to PD-10 size exclusion cartridges (GE Healthcare), eluted with isotonic HEPES and collected in 1 mL fractions. The radioactivity in the collected fractions was measured, and the peak fractions from the DOTA-containing samples were analyzed by radio-TLC to assess whether the Mn-DOTA complex was still intact.

Fonslet J., Tietze S., Jensen A.I., Graves S.A, & Severin G.W. (2017). Optimized procedures for manganese-52: Production, separation and radiolabeling. Applied radiation and isotopes : including data, instrumentation and methods for use in agriculture, industry and medicine, 121.

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Biochemical Reagent Acquisition Protocol

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Bovine serum, antimycotic antibiotic solution, 2,2-diphenyl-1-pikrilhydrazil (DPPH), dimethyl sulfoxide (DMSO), fetal Bovine serum, 4-(dimethylamino) benzaldehyde, hydrolysate, Ehrlich solution, borax-sulfuric acid mixture, and Dulbecco’s Modified Eagle Medium (DMEM) were all obtained from the US company Sigma-Aldrich.

Wang L., Zhang L., Zheng G., Luo H., El-kott A.F, & El-kenawy A.E. (2021). Equisetum arvense L aqueous extract: a novel chemotherapeutic supplement for treatment of human colon carcinoma. Archives of Medical Science : AMS, 19(5), 1472-1478.

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Evaluating Anti-Parasitic Potency In Vitro

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E. histolytica strain HM1-IMSS was grown in TYI-S-33 medium supplemented with 10% heat-inactivated bovine serum (Sigma Adrich, Toluca, Mexico). G. lamblia strain IMSS:8909:1 trophozoites used in all experiments were cultivated axenically at 37 °C in TYI-S-33 modified medium supplemented with 10% calf serum and bovine bile. In vitro susceptibility tests were performed using E. histolytica (6 × 10³) or G. lamblia (5 × 10⁴) trophozoites were incubated for 48 h at 37 °C in the presence of different concentrations (2.5–200 μg/mL) of pure compounds in DMSO at 2%. After incubation, the trophozoites were detached by chilling and 50 μL samples of each tube were subcultured in fresh medium for another 24 h. The final number of parasites was determined in Neubauer. Then, data were analyzed using probit analysis. The percentage of trophozoites surviving was calculated by comparison with the growth in the control group. The plot of probit against log concentration was made, the best straight line was determined by regression analysis, and the IC₅₀ values were calculated. The regression coefficient, its level of significance (p < 0.05 indicates significant difference between groups), and correlation coefficient were calculated and 95% confidence interval (CI) values determined. ADMET characteristics of selected compounds are shown in Table S3 [34 (link)].

Juárez-Saldivar A., Barbosa-Cabrera E., Lara-Ramírez E.E., Paz-González A.D., Martínez-Vázquez A.V., Bocanegra-García V., Palos I., Campillo N.E, & Rivera G. (2021). Virtual Screening of FDA-Approved Drugs against Triose Phosphate Isomerase from Entamoeba histolytica and Giardia lamblia Identifies Inhibitors of Their Trophozoite Growth Phase. International Journal of Molecular Sciences, 22(11), 5943.

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Culturing Human Ovarian Cancer Cells

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All reagents necessary for cell cultures (RPMI-1640, bovine serum, L-glutamine and penicillin-streptomycin) were purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany) unless indicated otherwise.
Cell culture. The human A2780 ovarian cancer cell line was obtained from the European Collection of Authenticated Cell Cultures (Salisbury, UK). A2780 cells were cultured in RPMI 1640 medium, supplemented with L-glutamine, penicillin-streptomycin (10 U/ml; 100 µg/ml) and 10% fetal bovine serum, in a humidified atmosphere of 95% air and 5% CO 2 at 37˚C.

Kapka-Skrzypczak L., Popek S., Sawicki K., Drop B., Czajka M., Jodłowska-Jędrych B., Matysiak-Kucharek M., Furman-Toczek D., Zagórska-Dziok M, & Kruszewski M. (2019). IL‑6 prevents CXCL8‑induced stimulation of EpCAM expression in ovarian cancer cells. Molecular medicine reports, 19(3).

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In Vitro Hepatoprotective Assay

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The in vitro hepatoprotective assay was based on the protection of human liver-derived HepG2 cells against CCl₄-induced damage. We procured HepG2 cell line from American Type Culture Collection no. (ATCC#HB-8065) (American Type Culture Collection (ATCC), Manassas, VA, USA). Human liver hepatocellular carcinoma cells (HepG2) were grown in DMEM media supplemented with 10% bovine serum, 0.1% antimycotic solution from (Sigma-Aldrich Co., St Louis, MO, USA) at 37°C in a humidified chamber with 5% CO₂. The cells were then treated with medium containing 1% CCl₄ along with or without the test compounds. Silymarin was used as the reference drug. Stocks of all compounds (1.0 mg/mL) were made in DMSO and further dilutions (100 µg/mL) were prepared in culture media. Cells were treated with test compounds, separately; with dose of 25 µg/mL of each compound. Treated cells were incubated in complete growth media for 48 hours. As a positive control, cells were also tested with the reference drug, silymarin. Cytotoxicity was assessed by calculating the viability of the HepG2 cells by MTT reduction assay.25 (link)

Bhat M.A., Al-Omar M.A., Khan A.A., Alanazi A.M, & Naglah A.M. (2019). Synthesis and antihepatotoxic activity of dihydropyrimidinone derivatives linked with 1,4-benzodioxane. Drug Design, Development and Therapy, 13, 2393-2404.

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Candida Biofilm Formation Protocol

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The biofilms were formed according to the methodology described by Tampakakis et al. [10 (link)]. In brief, C. albicans, C. glabrata and C. krusei were grown in YPD medium (2% dextrose, 2% Bacto Peptone, 1% yeast extract) overnight at 30°C and diluted to an OD600 of 0.5 in Spider medium. To form the mixed groups, 1 mL of each Candida species was added to a well of a sterile 12-well plate containing a silicone pad measuring 1.5 × 1.5 cm that had been pretreated overnight with bovine serum (Sigma-Aldrich). For single biofilms, 1 mL or 2 mL of each species of Candida were added.
The inoculated 12-well plate was incubated with gentle agitation (150 rpm) for 90 min at 37°C for adhesion to occur and the standardized samples were washed with 2 mL PBS, and incubation was continued for 60 h at 37°C at 150 rpm in 2 mL of fresh Spider medium. A negative control group of Spider medium without any fungal cells was also included in this study. The silicone pads with biofilm were removed from the wells, dried overnight, and weighed the following day. The total biomass (mg) of each biofilm was calculated by subtracting the weight of the platform material prior to biofilm growth from the weight after the drying period and adjusting for the weight of a control pad unexposed to cells.

Rossoni R.D., Barbosa J.O., Vilela S.F., dos Santos J.D., de Barros P.P., Prata M.C., Anbinder A.L., Fuchs B.B., Jorge A.O., Mylonakis E, & Junqueira J.C. (2015). Competitive Interactions between C. albicans, C. glabrata and C. krusei during Biofilm Formation and Development of Experimental Candidiasis. PLoS ONE, 10(7), e0131700.

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Culturing Spodoptera frugiperda Sf9 cells and Escherichia coli BL21 Transetta

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The Spodoptera frugiperda Sf9 cell line was cultured in the ESF 921 insect cell culture medium (Expression Systems, Davis, CA, USA; 96-001) containing 1.5% bovine serum (Sigma-Aldrich, Saint Louis, MO, USA; F8318), 100 IU/mL penicillin, and 100 μg/mL streptomycin sulfate at 28°C. The recombinant Escherichia coli BL21 Transetta (DE3) (TransGen Biotech, Beijing, China; CD801) strain was cultured in Luria-Bertani broth medium supplemented with 100 mg/mL of ampicillin at 37°C.

Gao J., Song Y.J., Wang H., Zhao B.R, & Wang X.W. (2023). Mindin Activates Autophagy for Lipid Utilization and Facilitates White Spot Syndrome Virus Infection in Shrimp. mBio, 14(2), e02919-22.

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Affinity Purification of C3 Complement Protein

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Pull-downs used to identify C3 (Fig. 1c) were performed as previously described^{22 (link)}. In brief, glutathione-sepharose 4B (GE Healthcare, 140 μl slurry/pull-down) was incubated with GST-tagged protein (1 mg/pull-down) for 30 min, washed with PBS and incubated with bovine serum (Sigma-Aldrich, 1 ml/pull-down) or a PBS control for 60 min. ISG65 E1125A produced in E. coli was used in these experiments. The beads were rapidly washed with three PBS washes with a total time of 5 min. Bound protein was eluted by adding SDS-polyacrylamide gel electrophoresis sample buffer and heating at 50 °C. The pull-downs using GST-ISG65A as bait were performed in three replicates. Pull-downs used to identify C3 from different mammalian species (Fig. 1d) were performed by binding avi-tagged ISG65 E1125G to Pierce Streptavidin Agarose beads (Thermo Fisher). Serum from various mammalian species (Sigma) was added to ISG65-streptavidin beads via gravity flow. The beads were then washed three times with HBS pH 7.4, and C3 bound to ISG65 eluted with 0.1 M sodium acetate pH 4, 500 mM NaCl.

Macleod O.J., Cook A.D., Webb H., Crow M., Burns R., Redpath M., Seisenberger S., Trevor C.E., Peacock L., Schwede A., Kimblin N., Francisco A.F., Pepperl J., Rust S., Voorheis P., Gibson W., Taylor M.C., Higgins M.K, & Carrington M. (2022). Invariant surface glycoprotein 65 of Trypanosoma brucei is a complement C3 receptor. Nature Communications, 13, 5085.

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Comprehensive Antioxidant Screening Protocol

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Standard phenolic compounds (purity > 97%), including phenolic acids (benzoic, caffeic, ferulic, p-coumaric, m-hydroxybenzoic, p-hydroxybenzoic) and flavonoids (apigenin, D-catechin, epicatechin, kaempferol, naringenin, quercetin), Trolox (6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid), DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)), and HPLC-grade solvents and reagents such as acetonitrile, ammonium formate, bovine serum, formic acid, and glucose were purchased from Sigma Aldrich (Poznań, Poland). Analytical-grade reagents such as acetic acid, Folin–Ciocalteau (F-C) reagent, methanol, phosphate buffer, potassium dithionite, sodium carbonate, and sodium hydroxide were supplied by Chempur (Piekary Śląskie, Poland).

Topka P., Poliński S., Sawicki T., Szydłowska-Czerniak A, & Tańska M. (2023). Effect of Enriching Gingerbread Cookies with Elder (Sambucus nigra L.) Products on Their Phenolic Composition, Antioxidant and Anti-Glycation Properties, and Sensory Acceptance. International Journal of Molecular Sciences, 24(2), 1493.

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Bovine Serum Antioxidant Assay

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Bovine serum, antimycotic antibiotic solution, 2,2-diphenyl-1-picrylhydrazyl (DPPH), dimethyl sulfoxide (DMSO), decamplmaneh fetal, 4-(dimethylamino) benzaldehyde, hydrolysate, Ehrlich solution, borax-sulfuric acid mixture, and DMED were all obtained from the US company Sigma-Aldrich.

Liu Z., Zhang Z., Du X., Liu Y., Alarfaj A., Hirad A., Ansari S.A, & Zhang Z. (2021). Novel green synthesis of silver nanoparticles mediated by Curcumae kwangsiensis for anti-lung cancer activities: a preclinical trial study. Archives of Medical Science : AMS, 19(5), 1463-1471.

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