Amphotericin b

Manufactured by BD

Sourced in United States

Amphotericin B is a broad-spectrum antifungal medication used in the treatment of severe fungal infections. It is a polyene macrolide antibiotic produced by the bacterium Streptomyces nodosus. Amphotericin B is primarily used to treat invasive fungal infections, such as aspergillosis, candidiasis, and cryptococcosis.

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Lab products found in correlation

Trimethoprim, by BD (4 mentions) Nalidixic acid, by BD (4 mentions) Polymyxin b, by BD (3 mentions) Azlocillin, by BD (3 mentions) Streptomycin, by BD (2 mentions) Penicillin, by BD (2 mentions) Gentamicin, by BD (2 mentions) Shields and sang m3 insect medium, by Merck Group (1 mentions) Human insulin, by Merck Group (1 mentions) Streptomycin, by Thermo Fisher Scientific (1 mentions) Penicillin, by Thermo Fisher Scientific (1 mentions) Yeast extract, by Merck Group (1 mentions) Centriprep centrifugal filter unit, by Merck Group (1 mentions) Fetal bovine serum (fbs), by Merck Group (1 mentions) Amphotericin b, by Thermo Fisher Scientific (1 mentions) Bacto peptone, by BD (1 mentions) Penicillin, by GE Healthcare (1 mentions) Streptomycin, by GE Healthcare (1 mentions) Penicillin streptomycin, by BD (1 mentions) Mycosensor pcr assay kit, by Agilent Technologies (1 mentions)

11 protocols using amphotericin b

Maintaining Cl.8 and S2 Cell Lines

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The Clone 8 (Cl.8) cell line was maintained in Shields and Sang M3 Insect medium (Sigma-Aldrich), supplemented with 2% heat-inactivated Fetal Bovine Serum (FBS) (Sigma-Aldrich), 2,5% fly extract, 5 µg/ml human insulin (Sigma-Aldrich), 0.25 µg/ml Amphotericin B (Gibco, Life Technologies), 100 U/ml penicillin and 100 μg/ml streptomycin (Gibco, Life Technologies). The Schneider 2 (S2) cell line was also maintained in Shields and Sang M3 Insect medium supplemented with 1 g/l yeast extract (Sigma-Aldrich) and 2.5 g/l bactopeptone (BD Biosciences), 10% heat-inactivated FBS, 0.25 µg/ml Amphotericin B, 100 U/ml penicillin and 100 μg/ml streptomycin. The cells were subcultured weekly and maintained at 25 °C.
To study the presence of miRNAs in EVs or bound to EAgo-1, optimized cell culture media lacking vesicles and Ago proteins were required. EV-depleted FBS and fly extract were obtained by ultracentrifugation (110,000 g, 4 h, 4 °C). In addition, the fly extract was depleted of Ago proteins by using a 30 kDa MWCO Centriprep™ Centrifugal Filter Units (Millipore®).

Van den Brande S., Gijbels M., Wynant N., Santos D., Mingels L., Gansemans Y., Van Nieuwerburgh F, & Vanden Broeck J. (2018). The presence of extracellular microRNAs in the media of cultured Drosophila cells. Scientific Reports, 8, 17312.

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Porcine Adipose Tissue Harvesting

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The study was performed with the permission of the Local Ethics Committee (no. 30/2013). Ten female domestic pigs included in this study weighed between 30 and 40 kg (age 10 to 12 wk). Adipose tissue was harvested from subcutaneous tissue of the abdominal wall. Adipose tissue immediately after resection was transferred to containers with Dulbecco’s Modified Eagle’s Medium/ Ham’s F12 (GE Healthcare Life Science, Logan, UT, USA) supplemented with antibiotics: penicillin (100 U/ml, GE Healthcare Life Science), streptomycin (100 μg/ml, GE Healthcare Life Science), amphotericin B (5 µg/ml, BD Bioscience, Franklin Lakes, NJ, USA) and transported to the laboratory.

Pokrywczynska M., Maj M., Kloskowski T., Buhl M., Balcerczyk D., Jundziłł A., Szeliski K., Rasmus M, & Drewa T. (2020). Molecular Aspects of Adipose-Derived Stromal Cell Senescence in a Long-Term Culture: A Potential Role of Inflammatory Pathways. Cell Transplantation, 29, 0963689720917341.

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Isolation and Culture of Mammary Tumor Cells

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Single cell suspensions were established from spontaneous mammary tumors arising in FVB MMTV-PyVmT females as previously described (43 (link)). Tumors were minced and incubated at 37^oC for 2 hours in 5 ml of Ham’s F12K medium containing 1 mg/ml collagenase (Roche), 2 mg/ml soybean trypsin inhibitor (Sigma) and 2% bovine serum albumin (BSA). After addition of fetal bovine serum (FCS)-containing medium, the suspension was passed through a 70 mm nylon filter (Fisher Scientific). Single cells were pelleted by centrifugation and resuspended in PBS/2% BSA for immediate flow cytometric analysis. Stained cells were analyzed by a FACS Canto cytometer (BD Biosciences). Antibodies for flow cytometric analysis are listed in the Table S3. For establishment of MEC cell lines, cells were cultured in DMEM: F12 (1:1) medium mix containing 5% FCS, 2.5 mg/ml amphotericin B, 10 mg/ml penicillin–streptomycin and MITO+ (BD Biosciences). Non-epithelial cells were removed by differential passaging and epithelial origin of established cell lines were confirmed by CD326 (Biolegend) staining after five passages (CD326⁺ > 98%). The cell lines have been tested as mycoplasma-free using mycosensor PCR assay kit (Agilent Technologies) with the latest test in 2018. The cell line authentication was not routinely performed. The cells were used within two month after thawing.

Fan J.B., Miyauchi S., Xu H.Z., Liu D., Kim L.J., Burkart C., Cheng H., Arimoto K.I., Yan M., Zhou Y., Győrffy B., Knobeloch K.P., Rich J.N., Cang H., Fu X.D, & Zhang D.E. (2020). Type I Interferon Regulates a Coordinated Gene Network to Enhance Cytotoxic T Cell-Mediated Tumor Killing. Cancer discovery, 10(3), 382-393.

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Nasal Swab Sampling for Leprosy

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From May through October 2012, 20 clinically confirmed MB leprosy cases - detected in the context of the national leprosy control programme activities - were enrolled for clinical validation (inclusion criteria: > 5 years of age, > 5 skin lesions, no previous MDT). The cases originated from the regions Maritime (n = 12), Plateaux (n = 6), Centrale (n = 1) and Kara (n = 1).
Eight nasal swab samples per patient (n = 160) were collected with custom-made swabs (Bio-Budget, Krefeld, Germany). Two samples each (one per nostril) were stored in 700 μl cell lysis solution (CLS, Qiagen, Hilden, Germany) and 400 μl tissue lysis buffer (TLS, Bio-Budget) respectively for RLEP qPCR. Two other samples each (one per nostril) were stored in 500 μl PANTA transport medium (comprising Polymyxin B, Amphotericin B, Nalidixic acid, Trimethoprim, Azlocillin; BD, Heidelberg, Germany, Additional file 1: Protocol 5) and 500 μl RNAlater stabilization reagent (Qiagen) respectively for 16S rRNA RT qPCR.

Beissner M., Woestemeier A., Saar M., Badziklou K., Maman I., Amedifou C., Wagner M., Wiedemann F.X., Amekuse K., Kobara B., Herbinger K.H., Kere A.B., Löscher T, & Bretzel G. (2019). Development of a combined RLEP/16S rRNA (RT) qPCR assay for the detection of viable M. leprae from nasal swab samples. BMC Infectious Diseases, 19, 753.

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Evaluating Probiotic Strain Effects on Intestinal Epithelium

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The human colonocyte-like cellular line HT29-MTX (Lesuffleur, Barbat, Dussaulx, & Zweibaum, 1990) was used to test the capability of the two Lb. paraplantarum strains to counteract the effect of pathogens upon the intestinal epithelium. For this purpose, 1x10 5 HT29-MTX cells were seed in 48-wells microplates (BD Falcon, BD Biosciences, NJ, USA) using complete-DMEM, i.e., DMEM medium supplemented with 10% foetal bovine serum and with a mixture of antibiotics (50 g/ml penicillin, 50 g/ml streptomycin, 50 g/ml gentamicin and 1.25 g/ml amphotericin B). All reagents were purchased from Sigma (Sigma Chemical Co., St. Louis, MO, USA). Microplates were incubated at 37ºC, 5% CO 2 in the CO2-Series Shel-Lab incubator (Sheldon Manufacturing Inc. OR, USA) until reach the differentiated and confluent (monolayer) state (12± 1 days post-seeding, about 1x10 7 cells/ml). For cocultivation with bacterial strains, HT29-MTX monolayers were incubated under same conditions in the HERAcell 240 incubator (Thermo Electron LED GmbH, Langenselbold, Germany).

Zivkovic M., Hidalgo-Cantabrana C., Kojic M., Gueimonde M., Golic N, & Ruas-Madiedo P. (2015). Capability of exopolysaccharide-producing Lactobacillus paraplantarum BGCG11 and its non-producing isogenic strain NB1, to counteract the effect of enteropathogens upon the epithelial cell line HT29-MTX. Food research international (Ottawa, Ont.), 74.

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Ascitic Fluid Tuberculosis Detection

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Ascitic fluid samples of all patients were subjected to biochemical and cytological investigations at RMLH, New Delhi. The left over ascitic fluid was transferred to the AIIMS laboratory and processed for liquid culture and Xpert on the same day. Briefly, liquid culture for M. tuberculosis was performed from the sediment obtained after centrifugation of ascitic fluid in Middlebrook 7H9 broth enriched with oleic acid, albumin, dextrose, catalase (OADC) and an antibiotic cocktail; Polymyxin B, Amphotericin B, Nalidixic acid, Trimethoprim and Azlocillin (PANTA) [Becton and Dickinson (BD), Franklin Lakes, New Jersey, United States] (Fig 1). The supernatant was stored at -80°C and used later for cfMTB-DNA extraction (Fig 1). Quantification of cfMTB-DNA was performed targeting the devR region of M. tuberculosis by qPCR.

Sharma P., Anthwal D., Kumari P., Gupta R.K., Lavania S., Sharma N., Sharma L.K., Rath D., Soraganvi P.K., Sharma A., Gadpayle A.K., Taneja R.S., Tyagi J.S, & Haldar S. (2020). Utility of circulating cell-free Mycobacterium tuberculosis DNA for the improved diagnosis of abdominal tuberculosis. PLoS ONE, 15(8), e0238119.

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Culturing and Validating Mycobacterium tuberculosis

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M. tuberculosis H37Rv and the clinical Beijing and LAM genotype families used in this study were grown in Middlebrook 7H9 (Becton-Dickenson, Franklin Lakes, NJ, USA) media supplemented with 0.2% glycerol (Sigma, Springfield, MO, USA), 10% Middlebrook oleic acid-albumin-dextrose-catalase (OADC, Becton-Dickenson) and 0.05% Tween 80 (Sigma, Missouri, USA) to mid-log phase (OD_600nm 0.5–0.8) at 37 °C without shaking. Five individual strains from the Beijing family and five from the LAM family were thawed from −80 °C freezer stocks, streaked on BBL™ Middlebrook 7H11 (Sigma, Springfield, MO, USA) media and incubated for 3 weeks at 37 °C. Single colonies were selected and sub-cultured for 7 days at 37 °C in 5 mL of Middlebrook 7H9 media supplemented with OADC and Polymyxin B, Amphotericin B, Nalidixic acid, Trimethoprim and Azlocillin (PANTA, Becton-Dickenson, Franklin Lakes, NJ, USA) reconstituted in OADC, to ensure the strains were free of contaminants. Once the cultures reached mid-log phase (OD_600nm 0.5–0.8), the cells were used as a preculture to regrow the strains, again in Middlebrook 7H9 media supplemented with OADC and PANTA, to ensure any remaining contamination was killed. The mid-log liquid culture was then spread on BBL™ Middlebrook 7H11 plates supplemented with OADC to confirm that the Beijing and LAM strains were free of contamination.

Gordhan B.G., Padarath K., Sewcharran A., McIvor A., VanNieuwenhze M.S., Waja Z., Martinson N, & Kana B.D. (2024). Clinical Strains of Mycobacterium tuberculosis Representing Different Genotype Families Exhibit Distinct Propensities to Adopt the Differentially Culturable State. Pathogens, 13(4), 318.

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Isolation of Aortic Endothelial Cells

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Porcine aortic endothelial cells were isolated from GalTKO.hCD46.Neu5GcKO experiments (n=4) and GalTKO.hCD46 historical controls (n=3) via scraping and plated on 0.5% gelatin coated culture flasks (Sigma-Aldrich, St. Louis, MO, USA). Culture media consisted of DMEM (Gibco-Life Technologies, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (Atlanta Biological, Lawrenceville, GA, USA), gentamicin, amphotericin B, and endothelial cell growth supplement (Becton Dickinson, San Jose, CA, USA). Human aortic endothelial cells (HAECs) were purchased from Lonza (Allendale, NJ, USA) and cultured in presence of Medium 200 + LSGS Kit supplemented with 10% fetal bovine serum (Thermo Fisher Scientific, Waltham, MA, USA).

Cimeno A., Hassanein W., French B.M., Powell J.M., Burdorf L., Goloubeva O., Cheng X., Parsell D.M., Ramsoondar J., Kuravi K., Vaught T., Uluer M.C., Redding E., O’Neill N., Laird C., Hershfeld A., Tatarov I., Thomas K., Ayares D., Azimzadeh A.M., Pierson RN I.I.I., Barth R.N, & LaMattina J.C. (2017). N-glycolylneuraminic acid knock out reduces erythrocyte sequestration and thromboxane elaboration in an ex vivo pig-to-human xenoperfusion model. Xenotransplantation, 24(6), 10.1111/xen.12339.

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Culturing and Quantifying Mycobacterium avium subsp. paratuberculosis

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The MAP laboratory strain CAPM 6381 (The Collection of Animal Pathogenic Microorganisms, Veterinary Research Institute, Czechia) and field isolates 7082 (white deer) and NL-3 (cattle) were used throughout the whole study. All strains and isolates were of RFLP type C1.
All strains and isolates were grown on Herrold’s egg yolk medium (HEYM) with penicillin G, chloramphenicol, and amphotericin B (Becton Dickinson, Franklin Lakes, NJ, United States), supplemented with 2 μg/ml Mycobactin J (Allied Monitor, Fayette, MO, United States) at 37°C for 2–3 months until visible colonies were observed. Afterward, grown colonies were harvested using a loop and resuspended in 1.5 ml of Middlebrook 7H9 (M7H9) broth supplemented with 10% (vol/vol) Middlebrook OADC enrichment (both Becton Dickinson, Franklin Lakes, NJ, United States). In order to homogenize the MAP suspensions, 12 1-mm zirconia/silica beads (Biospec, Bartlesville, OK, United States) were added followed by vortexing for 30 s. To remove the MAP clumps, the suspension was centrifuged at 100 × g for 30 s and the upper cell fraction was resuspended in fresh M7H9 broth. The suspension was diluted to OD₆₀₀ ≈ 0.15–0.20, which corresponds to approximately 10⁸ MAP cells/ml of suspension (Kralik et al., 2011 (link)).

Kralik P., Babak V, & Dziedzinska R. (2018). The Impact of the Antimicrobial Compounds Produced by Lactic Acid Bacteria on the Growth Performance of Mycobacterium avium subsp. paratuberculosis. Frontiers in Microbiology, 9, 638.

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Isolation of Mycobacterial Pathogens from Wildlife

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Sample collection was conducted by veterinary wildlife specialists to avoid contamination of samples during transportation and at the laboratory (Pate et al. 2016) . The fecal and tissue samples were immediately processed using 2% sodium hydroxide (Hibiya et al. 2010; Ito et al. 2018b ). The inocula obtained from both tissues and feces were inoculated onto a 2% Ogawa slant culture tube (Kyokuto Pharmacy, Tokyo, Japan) and into BBLe (Difco Laboratories, Sparks, Maryland, USA) mycobacterium growth indicator tube liquid media supplemented with polymyxin B, amphotericin B, nalidixic acid, trimethoprim, and azlocillin (Becton Dickinson, Franklin Lakes, New Jersey, USA) and incubated at 37 C for 4 wk. Each single colony formed on the slant tube was subcultured on Middlebrook 7H11 agar (Difco) to obtain distinct colonies (Ito et al. 2018b) .

Odoi J.O., Ohya K., Moribe J., Takashima Y., Sawai K., Taguchi K., Fukushi H., Wada T., Yoshida S, & Asai T. (2020). ISOLATION AND ANTIMICROBIAL SUSCEPTIBILITIES OF NONTUBERCULOUS MYCOBACTERIA FROM WILDLIFE IN JAPAN. Journal of wildlife diseases, 56(4).

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