Nonwovens were initially hydrated with 70% ethanol, washed three times with saline, and soaked in growth media. Wet nonwovens were then placed on the tissue culture well plate, and the adipose tissue fragments, which were minced into approximately 2 mm squares, were placed at the center of the nonwovens (Fig. 1 B). Medium was finally added to cover the top of the fragment. Nonwovens were cultured with Nutristem MSC (SARTORIUS, Göttingen, Germany) containing 2% fetal bovine serum (FBS, Lonza, Basel, Switzerland) and 1% antibiotic solution (Nacalai Tesque, Kyoto, Japan) (referred as growth media) at 37 °C in an incubator with humidified 5% CO2. The medium was changed every 3 days. Tissue attachment on nonwoven was microscopically assessed with three levels as follows: 1, full contact; 0.5, partial contact; 0, detached.
Antibiotic solution
Manufactured by Nacalai Tesque
Sourced in Japan
The Antibiotic solution is a laboratory product designed for use in various scientific applications. It serves as a concentrated solution containing one or more antibiotics. The core function of this product is to provide a source of antibiotics for research, analysis, or other laboratory procedures that require the use of antimicrobial compounds.
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3 protocols using antibiotic solution
1
Adipose Tissue Culture on Nonwovens
2
Isoquercitrin Regulation of AMPK Activity
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Isoquercitrin was purchased from the National Institutes for Food and Drug Control (Beijing, China). Dulbecco’s modified Eagle’s medium (DMEM) and antibiotic solution was obtained from Nacalai Tesque (Kyoto, Japan). AMPK inhibitor, compound C was pucharsed from Santa Cruz Biotechnology, Inc. (Texas, USA), and 5-aminoimidazole-4-carboxamide 1-ribofuranoside (AICAR) was purchased from Cell Signaling Technology, Inc. (Beverly, USA).
3
Mouse Preosteoblastic Cell Culture
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The mouse preosteoblastic cell line MC3T3-E1 (purchased from RIKEN, Saitama, Japan) was cultured in alpha minimum essential medium (Gibco BRL, Rockville, MD, USA) supplemented with 10% heat-inactivated fetal bovine serum and 1% antibiotic solution (Nacalai Tesque, Kyoto, Japan) at 37℃ and 5% CO2.
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