On days 7 and 21, samples were fixed in 2.5% glutaraldehyde (Merck, Gillingham, UK) in 0.1 M cacodylate (Merck, Gillingham, UK) overnight at 4 °C. The samples were then washed with deionized water three times and dehydrated with an ethanol series (20 min each, 50%, 70%, 90% and 100%) (Fisher, Loughborough, UK) and with 1:2 and 2:1 HMDS (hexamethyldisilazane): EtOH and 100% HMDS (overnight) (Merck, Gillingham, UK). The samples were then mounted on stubs and gold-coated (Polaron e500) (Quoram Technologies, Laughton, UK) before imaging at 20.00 kV by SEM.
0.1 m cacodylate
Manufactured by Merck Group
Sourced in United Kingdom
0.1 M cacodylate is a buffer solution commonly used in biological and chemical laboratories. It provides a buffering system for maintaining a specific pH level, typically around 7.2-7.4, which is suitable for a variety of applications involving cellular and molecular studies.
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Lab products found in correlation
4 protocols using 0.1 m cacodylate
1
SEM Sample Preparation Protocol
2
Sample Preparation for SEM Imaging
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On days 7 and 21, samples were rinsed with 0.1 M cacodylate buffer and fixed with 2.5% glutaraldehyde in 0.1 M cacodylate (Merck Life Science, Gillingham, UK) overnight at 4 °C. Following fixation, the samples were washed 3 times with deionised water and dehydrated in an ethanol series (20 min each at 50%, 70%, 90%, and 100%) followed by treatment with 1:2 and 2:1 HMDS (hexamethyldisilane): EtOH (20 min) and 100% HMDS (overnight) (Merck Life Science, Gillingham, UK). The samples were then mounted on stubs and gold- sputtered coated using a Polaron e500 (Quorum Technologies, Lewes, UK). Imaging was performed with an SEM with an accelerating voltage of 20 kV (EVO ma10, Zeiss, White Plains, NY, USA).
3
PLLA/PLGA Scaffold Characterization
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PLLA/PLGA scaffolds with and without cells were scanned in the Quanta 200 microscope (FEI). Cell-seeded scaffolds were fixed in 2.5% glutaraldehyde in 0.1 M cacodylate (Sigma-Aldrich) for 5 min and dehydrated with increasing ethanol (GADOT-GROUP) gradient of 70, 85, 95, and 100% for 5 min each. The scaffolds were then soaked in hexametyldisilazane (Sigma-Aldrich) for 5 min and, afterward, were left to air dry ON at RT. Before the scanning, all the samples were coated with a gold-palladium mixture.
4
Scaffold Surface Characterization by SEM
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After 1 week of culturing, the scaffolds were prepared as described previously [5]. Briefly, the scaffolds were washed once with PBS solution and fixed with 2.5% glutardialdehyde (75% solution in water, Sigma-Aldrich, Germany) buffered with 0.1 M cacodylate (Sigma-Aldrich, Germany), pH 7.4, for 2 h at 4 °C. The samples were then dehydrated by passing through a series of increasing concentration of ethanol (25, 50, 75, 95% each for 15 min). The analysis was performed on a LEO Zeiss Gemini 1530 scanning electron microscope (SEM).
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