Anesthetized animals were perfused with PBS for 2 min, followed by perfusion with EM fixative (2.5% glutaraldehyde (Sigma, G5882) and 0.1 M sodium cacodylate (Sigma, C4945) in 4% PFA (Sigma, P6148), pH adjusted to 7.4) for 2 min. The tissue was post-fixed in EM fixative for 2 h, followed by decalcification with 1:1 1 M sodium cacodylate buffer (pH 7.4) and 0.5 M EDTA (pH 8) at room temperature for five days with daily exchange of the decalcification solution. Fixed penile tissue was embedded in a gelatin-albumin matrix for vibratome sectioning. The coronal penile sections (200 µm) were kept in sodium cacodylate buffer until processing. For SEM imaging, vibratome sections were washed in 0.1M sodium cacodylate buffer (pH 7.4), post-fixed in 1% w/v unbuffered OsO4 for 1 h, washed three times in water, dehydrated in EtOH (20–100%, 20% steps) and then EtOH–acetone (70:30, 50:50, 30:70, 100%; 15 min steps), critical point dried in an Agar E3000 critical point dryer (Quorum Technologies Ltd., East Essex, UK), mounted on stubs and coated with Au with an Emitech K550X sputter device (Emitech Ltd., Kent, UK) before examining using a Philips XL 30 ESEM (Philips, Eindhoven, Netherlands) operated at 15 kV accelerating voltage. Images were recorded digitally.
Agar e3000 critical point dryer
Manufactured by Quorum Technologies
Sourced in United Kingdom
The Agar E3000 is a critical point dryer, which is a laboratory equipment used to dry samples while preserving their physical structure. The device utilizes a process called critical point drying to remove liquid from the sample without causing damage or deformation.
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Lab products found in correlation
2 protocols using agar e3000 critical point dryer
1
Penile Tissue Preparation for SEM
2
Scanning Electron Microscopy of Enzymatic Hydrolysis
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Scanning electron microscopy (SEM) was used to follow the progressive hydrolysis of TW G-layers as well as any hydrolytic effects of the cellulase complement on the cellular structure of other cell wall layers (i.e., secondary cell walls/primary, middle lamella regions). Entire semi-thin sections of stems after enzymatic treatment and controls were fixed in 3% v/v glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.2) for 3 h at room temperature. After washing in the same buffer (3 × 20 min), sections were post fixed in 1% w/v osmium tetroxide for 1 h. Following washing in water (3 × 30 min; and overnight), sections were dehydrated in a progressive series of ethanol [16 (link)] and finally acetone. Samples were dried in an Agar E3000 critical point dryer (Quorum Technologies Ltd., East Essex, UK) using liquid CO2 as the drying agent, mounted on stubs, and coated with gold using an Emitech K550X sputter device (Emitech Ltd., Kent, UK) [16 (link)]. Observations were made using a Philips XL 30 ESEM (Philips, Eindhoven, Netherlands) operated at 10–20 kV with images recorded digitally.
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