Sep 03, 2021
MicroCT protocols for scanning egg capsules of Hexaplex trunculus
- 1Hellenic Centre for Marine Research
- MicroCT Lab HCMR
Protocol Citation: Eva Chatzinikolaou, Kleoniki Keklikoglou 2021. MicroCT protocols for scanning egg capsules of Hexaplex trunculus. protocols.io https://dx.doi.org/10.17504/protocols.io.bxw5ppg6
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: September 02, 2021
Last Modified: September 03, 2021
Protocol Integer ID: 52925
Keywords: microCT, gastropods, egg capsules, Hexaplex trunculus, scanning
Funders Acknowledgement:
ECCO project, Hellenic Foundation for Research and Innovation ( HFRI)
Grant ID: project ID 343
Disclaimer
DISCLAIMER – FOR INFORMATIONAL PURPOSES ONLY; USE AT YOUR OWN RISK
The protocol content here is for informational purposes only and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to protocols.io is not peer reviewed and may not have undergone a formal approval of any kind. Information presented in this protocol should not substitute for independent professional judgment, advice, diagnosis, or treatment. Any action you take or refrain from taking using or relying upon the information presented here is strictly at your own risk. You agree that neither the Company nor any of the authors, contributors, administrators, or anyone else associated with protocols.io, can be held responsible for your use of the information contained in or linked to this protocol or any of our Sites/Apps and Services.
Abstract
Micro-computed tomography (micro-CT) is a high-resolution 3D-imaging technique which is now increasingly applied in biological studies focusing on taxonomy and functional morphology. The creation of virtual representations of specimens can increase availability of otherwise underexploited and inaccessible samples. This protocol aims to standardise micro-CT scanning procedures for the egg capsules of the marine gastropod species Hexaplex trunculus.
Sample preparation
Sample preparation
1w 5d 1h 4m
1w 5d 1h 4m
Fixation of egg capsules of Hexaplex trunculus in 5% formaldehyde buffered with seawater.
5d
Wash egg capsules with distilled water and dehydrated them with ethanol in gradually increasing concentrations (20%, 50%, 70%, 96%).
1h
Staining of egg capsules with 1% iodine in 96% ethanol.
1w
Washing of stained samples with 96% ethanol.
2m
Placement of samples inside a plastic Falkon tube with 96% ethanol as a scanning medium.
2m
microCT scanning
microCT scanning
3h 38m
3h 38m
Equipment
SkyScan 1172
NAME
micro-computed tomographer (microCT)
TYPE
Bruker
BRAND
10L01170
SKU
LINK
Detail detectability: <1 μm
Low contrast resolution (10% MTF): 5 μm
Pixel size at maximum magnification: <0.8 μm
X-ray source:
Sealed microfocus X-ray tube, air cooled, >10,000h lifetime;
Spot size <5μm @ 4W, 20-100kV, 0-250 μA (10W max)
X-ray detector (camera): 11 Megapixel (4000X2300) 12-bit digital CCD-camera with fibre optic coupling to scintillator
Maximum object size: 50 mm in diameter using offset scan
Radiation safety: < 1 µSv/h at any point on the instrument surface
SPECIFICATIONS
3h 38m
Scanning parameters for embryos and juvenile Hexaplex trunculus
Voltage: 80 kV
Current :124 μA
Filter: aluminium
Pixel size: 13.79 μm
Camera binning: 2 × 2
Exposure time: 1435 ms
Rotation: 360°
Rotation step: 0.40º
Frame averaging: 3
3h 38m
Images reconstruction
Images reconstruction
30m
30m
Projection images were reconstructed into cross sections using the SkyScan’s NRecon software (Bruker, Kontich, Belgium).
30m
Upload projection images.
2m
Perform X-Y alignment.
2m
Reconstruction parameters
Smoothing: 2
Ring artifact correction: 20
Beam hardening correction: 59%
Attenuation coefficients: 0 - 0.064
Save as: 16-bit TIFF images
26m