MicroCT protocols for scanning of embryos and juvenile Hexaplex trunculus
Sep 03, 2021

MicroCT protocols for scanning of embryos and juvenile Hexaplex trunculus

  • 1Hellenic Centre for Marine Research
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only and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to protocols.io is not peer reviewed and may not have undergone a formal approval of any kind.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.
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 embryos and juveniles of the marine gastropod species Hexaplex trunculus.
 
Eva Chatzinikolaou, Kleoniki Keklikoglou 2021. MicroCT protocols for scanning of embryos and juvenile Hexaplex trunculus. protocols.iohttps://dx.doi.org/10.17504/protocols.io.bxw4ppgw
ECCO project, Hellenic Foundation for Research and Innovation ( HFRI)
Grant ID: project ID 343
microCT, gastropods, embryos, juveniles, scanning, Hexaplex trunculus
 protocol  ,  
Sep 02, 2021
Sep 03, 2021
 
52924
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only and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to protocols.io is not peer reviewed and may not have undergone a formal approval of any kind.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.

Sample preparation

42m
1
10m
Removal of embryos of Hexaplex trunculus from their egg capsules.
2
30m
Anesthetization of embryos and juvenile gastropods with 7% MgCl2.
3
2m
Sample placement inside a plastic white pipette tip without any scanning medium (in the air).

microCT scanning

2h 23m
4
2h 23m

SkyScan 1172
micro-computed tomographer (microCT)
Bruker
10L01170
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


4.1
2h 23m
Scanning parameters for embryos and juvenile Hexaplex trunculus

Voltage: 59 kV
Current :167 μA
Filter: none
Pixel size: 2 μm
Camera binning: 1 × 1
Exposure time: 325 ms for embryos and 316 ms for juveniles
Rotation: 360° for embryos and 180° for juveniles
Rotation step: 0.20º for embryos and 0.25° for juveniles
Frame averaging: 3 for embryos and 5 for juveniles

Images reconstruction

20m
5
20m
Projection images were reconstructed into cross sections using the SkyScan’s NRecon software (Bruker, Kontich, Belgium)

5.1
2m
Upload projection images
5.2
2m
Perform X-Y alignment
5.3
16m
Reconstruction parameters

Smoothing: 2
Ring artifact correction: 20
Beam hardening correction: 59%
Attenuation coefficients: 0 - 1.127 for embryos and 0 - 1 for juveniles
Save as: 16-bit TIFF images