Age Estimation from Teeth in Longman’s Beaked Whales (Indopacetus Pacificus) Stranded in New Caledonia (South Pacific)

At a Glance

Metadata	Details
Publication Date	2021-05-01
Journal	Archimer (Ifremer)
Authors	Christina Lockyer, Claire Garrigue
Institutions	Écologie Marine Tropicale des Océans Pacifique et Indien
Citations	1
Analysis	Full AI Review Included

6CCVD Technical Analysis: Precision Material Sectioning for Advanced Biological Microscopy

Research Paper: Age Estimation from Teeth in Longman’s Beaked Whales (Indopacetus pacificus) Stranded in New Caledonia (South Pacific)

This analysis translates the high-precision requirements of biological hard-material sectioning (tooth age estimation) into specific material and engineering solutions offered by 6CCVD. The reliance on low-speed circular diamond saws for thin sectioning underscores the necessity of high-quality Single Crystal Diamond (SCD) and Polycrystalline Diamond (PCD) materials for specialized tooling applications.

Executive Summary

This research highlights the critical role of ultra-precise material processing and cutting technology in high-resolution bio-analysis, achieved through the following technical parameters:

Precision Sectioning Requirement: Successful age estimation relied on generating ultra-thin sections, specifically 150 µm (untreated) and 10-25 µm (decalcified), from highly mineralized teeth.
Essential Tooling: Sectioning was performed using an Isomet low-speed circular diamond saw, demonstrating the mandatory nature of diamond materials for minimizing sample damage in hard tissue preparation.
Micro-Structural Resolution: Growth Layer Groups (GLGs) were successfully resolved, with cementum layers measuring 10-15 µm and dentine layers measuring 25 µm, requiring pristine optical quality in the prepared sections.
Optical Clarity Demand: Examination utilized both plain and polarized transmitted light microscopy, mandating ultra-flat and highly parallel section surfaces, achievable only with superior material finishing protocols.
6CCVD Value Proposition: The exacting demands of low-damage, precision cutting directly align with 6CCVD’s expertise in providing custom SCD/PCD material for high-performance, ultra-wear-resistant cutting tools and precision optical platforms.
Outcome Optimization: The stained, decalcified 10-25 µm sections provided higher, clearer GLG counts, proving that maximizing material processing quality leads to superior scientific data integrity.

Technical Specifications

The following parameters derived from the paper illustrate the extreme requirements for precision material preparation, directly relevant to diamond cutting tooling design:

Parameter	Value	Unit	Context
Sectioning Tool Type	Low-Speed Circular	Diamond Saw	Used for initial thin sections and thick wafering
Untreated Thin Section Thickness	150	µm	Standard preparation method 1 (Polarized Light)
Thick Wafer Thickness	2.0 - 2.5	mm	Intermediate section for Method 2 (Decalcification)
Decalcified Thin Section Thickness	10 - 25	µm	Final preparation method 2 (Stained/Plain Light)
Dentine GLG Layer Thickness	25	µm	Observed micro-structure resolution
Cementum GLG Layer Thickness	10 - 15	µm	Observed micro-structure resolution (thinnest feature)
Decalcification Solution	RDO™	N/A	Proprietary rapid decalcifying solution
Decalcification Time	Up to 15	hours	Duration required for adequate softening
Freezing Stage Operating Temperature	~ -12	°C	Required for microtome sectioning of soft tissue
Microscope Magnification Range	x7 - x40	N/A	Used for examining thin sections

Key Methodologies

The experiment utilized a two-method preparation approach, both relying on high-precision sectioning to achieve optical analysis compatibility:

Mounting and Wafering: Extracted teeth were cleaned, mounted on wood blocks using standard hobby glue, and subsequently cut using a Buehler Isomet low-speed circular diamond saw.
Method 1: Untreated Thin Sections: Centrally cut sections approximately 150 µm thick were created using the diamond saw. These sections were examined directly using transmitted plain light and polarized light microscopy.
Method 2a: Thick Wafering and Decalcification: A thicker central section (wafer) measuring 2.0-2.5 mm was removed adjacent to the first cut. This wafer was rapidly decalcified in RDO™ solution for up to 15 hours.
Method 2b: Final Thin Sectioning and Staining: The decalcified wafer was then ultra-thin sectioned at 10-25 µm using a portable benchtop freezing microtome (operating at approximately -12 °C).
Staining and Mounting: Sections were stained with Ehrlich’s Acid Hæmatoxylin (EAH), treated with ammonia solution to achieve a purple-blue color, rinsed, and permanently mounted under coverslips using DPX (a distyrine/xylene mixture).

6CCVD Solutions & Capabilities

This research demonstrates a clear requirement for specialized, high-performance tooling capable of consistently producing thin sections (down to 10 µm range) from ultra-hard, mineralized substrates. 6CCVD is uniquely positioned to supply the foundational diamond material necessary for these demanding applications, ensuring minimal sub-surface damage (SSD) and optimal optical quality.

Applicable Materials

The precision and low wear required for the Isomet diamond saw blades used in this research necessitate the highest quality diamond material. 6CCVD offers the base material for superior tooling:

Industrial Grade PCD Substrates: Ideal for manufacturing durable, highly uniform circular saw blades and grinding wheels required for reproducible sectioning of dense mineralized tissues. Our PCD material guarantees exceptional abrasion resistance and thermal stability during low-speed cutting.
Optical Grade SCD (Single Crystal Diamond): While the paper utilized standard tooling, researchers extending this work into nanoscale analysis (e.g., using Focused Ion Beam milling on specific GLG layers) would require optically perfect SCD platforms for substrate mounting or window applications. 6CCVD delivers SCD with Ra < 1 nm polishing for superior transmitted light microscopy, even at high magnification.
Custom BDD (Boron-Doped Diamond): For applications that require electrochemical analysis of tissue layers or require specific resistivity for advanced imaging, BDD substrates can be tailored to specification.

Customization Potential

6CCVD’s core capabilities directly address the limitations and specific dimensions required by precision sectioning science:

Requirement (Paper Context)	6CCVD Solution	Technical Capability
Tooling Material Source	SCD/PCD Material Supply	Plates/wafers up to 125 mm for custom blade manufacturing.
Ultra-Thin Sectioning	Substrate Thickness Control	SCD/PCD materials supplied at precise thicknesses, from 0.1 µm to 500 µm, ensuring consistency for tool fabrication.
High Clarity Surfaces	Precision Polishing	Guaranteed polishing quality: Ra < 1 nm (SCD) and Ra < 5 nm (Inch-size PCD), critical for transmitted polarized light analysis.
Advanced Sample Integration	Custom Metalization	In-house capability for depositing Au, Pt, Pd, Ti, W, Cu layers onto diamond substrates, supporting SEM/TEM preparation or integrating sections into electronic testing rigs.
Complex Geometries	Laser Cutting Services	Precision machining of diamond materials for customized tool inserts, specific apertures, or non-standard blade geometries used in microtome or wafering equipment.

Engineering Support

6CCVD recognizes that successful age estimation and micro-analysis of ultra-hard materials like bone and teeth rely heavily on optimized material properties and processing recipes.

Our in-house PhD team can assist researchers and engineers in selecting the optimal diamond substrate specifications (grade, orientation, doping) for manufacturing high-end micro-sectioning tools or developing specialized material platforms for similar bio-mineralization or hard tissue micro-analysis projects.
We offer technical consulting on optimal polishing specifications to minimize sub-surface damage, ensuring maximum optical fidelity for examining fine structures such as the 10 µm cementum GLG layers observed in this research.

For custom specifications or material consultation, visit 6ccvd.com or contact our engineering team directly.

View Original Abstract

Seven Longman’s beaked whales mass stranded in New Caledonia in November 2013, of which 4 ultimately died, in a first worldwide event reported for this poorly known Ziphioid species. Teeth were extracted, collected and thoroughly cleaned of gum tissue from 3 females ranging from juvenile to adult and one adult male. These were sectioned (crown-root) and prepared using two different methods and examined under microscope magnification when Growth Layer Groups (GLGs) in both dentine and cement were successfully identified. The methods employed for aging included 1) sectioning centrally at approx. 150 µm through crown and root on an Isomet circular diamond saw and examining under a microscope using both transmitted polarised light and plain light; and, 2) thick sectioning (wafering) at approx. 2.5 mm and subsequent decalcifiation in RDO™ (a proprietory brand, Illinois, USA) and then thin sectioning the wafer at 10-25 µm and staining with Ehrlich’s acid haematoxylin. GLGs were investigated in both dentine (25 micron) and cementum (10-15 µm). Layering was evident in both tissues but higher counts were more evident in thin stained sections of cementum. Although dentinal GLGs in untreated tooth sections have been used successfully for aging in Ziphioid species Hyperoodon ampullatus (Christensen 1973, Feyrer et al., 2020), it is believed this is the first time that teeth have been used for estimating age from GLGs in this tropical species.\n

Tech Support

Original Source

DOI: https://doi.org/10.34331/cpops.2020f010