Overview
ISO/TS 7833:2024 - Nanotechnologies - Extraction method of nanomaterials from lung tissue by proteinase K digestion - specifies an enzymatic extraction method using proteinase K (PK) to recover nanomaterials deposited in lung tissue. The Technical Specification describes the advantages of PK digestion over aggressive chemical digestion (reduced particle dissolution and structural damage) and focuses on separating particulate nanomaterials from their ionic counterparts for subsequent instrumental analysis.
Key topics and technical requirements
- Scope: Extraction of nanomaterials from lung tissue and separation of nanoparticles from dissolved ionic species; method is applicable to particles that remain insoluble during PK digestion.
- Optimal PK system:
- Recommended digestion buffer (tested formulation): 30 mM Tris‑HCl, 10 mM EDTA, 1% SDS, 5 mM CaCl2, pH 8.0.
- Optimal PK activity reported in the document: 10 µg PK (≈ 0.2–0.3 U) under the described test conditions.
- Incubation condition example: 56 °C ± 1 °C (heat block or water bath), typical digestion times in tests were 24 h.
- Sample preparation & equipment:
- Drying/weighting of lung tissue, slicing (~2 mm), homogenisation (micro ball mill), optional lyophiliser.
- Vessels resistant to protein adsorption (1.5–50 ml tubes), Petri dishes for drying.
- Centrifugation capable of ≥21 000 g (microcentrifuge/ultracentrifuge) to pellet nanoparticles; bath sonicator (≈400 W, 40 kHz) for dispersion.
- Procedure outline:
- Sampling and drying, homogenisation/powderisation, PK digestion of tissue, collection of nanoparticle pellet and preparation for instrumental analysis.
- Methodological considerations:
- Guidelines on separating particulate and ionic fractions, nanomaterials types applicable (e.g., carbon-based materials and certain metal oxides), and the impact of blood content or organ-specific variability.
Applications and users
- Primary applications:
- Lung burden analysis in inhalation toxicology and toxicokinetic studies.
- Recovery and characterization of retained nanomaterials prior to ICP‑MS, sp-ICP‑MS, TEM, EDS, UV‑Vis or other instrumental analyses.
- Who should use this standard:
- Nanotoxicology laboratories, regulatory testing labs, contract research organisations (CROs), materials scientists, and researchers conducting inhalation or organ-distribution studies of engineered nanomaterials.
Related standards and guidance
- ISO 80004‑1 (Nanotechnologies - Vocabulary)
- ISO/TR 22019 (toxicokinetic considerations for nanomaterials)
- OECD inhalation guidance (TG 412, TG 413) and ongoing toxicokinetic guidance initiatives (related to TG 417 discussions)
ISO/TS 7833:2024 provides a practical, validated PK-based protocol to extract particulate nanomaterials from lung tissue while minimising artefacts from chemical digestion - a key tool for accurate lung burden and toxicokinetic assessments.