³H-Radiolabeling Service

The service for ³H-radiolabeling offers quantitative information necessary for determining the absorption, distribution, metabolism, and excretion (ADME) profile of an investigational compound, which is essential for the progression of development programs. Alfa Cytology has the customized radiochemistry capabilities together with advanced synthesis to perform the ³H-labeling of a wide range of molecular entities, and to provide highly pure, well-characterized reagents appropriate for important preclinical studies.

Introduction to ³H Isotopes

Tritium, abbreviated as ³H or T, is a radioactive isotope of hydrogen. It has a nucleus that comprises one proton and two neutrons. It has a long, 12.3-year half-life and decays by emitting a beta particle (β^-) of low energy. ³H is not used for in vivo imaging since its beta radiation does not have enough energy to penetrate tissues. Because of its hydrogen chemical identity, ³H is a perfect and highly stable tracer in laboratory-based applications, and, thus, it has become indispensable in quantitative in vitro receptor binding assays and preclinical metabolic studies.

³H Labeling for Pharmaceutical Research

The ³H-radiolabeling service offers a definitive approach to the quantitative assessment of the behavior within biological systems. With the introduction of a tritium isotope, the absorption, distribution, metabolism, and excretion (ADME) profile can be fully delineated. The pharmacokinetic and metabolic information generated is essential to the complete disposition understanding of a molecule and forms the basis of the integral data that informs critical preclinical drug development decisions.

Fig 1. Schematic of a ³H-labeled molecule interacting with a target cell.

Advantages of ³H Labeling

Our Services

Alfa Cytology offers top-tier ³H-radiolabeling services, which facilitate the accurate tracking of compounds across a range of various research scopes. The service offers a comprehensive study of pharmacokinetics, metabolism, and environmental fate of compounds with high sensitivity, precision, and consistency.

Custom ³H Conjugation Service

Alfa Cytology delivers custom ³H conjugation services, generating precisely radiolabeled small molecules and biomolecules tailored to defined study objectives and research workflows.

Small-Molecule Radionuclide Conjugate (SMRC)

Antibody-Radionuclide Conjugate (ARC)

Peptide-Radionuclide Conjugate (PRC)

siRNA-Radionuclide Conjugate

Workflow for Developing ³H-Radiolabeled Small Molecule Conjugates

³H Custom Small Molecule Labeling

• Strategy & Reagent Selection: This process starts with an in-depth examination of a target molecule's structure in order to determine the best positions for attachment, optimizing for stability and biological function. Suitable tritiated precursors are chosen next, focusing on specific activity and synthetic accessibility for a well-defined labeling approach.
• Precursor Synthesis & Characterization: This step consists of preparing the non-radioactive precursor molecule and characterizing it wholly analytically. This key step validates the suitability of the precursor for the following tritium incorporation reaction.
• Protocol Optimization: Focus on the crucial parameters and systematically adjust reagent quantities, reaction duration, and temperature to obtain the best radiochemical yield and the targeted specific activity.
• Custom Radiosynthesis: Labeling reactions are conducted in specialized, controlled environments. For safety and product consistency, state-of-the-art equipment and safety protocols are employed during labeling reactions.

Purification & Quality Control

• Purification & Isolation: For purification and isolation of the target ³H-labeled product, use methods such as high-performance liquid chromatography (HPLC).
• Structure Confirmation: To confirm the product structure, compare to a "cold" (non-radioactive) reference standard and use mass spectrometry (MS) and Nuclear Magnetic Resonance (NMR) techniques.
• Quality Control (QC) Certification:
  • Chemical Purity: Evaluates chemical purity and molecular integrity using analytical techniques such as HPLC-UV.
  • Radiochemical Purity: Assessed using radio-HPLC and radio-SEC-HPLC to confirm the absence of free radionuclide and high molecular weight aggregates.
  • Stability Analysis: Predicts long-term shelf-life from accelerated stability studies under stressed conditions, such as high temperatures. To confirm expiration dating and validate proposed storage conditions, conduct real-time stability studies on the final formulated conjugate.
  • Specific Activity: The specific activity is accurately determined through a joint approach of spectrophotometric protein quantification and radiometric detection.

In Vivo Study

At designated time points after administration, make sure to gather the complete suite of biological samples as outlined below:

• Blood/Plasma: For pharmacokinetic (PK) analysis.
• Urine & Feces: For mass balance and evaluation of potential pathways for excretion.
• Bile: To assess hepatobiliary excretion for selected species.
• Tissues: At the terminal timepoint of the preclinical studies for quantitative whole-body autoradiography (QWBA).

Bioanalytical Sample Analysis

• Total Radioactivity Measurement: Accurately measure the total ³H radioactivity in all biological samples using a liquid scintillation counter (LSC). This data is fundamental for calculating mass balance (recovery) and constructing PK profiles.
• Metabolite Profiling: Analyze extracts from plasma, urine, and feces using HPLC with in-line radio-detection. This generates a "radiochromatogram" that reveals the parent drug and the relative abundance of all major metabolites.
• Metabolite Identification: Integrate the radio-chromatography data with high-resolution mass spectrometry (LC-MS/MS) to interpret the chemical structures of the major metabolites identified.
• Quantitative Whole-Body Autoradiography (QWBA): In preclinical studies, freeze the animal carcass, prepare thin whole-body sections, and use phosphor imaging to visualize and quantify the ³H signal, creating a detailed map of the drug's distribution across all organs and tissues.

Applications of ³H-Radiolabeling

The use of ³H radiolabeling has proliferated due to its versatility and high sensitivity. Across the fields of chemistry, biology, pharmacology, and environmental science, its applications continue to grow. After the incorporation of tritium at hydrogen sites, organic compounds can be assessed and traced quantitatively through various intricate systems.

At Alfa Cytology, we provide ³H radiolabeling service through our unique radiochemistry capabilities and integrated radiochemistry R&D workflows for method development, tracer synthesis, and optimization. Thorough, study-driven support fuels research in pharmacokinetics, metabolism, and environmental fate. For more information, feel free to reach out.

For research use only. Not intended for any clinical use.