³²P-Radiolabeling Service

³²P-radiolabeling is a foundational and highly sensitive method in the field of molecular biology, which uses the high-energy beta-emitting isotope Phosphorus-32 to be incorporated into nucleic acids or proteins for tracing and detection in a variety of biological assays. With deep expertise and strict procedures, Alfa Cytology's scientific personnel are positioned to offer dependable, high-purity ³²P-radiolabeling services for your particular proteins, oligonucleotides, and other molecular substrates.

Introduction to ³²P Isotopes

Phosphorus-32 (³²P) is a radioactive isotope used as a sensitive tracer in many areas of life sciences research. ³²P has a half-life of 14.3 days and decays with the emission of a high-energy beta particle that is readily detected. Consequently, ³²P is very useful for labeling a variety of nucleic acids and proteins, such as those used in assays to study protein phosphorylation, kinase reactions, and in hybridized nucleic acids. These nucleic acid hybridization studies can be visualized with autoradiographs and quantitatively analyzed with scintillation counters.

³²P-Labeling for SPECT Imaging

³²P labeling is a fundamental biochemistry method for detecting biological macromolecules, which incorporates radioisotope Phosphorus-32 in them. Enzymatic reactions are commonly used in this procedure. For example, T4 Polynucleotide Kinase is used in the process of 5' labeling of DNA and RNA, or protein kinases to transfer the radiolabeled gamma-phosphate from [γ-³²P]ATP onto a target protein. These reactions generate high-specific-activity probes, which are vital for a variety of downstream processes, including nucleic acid hybridization, kinase assay, and autoradiographic imaging.

Fig.1 Chemical structure of the final 3'-radiolabeled product. (COFSKY J C, et al., 2020)

³²P Radiopharmaceutical

Disclaimer: Alfa Cytology focuses on providing preclinical research services. This table is for information exchange purposes only. This table is not a treatment plan recommendation. For guidance on treatment options, please visit a regular hospital.

Our Services

At Alfa Cytology, skilled biochemists and molecular biologists with years of experience carefully think through every detail of the job in order to provide top-quality and high-specific-activity ³²P radiolabeling services. We have mastered the art of radiolabeling proteins, peptides, and nucleic acids and provide tailored solutions for your trusted, reproducible results.

Custom ³²P Conjugation Service

Expertly performing ³²P conjugation for a wide spectrum of biological molecules, including proteins, peptides, and nucleic acids (DNA, RNA, and oligonucleotides).

Small-Molecule Radionuclide Conjugate (SMRC)

Antibody-Radionuclide Conjugate (ARC)

Peptide-Radionuclide Conjugate (PRC)

siRNA-Radionuclide Conjugate

Workflow for Developing ³²P-Radiolabeled Nucleic Acids Conjugates

³²P Custom Nucleic Acids Labeling

• Strategy & Reagent Selection: For every specific case, a unique labeling strategy is developed whereby a specific ³²P-labeled nucleotide is chosen (e.g. [γ-³²P] ATP is used in the case of end-labeling) and a specific enzyme is used, such as for 5'-end labeling, T4 Polynucleotide Kinase (PNK) is used or for internal labeling by nick translation, the DNA Polymerase is used.
• Substrate Preparation: The target oligonucleotide is designed with attention to secondary structure and possible labeling positions. Then, synthesis or acquisition occurs, followed by preparation in an appropriate nuclease-free buffer to act as the labeling substrate.
• Protocol Optimization: The efficiency and specific activity of radiolabeling of the nucleic acid probe are maximized and accomplished through the finely finessed and perfected protocols of the enzymatic labeling procedure, which optimized key factors like the ratio of enzyme to substrate, the pH of the buffer, temperature, and the time of the enzyme reaction.
• Custom Radiosynthesis: The process of maximizing yield and specific activity is performed under optimized reaction conditions. This includes the enzymatic transfer of the ³²P-radionuclide to the nucleic acid and immediately terminating the reaction afterward.

Purification & Quality Control

The compound is confirmed for successful incorporation of ³²P after labeling. Verification is done using liquid chromatography, mass spectrometry, and scintillation counting to confirm correct isotopic labeling, purity, and stability of the compound. All analytical processes ensure the compound meets biological study standards.

In Vitro Evaluation

In vitro tests are performed to evaluate the biological properties of the ³²P-labeled radiopharmaceutical. This includes studying its interaction with target receptors, cellular uptake, metabolic stability, and other pharmacological properties.

In Vivo Evaluation

The first step in this crucial stage focuses on performing PK studies and quantitative biodistribution analyses in order to calculate the agent's uptake in each organ and its overall clearance. In due course, relevant disease models are subjected to SPECT imaging in order to ascertain the agent's uptake, confirming target engagement, and in vivo blocking studies ruling in the diagnostic capability of the agent.

Applications of ³²P-Radiolabeling

Harnessing the unique theranostic potential of Phosphorus-32 to visualize and eliminate cancer cells with unparalleled precision.

Endowed with immense knowledge in molecular biology, the scientific team in Alfa Cytology specializes in the ³²P enzymatic labeling of nucleic acids and proteins with remarkable precision. Get in touch with us and tell us about your project, and we will devise a specific labeling and purification strategy to produce high-sensitivity probes tailored for your downstream applications.

Reference

1. COFSKY J C, DOUDNA J A. Attachment of a (³²)P-phosphate to the 3' Terminus of a DNA Oligonucleotide [J]. Bio-protocol, 2020, 10(20): e3787.

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