Autoradiograph

Autoradiography or radiography ( common abbreviation AURA ) refers to the visualization of a chemical component by radioactive isotopes, originally by blackening of a photographic film, now increasingly using a radiation detector. The recording obtained is called the autoradiogram.

Applications

Autoradiography was for three decades to be an integral part of the DNA sequence analysis as described by Sanger. Since the last years of the 20th century, however, increased fluorescent are used instead of radiolabeled DNA nucleotides for sequence analysis.

Autoradiography is furthermore used for the production of recombinant proteins, analysis of enzymatic reactions, or identification of the enzyme substrates.

Moreover, it is applied in pharmacokinetics to create eg Liberation Absorption Distribution Metabolism Excretion studies ( LADME ).

The combination of autoradiography and used as a neutron activation Neutronenautoradiografie to examine local elementary compositions in paintings.

Implementation

All applications autoradiography common is the need to inject radioactive isotopes in the molecules to be analyzed. Due to the frequent occurrence of the corresponding stable isotope in biomolecules, and their relatively low hazard, the radioisotopes 14 C (carbon), 35S (sulfur), 32 P (phosphorus ), and 3 H (tritium ), are often used.

Example 1 - Marking of recombinant proteins

A bacterial strain transformed with an expression plasmid is combined with a nutrient medium containing the radioactively labeled amino acid 35 S-methionine. 35S -methionine is incorporated into the recombinant protein encoded on the plasmid. A bacterial extract is separated by SDS-PAGE, the gel is dried, and a film is placed. For " exposure " of the film by the outgoing of the isotopes beta ray is displayed on the developed film, the position of the recombinant protein.

Example 2 - Analysis of enzyme activity

ATPase, ie an enzyme which cleaves the nucleotide ATP, is incubated with radiolabeled 32P ATP. The mixture is separated by thin layer chromatography after different times, the chromatographic plate is dried and placed on film. The density of the film by means of 32P - phosphate reflects the activity of the enzyme.