Autoradiography

Autoradiography of biomolecules

Autoradiography is a class of techniques in which biomolecules are labelled with radioisotopes such that their presence and properties can be detected in an autoradiogram ("self-picture"). Radioisotopes such as ¹⁴C, ³²P, ³⁵S emit radiation that exposes photographic or X-Ray film. Substitution of ³²P into the backbone of DNA, or ³⁵S into the sulfur-containing amino acids allows labelled protein molecules to be detected.

In the particular experiment shown, a DNA sequencing gel is dried onto filter paper, which is placed inside a light-proof X-Ray cassette box (left) and overlain with a sheet of X-ray film. The cassette is sealed shut and left for several hours to several days. The film records the positions of the radio-labelled DNA termination fragments. The DNA sequencing "ladder" is read directly from the autoradiogram.

In other applications, a radio-isotope labelled Southern Blot filter DNA exposes the film, which when developed shows a pattern of black bands that indicate the positions of labelled DNA in the blot. Alignment marks in three of the four corners are made with radio-luminescent ink that also exposes the film. The blot can be aligned with the film by means of the three index marks (with thee marks, only one alignment is possible). In related experiments, the alignment can be used to locate particular DNA bands at precise positions on the blot, which can then be recovered for further experiments.