AUTORADIOGRAPHY

AUTORADIOGRAPHY Aswini Krishna.N.K.AUTORADIOGRAPHYA method used to locate radioisotpe labeled materials which have been seperated in gel or present in blots.The location of radiolabeled material is determined by overlying the test material with a photographic film that is sensitive to the radioisotope.PRINCIPLE When a radiation emitted by the radiolbeled material interacts with the photographic film the silver ions present in the film is coverted into metallic silver and thus forming the image. Autoradiography Incubate tissue withradioactive ligandExpose to filmor emulsionIsotope will emit radiation (usually beta)Radiation will hit silver grains in emulsion and expose them Suitable isotopesWeak beta emitting particles are used.Low energy of the negatrons produce short track length and a discrete image will result.

Emulsion or FilmPhotographic emulsion composed of gelatin to which silver halide crystals are embedded.Usually silver bromide crystals are used.These emulsion are very sensitive to radioisotopes.

PHOTOGRAPHIC IMAGEIf the charged particles are passed through the film they are liable to interact with silver bromide crystals.By ionisation ,silver bromide crystal has acquired additional electrons and some silver ions become silver atoms.Thus The LATENT IMAGE is produced .The crystals become easily susceptible to reducing agents which converts more silver ions to silver atoms.Small nuclei of metallic silver constitute the FINAL IMAGE.Direct autographyIn direct autography , the sample is placed in intimate contact with the film.The radioactive emissions produced black areas on the developed autoradiograph.It is suitable for the detection of weak beta emitting radionuclides .But it is not suited to the detection of highly energetic beta particles.

Indirect autoradigraphyThe technique by which emitted energy is converted into light by means of a scintillator,using fluorography.

2 Types: In-vivo autoradiography - receptors are labelled in intact living tissue by systemic administration of the radioligand (PET)In-vitro autoradiography - slide-mounted tissue sections are incubated with radioligand so that receptors are labelled under very controlled conditions

Autoradiography:Uses Map anatomical location of radiolabelled ligands to visualize and quantify receptors in tissueTrace neurons by axonal transport of radioactively labelled amino acids, certain sugars, or transmitter substancesMeasure DNA production (e.g., 3H-thymidine)

Advantages:- Highly specific tool to pharmacologically characterize receptors in tissue.- Provides location of receptor in tissue- Enables characterization of receptors in different tissues between different animals or brain regions- Technically easy

Disadvantages:- There are no biochemical or physiological criteria to assess the binding specificity (i.e., to determine whether the binding site really corresponds to an actual receptor)- The presence of a high-affinity radiolabelled receptor does not necessarily imply that the receptor has physiological significance- Ligands are not always very specific

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