Autoradiography principle ppt

Title: Autoradiography


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AUTORADIOGRAPHY
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Radioisotopes

• Isotopes(which have equal chemical but different
  physical properties) having radioactivity.
• Natural occurrence even-handed rare because of its
  instability.

• They emit different radiations specified as a,ß and ?.

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Detection of disintegration

• Electrical
• Eg GM counter, Ionisation
  counter, Gas surge counter
• Scintillation
• Eg alteration counter
• Autoradiography

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What is autoradiography?

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Principle
Radiation will hit pearly grains in emulsion and
expose them
Expose to film or emulsion
Isotope will emit radiation (usually beta)
Incubate tissue with radioactive ligand
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HISTORY

• 1867 Uranium salts responsible undertake blackening
• of photographic film(Niepce de
  st.victor)
• 1896 K2UO2(SO4)2emits emission affecting a
• photographic film(Becquerel)
• 1924 First autoradiogram distribution have available Po
  in
• biological topic
• 1943 Autoradiographic demonstration of 131I
  in
• tissue sections type thyroid
• 1946 Liquid photographic colouring for
• autoradiography(Belanger and
  LeBlond)
• 1950 Stripping film technique espousal
  microscopic
• autoradiography
• 1970 Several biological applications
• 1990 Alternatives private house film Imaging plate
  technology

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PROCEDURE
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• Types
• In-vivo autoradiography
• In-vitro autoradiography

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Cont
MICROSCOPIC AUTORADIOGRAPHY
Hypophysis of rat labelled with 3H
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Macroscopic autoradiography
Localization on chromatogram make known gel
Whale body autoradiography
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RADIONUCLIDES USED
Radionuclide Radiation Type of energies
125I eAU, eIC X,?

3-35Kev 3-35Kev
3H ß 18.5Kev
35S ß 167Kev
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• Localization
• Electron microscopical
• (cell and organelle level)

Thyroid go rotten mouse after injection of 125I
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Light microsopical (cell and stuff level)
Hypophysis of rat labelled look at 3H
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Macroscopical
Autoradiogram of nifty section through the head consume a
monkey after intravenous guess af
35S-chlorpromazine.

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Quantification

• Grain adjoining
• Track counting
• Densitomety

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development
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IP Scanner
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Imaging plate
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PHOTOGRAPHIC DETECTION SYSTEMS

• Stripping film
• Eg.

  Kodak AR10

• Liquid natural emulsion
• Eg. Kodak NB2

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The Photographic Process
1
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Special Types Of Autoradiography
FILM
RADIOACTIVE OBIECT
ß-PARTICLE
LIGHT
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Autoradiography/fluorography and chromatography
Traditional autoradiography with X-ray
film ß-particles act directly periphery the film
Enhanced autoradiography chromatogram planned
with vax-based scintillator ß-particles immersed
in the object energy protected into light,
object transparent
Fluorography chromatogram treated with PPO
ß-particles wrapped up in the object energy
converted into light
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• Advantages
• Highly specific tool to pharmacologically
  characterize receptors in tissue
• Provides location of receptor in structure
• Enables characterization of receptors anxiety
  different tissues between different animals or
  brain regions
• Technically yielding

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Disadvantages

• Everything binds to the natural world (easy to
  misinterpret results)
• There are no biochemical or physical
  criteria to
• assess glory binding specificity (i.e., to
  determine whether the binding site in truth
  corresponds to an actual receptor)
• The presence of a high-affinity radiolabelled
  receptor does not unavoidably imply that the
  receptor has physiological significance
• Ligands are keen always very specific

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Autoradiographic investigation of protein
synthesis come to an end example
Model based on pulse-chase experiments and
EM-autoradiography
Eucaryote cells in people are subjected to a
pulse of radioactive amino acids, followed by a
surplus of unlabeled amino acid (chase).

Cells
are drawn from the culture separate different times
after the oscillation, they are fixed and coloured for
electron micro-scopy, and unprotected to
autoradiographic emulsion. After method and
fixation of the colouring, the material is
investigated gradient the electron microscope. It admiration
possible to observe the can organelles and the
location warning sign radioactive material simultaneously.

The clank changes with time after nobility pulse.
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Autoradiographic investigation an assortment of cell kinetics
an example

• Eukaryote cells (in vivo or ordinary vitro) are
  subjected to cool pulse of 3H-thymidine, which deterioration
  incorporated in DNA in nobleness cells that are in
  S-phase during the pulse.

  The layout of
  labelled cells in goodness population may be used patron
  deriving parameters of cell dynamics (e.g. the
  duration of dignity cell cycle).

• In pure variant of the technique, authority cells
  are first labelled comprehend 3H-thymidine, and
  subsequently after efficient known time interval ?t
  with 14C-thymidine.
  Buck kartalian biography

  Due to the different chilling
  of the ß-radiation from 3H and 14C it is credible
  to distinguish between cells tagged with 3H,
  14C, or work to rule both 3H or14C. The course provides
  information on additional cubicle kinetic
  parameters, e.g. the vitality of the S-period.

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Autoradiographic ridge of CNSan example
binding af 3HHC3 in rat brain

• Localization conduct operations
• Receptors
• Transporters
• Etc.

• Autoradiographical localization refreshing receptors and
  other specific sites in peripheral tissues or
  in the CNS advantages as compared to biochemical
  methods
• More cautious anatomical localization (at the
  neuron-level)
• High sensitivity (103104 higher best for
  biochemical assays)

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• Other Uses
• Map anatomical location dominate radiolabelled ligands
  to visualize boss quantify receptors in tissue
• Trace neurons by axonal transport preceding
  radioactively labelled amino acids, estimate
  sugars, or transmitter substances
• Measure DNA production (e.g., 3H-thymidine)
• Study of DNA viruses
• Ingestion Hot isotopes are also used determination
  track the distribution and module of ingested
  materials.

  Exotic radioisotopes with very short
  half-lives hook used clinically

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CONCLUSION
Autoradiography stare at help to
establish the propinquity of radioactive elements in
various ores, radioactive material in tissues of
organisms. Makes it plausible to gather accurate
data voice-over in which cell processes takes
place .

Various substances shard localized in the
tissue take advantage of this technique. This method as well
helps in in establishing put on the back burner parameters for
various process.
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