Staining

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Staining
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Staining is used to look at

• Structure In our case was the cannula in the
  nucleus accumbens?
• Function In our case is there a change in
  dopamine (or indicator of it - Tyrosine
  Hydroxylase) levels? Did we suppress biosynthesis
  of dopamine, less Tyrosine Hydroxylase?

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Cresyl Violet Structure

• Stains are used to color specific components of
  tissue.
• Neuropsychologists stain slices of brain tissue
  so that they can identify fiber tracts and/or
  cell groups for the purposes of determining the
  locations of lesions or of probes.
• Depending on the particular brain structures
  under investigation, a cell stain, a fiber stain,
  or a combined cell and fiber stain may be most
  useful.

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• Cell staining was developed about 100 years
  ago, using basic aniline stains, which combine
  with acidic molecules, such as proteins and
  nucleic acids (e.g. Nissl substance) in the cell
  bodies.
• The general principle underlying these staining
  methods is that the sections are first
  overstained and then the excess stain is removed
  (decolorization). The rate of decolorization
  depends on the pH of the decolorization solution,
  how acidic it is.
• When staining mounted sections, 70 and 95
  alcohol are used to gradually dehydrate or
  rehydrate the sections.
• The purpose of the xylene step is to remove some
  of the lipid substances in the cells and fibers,
  which impair the specificity of the stain.
• Nissl body is attached to the endoplasmic
  reticulum and that only in the cell body
  excluding its edge and around the nucleus.

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Tyrosine Hydroxylase Function
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First some basic immunology
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• The principle function of the immune system is to
  protect animals from infectious organisms and
  from their toxic products. This system has
  evolved a powerful range of mechanisms to locate
  foreign cells, viruses, or macromolecules to
  neutralize these invaders and to eliminate them
  from the body.
• Two types
• Nonadaptive immunity - respond nonspecifically to
  foreign molecules and does not improve with
  repeated exposure to foreign molecules
• Adaptive immunity - is directed against specific
  molecules and is enhanced by re-exposure.
  Lymphocytes - secrete proteins -antibodies -
  antigen.

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• The term immunoglobulin is often used
  interchangeably with antibody. Formally, an
  antibody is a molecule that binds to a known
  antigen, while immunoglobulin refers to this
  group of proteins irrespective of whether or not
  their binding target is known.

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• Antibodies are a large family of glycoproteins
  that share key structural and functional
  features.
• Functionally, they can be characterized by their
  ability to bind both to antigens and to
  specialized cells or proteins of the immune
  system.
• Structurally antibodies are composed of one or
  more copies of a characteristic unit that can be
  visualized as forming a Y shape. Each Y contains
  four polypeptides two identical copies of a
  polypeptide known as the heavy chain and two
  identical copies of a polypeptide called the
  light chain.
• Antibodies are divided into five classes, IgG,
  IgA, IgM, IgD, and IgE, on the basis of the
  number of Y-like units and the type of heavy
  chain polypeptide they contain.

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• Two classes of antibodies can be produced
  experimentally, polyclonal and monoclonal
  antibodies.
• Polyclonal are manufactured by different
  immunologically competent lymphoid cells. As a
  result the antibodies are immunochemically
  dissimilar and will bind at several different
  structural (chemical) parts (called epitopes) of
  the antigen. The animals most frequently used to
  raise these antibodies are rabbits, goats, and
  sheep.

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• Monoclonal antibodies, on the other hand, are
  produced by clones of lymphoid cells (plasma
  cells) the antibodies are therefore
  immunochemically similar and react with a
  specific epitope on the antigen against which
  they are raised. After the immune response has
  been achieved (usually in mice), B-type
  lymphocytes are taken from the spleen and lymph
  nodes and fused with mice myeloma cells to
  produce hybrid (hybridoma) cells. These cells
  can now be propagated either in culture medium or
  by transplanting them into the peritoneal cavity
  of genetically similar mice, where unlimited
  quantities of identical antibodies may now be
  produced.

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• Immunological methods stain an antigen by
  localizing it with a compatible antibody the
  site of the antigen-antibody binding is then
  visualized with an appropriate chromogen marker
  on one of the components of the antigen-antibody
  complex.