IMMUNOLOGICAL ANALYSIS OF THE MAMMALIAN VISUAL

Project: Research project

Description

The long term objectives of this project are to gain an understanding of
the cellular and molecular mechanisms responsible for the development and
function of the mammalian visual system. Knowledge of these mechanisms is
essential to any understanding of congenital abnormalities in visual system
development and the wide variety of degenerative diseases that can affect
both the visual system and other areas of the central nervous system. To achieve these objectives it is necessary to have methods that allow the
identification of the cell types and the individual molecules of the visual
system. Two recent developments of molecular biology, namely the
introduction of monoclonal antibodies and recombinant DNA molecules, have
provided these methods. Antibodies that can identify the major subclasses
of cells in the rat retina have already been produced. The objectives of this proposal are to produce further antibodies that
recognise more discrete subclasses of cells of retina and visual cortex,
and to use cloned cDNA fragments to study the heterogeneity and development
of cell types in the visual system by using in situ hybridisation. To prepare the monoclonal antibodies, cells or membranes enriched in the
cell type of interest will be used to immunise mice. Spleen cells will be
taken from mice mounting an adequate immune response and fused with a mouse
plasmacytoma. In addition spleen cells will be immunised by in vitro
incubation with antigen and then fused. Hybrids will be selected by
standard techniques and those secreting antibodies detected by a
combination of radioactive binding assays and immunocytochemical assays. The antibodies will be used to study the in vivo development of cells of
the visual system by immunocytochemical analysis of tissue from animals of
various ages. The antibodies will also be used to purify defined
populations of cells by combinations of affinity separations and
fluorescence-activated cell sorting. Purified cells will be used both for
further immunisations and for studies in tissue culture. Monolayer cultures of cells from retina or visual cortex will be prepared
and the cell types identified by labelling with specific antibodies. By
using anatomical and electrophysiological methods the nature and
specificity of synaptic interactions between identified cells will be
studied.
StatusFinished
Effective start/end date5/1/844/30/93

Funding

  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health: $115,507.00
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health

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Antibodies
Retina
Visual Cortex
Spleen
Monoclonal Antibodies
Recombinant DNA
In Situ Hybridization
Molecular Biology
Immunization
Central Nervous System
Complementary DNA
Cell Culture Techniques
Cell Membrane
Antigens