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Production of Monoclonal antibodies. Antibodies React with specific molecule Antigens B lymphocytes recognize and respond to Ag Each B cell produces a a Ab directed to single epitope( antigenic determinant) Body- different B cells –mixture of Abs to the Ag

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slide2

Antibodies

  • React with specific molecule
  • Antigens
  • B lymphocytes recognize and respond to Ag
  • Each B cell produces a a Ab directed to single epitope( antigenic determinant)
  • Body- different B cells –mixture of Abs to the Ag
  • Polyclonal antibodies- as they recognize and respond to different sites on Antigen molecule
slide3

To produce a clone of cells that can produce Abs that recognizes a single, specific epitope of an antigen

  • Monoclonal antibodies
  • The production of monoclonal antibodies was pioneered by Georges Kohler and Cesar Milstein in 1975.
slide4

http://highered.mcgraw-hill.com/sites/0072556781/student_view0/chapter32/animation_quiz_3.htmlhttp://highered.mcgraw-hill.com/sites/0072556781/student_view0/chapter32/animation_quiz_3.html

hybridoma
Hybridoma
  • Immunize the mouse against the antigen of interest.
  • Multiple immunization is performed .
  • Intraperitoneal injections.
  • Test Bleeds –examine -Abs produce in body
  • If the host –producing Ab, spleen is removed to release B cells.
slide7

In culture –Mouse myeloma cell line+ Bcells from the host

  • Myeloma cells- tumor cells can divide indefinitely
  • In culture PEG ( polyethylene glycol) added
  • Two types of cell fuse- to form HYBRIDOMAS.
  • Separate the fused hybridoma cells.
  • the unfused B cells they die
  • In mixture you have the fused hybridomacells and myeloma cells.
slide8

Myeloma cell lines-Tumor cells

  • Lack an enzyme, so when placed in a culture medium which had HAT.
  • Only fused cells survive as B cells can produce the enzyme.
  • Conclusion- only hybridomas survive
  • Hybridomas are seperated and individually cultured one cell per well.
  • All the cells derived from a single cell are identical .
  • Culture for weeks
  • Screened for the presence of deisrable Abs.
  • Finally, the clones that produce desirable Absare grown in mass culture
  • frozen for storage.
slide9

Uses of monoclonal antibodies

  • Monoclonal bodies have a variety of academic, medical and commercial uses.
  • Antibodies are used in several diagnostic tests to detect small amounts of drugs, toxins or hormones
  • , e.g. monoclonal antibodies to human chorionic gonadotropin (HCG) are used in pregnancy test kits (Biotech, 1989).
  • Another diagnostic uses of antibodies is the diagnosis of AIDS by the ELISA test.
  • Antibodies are used in the radioimmunodetection and radioimmunotherapy of cancer, and some new methods can even target only the cell membranes of cancerous cells (Chaudhari et al, 1994). A new cancer drug based on monoclonal antibody technology is Ritoxin, approved by the FDA in November 1997 (Orrs, 1997).
slide10

Monoclonal antibodies can be used to treat viral diseases, traditionally considered "untreatable". In fact, there is some evidence to suggest that antibodies may lead to a cure for AIDS (P/S/L, 1997).

  • Monoclonal antibodies can be used to classify strains of a single pathogen, e.g. Neisseria gonorrhoeae can be typed using monoclonal antibodies (Wang et al, 1977).
  • Researchers use monoclonal antibodies to identify and to trace specific cells or molecules in an organism, e.g. developmental biologists at tyhe University of Oregon use monoclonal antibodies to find out which proteins are responsible for cell differentiation in the respiratory system (Fratella, 1998).