Antibodies are large molecular mass glycoproteins and, therefore, can function very effectively as immunogens.
There are 3 categories of epitopes (or antigenic determinants) associated with an Ig molecule.
Isotypic Determinants
These are CL and CH region determinants.
These epitopes distinguish heavy chain isotype and light chain type.
When an antibody from one species is injected into another species, isotypic determinants will be recognized as foreign and anti-isotypic antibody will be produced.
example: Goat anti- mouse IgG
Allotypic Determinants
These determinants are also CL and CH region determinants.
These determinants are present due to subtle amino acid differences encoded by different alleles for the different Igs.
An antibody to allotypic determinants can be produced by injecting antibodies from one member of the same species into a member of the species who possesses a different allotype of Ig.
[Also may be produced by mothers during pregnancy in response to paternal allotypic determinants on fetal Ig]
[May also be produced following blood transfusion]
Idiotypic Determinant
These determinants are VH and VL region determinants
Each determinant is referred to as an idiotope.
It can be the actual antigen binding site itself or a determinant outside of the antigen binding site.
The sum of all idiotopes of an Ig = idiotype
To get an anti-idiotypic antibody you must minimize isotypic and allotypic differences.
You have to use a syngeneic recipient or even the same animal.
Anti-idiotypic regulation of the antibody response is one mechanism by which antibody production can be controlled during a humoral response.
Dogma in biology for many years - 1 gene, 1 polypeptide, entrenched in the mind set of investigators.
The enigma of the variable and constant regions of the heavy and light chain puzzled people tremendously.
In 1965 - Dreyer and Bennett proposed that there must be at least two genes for one polypeptide chain in antibody formation. They proposed that there must be 100s or even thousands of different genes for the V regions but only 1 C region gene for each isotype.
In 1976, Tonegawa and Hozumi were the first to provide the experimental proof of the Dreyer and Bennett hypothesis.
Tonegawa and Hozumi analyzed the light chain genes from Myeloma cell DNA and compared the organization to that of light chain genes present in Whole mouse embryonic DNA.
Finding: In differentiated cells able to produce antibodies (i.e. myeloma cells and B lymphocytes) the V and C gene segments are brought together to form a V-C segment.
In other words, the Ig genes undergo gene rearrangement during B cell development. Ig gene rearrangement has been extensively studied and is known to be the primary mechanism for the generation of Ig diversity in humans and mice. These recombination events are the only known form of site specific DNA rearrangement in vertebrates.
Quote from Philip Leder (an important investigator in this field of research).
"The genes ultimately specifying the structure of each antibody are not present as such in germ cells, or in the cells of the early embryo. Rather than harboring a set of complete and active antibody genes, these cells contain bits and pieces of the genes; a kit of components.
The components are shuffled in the cells of the immune system called B lymphocytes as those cells develop and mature. The shuffling can lead to a different result in each of millions of lines of cells. Individual mutations amplify the diversity. The result is that in the mature descendents of each line, a unique gene is assebled, whose information is expressed in the form of unique antibody."
Example of Kappa Light Chain of Mouse
~300 V (variable) gene segments
one C segment
five J (joining) segments (one of these five is a non-functional pseudogene).
During B cell development, one V region segment is rearranged to join with one J region segment.
In the germline DNA, the kappa light chain genes are arranged as follows:
5'---L-Vk1---L-Vk2------L-Vkn-------Jk1-Jk2-Jk3-Jk4-Jk5-----Ck--3'
V-J joining forms the light chain variable region.
J segments are an extra set of gene segments separated in the genome from V segments.
The V segments and J segments are joined to form the functional V region of the L chain gene.
The J chain codes for last 13 amino acids in the variable region.
In addition, there are leader sequences just before each VL gene segment. The leader sequences are transcribed and translated but are excised from the protein before it is secreted.
During B lymphocyte development, DNA rearrangement occurs. One VL and one JL gene segment come together in the genome and the intervening sequences are deleted. All other cells of the body have the entire set of Ig genes. Only B lymphocytes rearrange them.
Following gene rearrangement, RNA splicing removes introns joining VLJL to C to produce the functional mRNA.
Remember:
introns - intervening sequences removed by RNA splicing
exons - sequences which remain to form mRNA
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