The reaction of the immune lymphocytes with the target antigens can be revealed by three principal assays. In one, graft rejection methods, the lymphocytes trigger a reaction that destroys a foreign tissue or organ transplanted to a recipient. In the second assay, the immune lymphocytes proliferate in response to an antigen: this is called the mixed lymphocyte reaction if the antigen is present in the membrane of a foreign lymphocyte; it is called  e T-Iymphocyte proliferation assay when a soluble antigen is picked up and presented by a specific cell, which is usually genetically identical with the lymphocyte. In the third assay, the cell-mediated lympholysis, assay, the activated lymphocytes differentiate into cytolytic cells which kill appropriate target cells.

There are four genetic situations in which immune cells and antibodies can be elicited. In the xenogeneic situation, the antigen derives from a different animal species than the responding animal. In the allogeneic situation, the donor is one individual and the responder another, genetically disparate individual of the same species. In the syngeneic situation, the donor and the recipient are genetically the same (for example two individuals of an inbred strain).

In the autologous situation, an individual responds to its own antigens. Responses in syngeneic and autologous situations occur only under pathological conditions when the individual fails to distinguish between self and nonself entities and regards its own molecules as foreign antigens (autoimmunity). Molecular methods

In modem immunogenetics research, the serological and histogenetical methods are combined with molecular methods in which the researcher isolates and works with the genes that code for the traits. Usually, the antigens are identified with antibodies or lymphocytes, then messenger ribonucleic acid (mRNA) coding for the antigen-bearing protein is isolated and used to identify the deoxyribonucleic acid (DNA) segment (gene) from which it has been transcribed. The isolated gene, in turn, can be inserted into a suitable cell and activated, and the expressed molecules used to elicit new sets of antibodies or immune cells for a refined analysis of the gene product. This approach of going back and forth from classical to molecular methods has proved to be very successful and has led to the elucidation of several complex genetic systems.

Animal immunogenetics relies heavily on the use of inbred, congenic, and recombinant inbred strains.Inbred lines result, when individuals that are more closely related to each other than randomly chosen individuals mate together, for many generations. In a randomly mating population, each individual is homozygous for most of its genes (carries identical alleles at each of the loci) but heterozygous at some 20 to 40% of loci. Random mating propagates this status, whereas inbreeding tends to increase the proportions of homozygous loci.

The reason for this increase is easy to understand. First, assume that the two parents of an initial mating are both Aa heterozygotes (where A and a are alleles at a given locus). The Aa Aa mating produces AA, Aa, and aa progeny. If brothers are mated with sisters from this litter, sooner or later two AA or two aa individuals will be selected by chance and mated together. When this happens, all the progeny in all subsequent brother x sister matings will always be AA or aa.

Homozygosity has been achieved at the A locus, and as long as the individuals in this genealogical line are not outcrossed to unrelated individuals, the line remains homozygous for this locus. The same thing happens sooner or later for other loci, B, C, D, and so onfor all those loci that were originally in a heterozygous state. The mathematical theory of inbreeding predicts that after 20 generations of brother x sister matings, more than 97% of the loci become homozygous. By convention, this figure is considered sufficient for a strain to qualify as inbred.

When the matings are not between brothers and sisters but between some more distant relatives, it takes proportionally longer to achieve this degree of homozygosity. Because each individual carries a certain load of deleterious genes (that is, genes that in a homozygous state lower the viability of their bearer), and because inbreeding also increases the homozygosity of these genes, an inbreeding depression may set in after a few inbreeding generations. In this phase of the inbreeding program, many individuals either die or do not breed.

However, if the program 'has been initiated on a sufficiently large scale, there are usually enough genetically healthy individuals to carry the program over this hurdle. After 20 generations of brother x sister mating, individuals within a group become genetically homogeneous, resembling one another the way identical twins do; an inbred strain (line) is thus established. A number of inbred strains have been developed in the laboratory mouse, rat, domestic fowl (chicken), guinea pig, Syrian hamster, and rabbit.

The advantage in working with inbred strains rather than outbred animals is that inbred strains restrict the variability of the conditions of an experiment. However, when two strains are compared and it is found that they respond differently to a treatment, it is not known to what gene this difference should be attributed. The strains may differ at as many genetic loci as two unrelated individuals in an outbred population do. To study the effect of single, defined genes, immunogeneticists have developed congenic lines. These lines always come in groups, the smallest group being a pair, which consists of a congenic line and its inbred partner strain.

The two are homozygous at more than 97% of their loci (that is, they are inbred) and are identical except, ideally, at one locusthe locus that is to be studied. At this differential locals, the two strains carry different alleles. This ideal situation is achieved only rarely, if ever; most of the time immunogeneticists are satisfied when the two strains in a pair differ in a short chromosomal segment that includes the differential locus. In addition to pairs, trios, quartets, or quintets of congenic lines can also be produced, in which all the lines differ from the inbred partner strain and from one another in that they carry different alleles at the same differential locus.

The simplest way of producing a pair of congenic lines involves two strains, 1 and 2, that differ at locus 0 (see illus.) Strain 1 is a dd homozygote and strain 2 is a OD homozygote. To study the effect of the 0 gene, a line is produced that is genetically identical with strain 1 but carries the 0 allele. To this end, the two strains are crossed, and the resulting F1 hybrid is backcrossed to strain 1. In the backcross generation, the 0 gene segregates so that half the progeny is Dd and the other half dd.

The animals are then typed and one of the Dd individuals is selected for further mating (backcrossing) to strain 1; this procedure is repeated for several generations. At the end, two Dd individuals are intercrossed and DD animals are obtained. By brother xsister mating of DD animals, a 1.2 line is obtained which is very similar to strain 1, but which carries the D instead of the d allele at the D locus. According to calculations, 99% of the loci no linked to D (that is, on a different chromosome than D) are of strain 1 origin when the backcross is repeated 12 times.

Fig A Scheme of congenic line production by repeated backcrossing to strain 1and selection in each segregating generation for the D allele donated by strain 2. The successive generations of matings of Dd individuals with strain H-1 are designated N₁, N₂, N₃ and so on. After 12 such generations, two Dd individuals are intercrossed and dd individuals are selected and mated among themselves. The intercrossing generations are then designated F₁, F₂, F₃ and so on

A. Generation

The largest number of congenic lines is available in the laboratory mouse. Lines which are known to differ from the inbred partner strain at a single locus (rather than a chromosomal segment) are called coisogenic. They arise when one of the loci in an inbred strain mutates and this changes to another allele. Individuals homozygous for the mutation are then coisogenic with the original strain.