The production of antibodies (Abs) in chickens and the extraction of specific Abs from egg yolk (IgY Abs) are increasingly attracting the interest of the scientific community, as demonstrated by the significant growth of the IgY literature. This review offers detailed and comprehensive information about IgY-technology, including: a) possibilities for hen keeping in accordance with the Three Rs principles; b) new insights into the IgY transfer mechanism from blood to yolk as a biological basis for the technology; c) the comparative characteristics of IgY Abs and IgG Abs; d) the high efficacy of the technique, in view of the extraordinary amount of IgY Ab produced by one hen in one year (between 20g and 40g IgY in total); e) comparisons between the efficacies of IgY Abs and IgG Abs (rabbit, sheep, mouse) in several immunological assays; f) immunisation protocols, as well as the most commonly used IgY-extraction procedures; g) new possibilities for application in human and veterinary medicine, including strategies for the treatment of Helicobacter pylori infection or fatal intestinal diseases in children, particularly in poor countries, for reducing the use of antibiotics, and, in Asia and South America, for producing Abs against snake, spider and scorpion venoms; and h) the use of IgY Abs in various fields of research, also taking into consideration recent developments in South America (particularly Argentina and Cuba) and in Asia.

Hens can be immunized by means of i.m. vaccination (Musculus pectoralis, left and right, injection volume 0.5-1.0 ml) or by means of Gene-Gun plasmid-immunization. Dependent on the immunogenicity of the antigen, high antibody-titres (up to 1:100,000 -1:1,000,000) can be achieved after only one or 3 -4 boost immunizations. Normally, a hen lays eggs continuously for about 72 weeks, thereafter the laying capacity decreases. This protocol describes the extraction of total IgY from egg yolk by means of a precipitation procedure (PEG. Polson et al. 1980). The method involves two important steps. The first one is the removal of lipids and the second is the precipitation of total IgY from the supernatant of step one. After dialysis against a buffer (normally PBS) the IgY-extract can be stored at -20°C for more than a year. The purity of the extract is around 80 %, the total IgY per egg varies from 40-80 mg, dependent on the age of the laying hen. The total IgY content increases with the age of the hen from around 40 mg/egg up to 80 mg/egg (concerning PEG precipitation). The laying capacity of a hen per year is around 325 eggs. That means a total potential harvest of 20 g total IgY/year based on a mean IgY content of 60 mg total IgY/egg (see Table 1).

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Polyclonal antibodies, widely used in research and diagnostics, are conventionally isolated from the blood of immunised mammals, especially rabbits. The fact that antibodies can also be detected in the yolk of eggs laid by immunised hens, led to the development of yolk antibody technology (IgY-technology) as an alternative method that is less stressful to animals. This technology has become a worthwhile alternative to the blood-dependent techniques. Furthermore, because of the phylogenetic distance between birds and mammals, avian antigens have a very specific immune response to highly conserved antigens of mammals, such as human erythrocyte antigens. To evaluate the humoral immune response of hens immunised with human red erythrocyte antigens, 22 White Leghorn hens were kept in cages and immunised with total red blood cells or stroma of the human rhesus positive (Rh+) system (D antigen) by weekly intramuscular and intravenous injections, without the use of an adjuvant. The haemag-glutination assay was used to evaluate the dynamics of the production of IgY antibodies against human erythrocyte antigens, and single radial immunodiffusion was used to evaluate the amount of total IgY in de-lipidated supernatants from egg yolk. The highest titres were observed four weeks after the first immunisation, and these remained stable for up to seven weeks for the intravenous route. Positive reactivity against human erythrocyte antigens A, B and O was demonstrated in de-lipidated supernatants from the egg yolks of immunised hens. The strongest reaction was observed against blood group O Rh+ (O+). Furthermore, the antibodies obtained recognised erythrocyte antigens that belong to neither the Rh system nor the Landsteiner-Weiner antigen. This finding opens the possibility of using hens instead of rabbits to produce polyclonal antibodies for human haemoclassification.

In the present work, a label immunoquantification procedure was developed in order to determine the number of markers introduced into DNA. A non-enzymatic, in vitro labeling method for introducing the p-bromobenzoyl radical (label), through transamination and acylation reactions of the cytidine nucleotides in calf thymus DNA, was carried out. Three spacer arms with different lengths were used for separating the label from the nucleotide and three labeled DNA were obtained. Anti-p-bromobenzoyl chicken IgY polyclonal antibodies were obtained. The antibodies detected the label, into three-labeled DNA, with different sensitivities, in relation to spacer arm length used. About 3-11 labels per 4 x 10(6) bases into thermally denatured DNA were immunoquantified.