In this article we will discuss about Antibodies:- 1. History of Antibodies 2. The Nature of Antibodies 3. Immunoglobulin G or IgG 4. Kinds of Antibodies.
History of Antibodies:
Von Behring and Kitasato in Berlin in 19th century recognised antibodies in the blood stream of an immunized animal against tetanus and diphtheria. They observed clumping or agglutination of microorganisms and precipitation of soluble antigens by the serum of the immunized animal.
They also practiced the serotherapy method. Serotherapy is the injection of antiserum, for the therapeutic use, from an immunized animal to non-immune patient. They also carried out the fragmentation of immunoglobulin’s by pepsin digestion.
Antibodies are synthesised under the influence of antigens which penetrate into the body and disturb the normal blood composition. Antibodies are specific substances in the bodies of the vertebrates secreted in the tissue fluids from the lymphoid cells that have been stimulated by foreign substances (antigens with which they react specifically).
Antibodies appear in the blood due to the infection or immunization by live (attenuated) or dead bacteria, rickettsiae, viruses, toxins or toxoids etc. Antibodies which occur under the influence of active immunization are named Immune antibodies in contrast to normal antibodies which are found in the sera of men and animals which had not been infected or exposed to immunization.
Normal and immune antibodies can render harmless the causal agents of infectious diseases.
The Nature of Antibodies:
1. Antibodies are globulins which have been altered under the influence of antigens;
2. Molecules of antibodies, like normal serum globulins, are probably asymmetrical;
3. Antibodies are found in globulin fractions;
4. The transformation of normal globulin into immune globulins is due to alteration in the spatial configuration of active atomic groups of the protein molecules. A change in protein metabolism is responsible for this process. The mechanism of antibody production is not yet clear, though many theories are put forth;
5. Antibodies do not differ greatly from normal globulins;
6. They have almost similar isoelectric points, viscosity, molecular weight (from 100,000 to 1,000,000) and are sensitive to the temperature and to other denaturating agents.
7. Antibodies are thermolabile and denatured on heating at 70°C for one hour;
8. The pH of the medium and other factors affecting proteins can also affect the activity of antibodies;
9. Antibodies are not denatured by precipitation with ethyl alcohol at low temperature (from 0 to + 4°C) but are denatured by alcohol at high temperature. Natural salts (magnesium sulphate, ammonium sulphate and sodium sulphate) can cause the precipitation of proteins, but do not denature antibodies.
Ethyl alcohol at low temperature and natural salts can be used for fractionation of immune sera to obtain them in a pure state.
There are five types of human immunoglobulin’s : IgG; Ig M; Ig A; Ig D, and Ig E—on the basis of polypeptide chain structure consisting of heavy chains and light chains joined by disulphide bond.
Immunoglobulin G or IgG:
Seventy-five per cent of total serum constitutes IgG. It is the major immunoglobulin component of serum and has a molecular weight of 150,000 in man. The molecule has two antibody combining sites (Fig. 10.1) which are called Fab (antibody binding) portions of the molecule.
Fab fragment has both light and heavy chains. Light chains are of two types. They are known as K or L (k or 1) chains.
The heavy chain or 
chain exists in four forms:
Ig G1 Ig G2; Ig G3 and Ig G4. The molecule has also Fc fragment for complement fixation.
Ig M, Ig A, Ig D, and Ig E
Like Ig G globulin, each of these classes of immunoglobulin’s in man contain k and 1 light chains. The heavy chains are unique for each of the types of immunoglobulin’s. Ig M contains H chain, Ig A alpha chain, Ig D 
-chain and Ig E 
-chain.
Structure and Function of Immunoglobulin’s:
In the fab portion, the heavy chain component (Fd portion) contains as much as 85 per cent of the antigen-binding ability. The light chain seems to act together with the heavy chain to form a stable antibody-combining site (Fig. 10.1). The Fc portion of the IgG molecule is responsible for activation of complement system.
On combination with antigen, the IgG molecule springs open at the hinge region exposing the Fc portion which can activate the complement. If Ig G molecule has not combined with antigen, it will be unfolded. A single IgM molecule can get attached to a red cell by multiple combining sites and can bring about lysis; but for the same effect, 1000 IgG molecules are required.
Agglutination is due to linkage of a particulate antigen (red blood cell or bacterial) by two Fab fragments of IgM. The IgM is of large size, so it is confined in the blood stream and plays an important role in the protection against blood invasion by microorganisms.
IgM deficiency may result into septicaemia. Diphtheria toxin, lysozyme or virus (polio virus) can be neutralized by IgG antibodies and IgG antibodies can neutralize more effectively than IgM antibodies.
Fc fragment of IgG molecule may assist the Fab fragments in neutralisation of virus infectivity and also helps in the selective transport of IgG. The characteristic feature of IgE class immunoglobulin is responsible for the hypersensitivity reaction in man which is due to the activity of Fc portion of IgG molecule.
IgM antibody specific for particulate antigen (bacteria) makes the antigen more susceptible to the phagocytosis by coating the surface of particulate antigen, so it is called as opsonizing antibody (Opsonin).
Selective Transport of IgG:
In pregnant women, IgG globulins can pass through the placenta and reach the foetal circulation; this process is not due to the mere filtration but due to the selective transfer of IgG molecules which is brought about by a part of Fc fragment of IgG heavy chains.
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chain of IgG and is not found in the µ chain of IgM and a chain of IgA. This mechanism of selective transport is mainly observed in primates. Similarly, immunoglobulin are absorbed from the colostrum through the intestinal epithelium of the ruminants.
IgA globulins are selectively secreted into saliva, respiratory, intestinal mucous secretions and into the colostrum. This is another mechanism of selective transport. There are two types of IgA: Serum IgA and dimer secretory IgA.
Dimer form of IgA is manufactured locally and has an attached secretory transport piece which is not found in serum IgA. This transport piece is added to the IgA molecule during its passage into the mucous secretions from the lamina propria of the intestinal and respiratory tract.
This secretory IgA would protect from the infection. The dimer form of IgA acquires the ability to fix the complement after attaching to Gram-negative organisms.
The monomeric form of IgA can also pass from the lamina propria to the blood stream via lymphatic’s so that the infection of the intestine may lead to increased serum IgA levels. Local application of vaccine may stimulate dimer IgA.
Kinds of Antibodies:
1. Opsonins:
These antibodies render the microorganisms more susceptible to phagocytosis by altering the surfaces of the antigens.
2. Cytolysins:
They help to dissolve the cells that stimulated their production.
3. Antitoxins:
Are formed due to the stimulation of exotoxins which are protein in nature and soluble in water. These antibodies are inactivated or neutralized by the exotoxins.
4. Haemolysins:
If a rabbit is injected several times with minute quantity of washed erythrocytes of another species of animal, e.g. sheep, the serum of the recipient rabbit acquires the property of destroying the erythrocytes of the sheep.
This is due to the formation of cytolysins that sensitize the sheep red blood cells to complement and cause the lysis of the sheep erythrocytes. The haemoglobin escapes into the surrounding fluid. This process is called haemolysis. The haemolytic cytolysins are called haemolysins.
5. Precipitins:
These antibodies combine with the molecules of soluble proteins and bring about a cloudiness in the fluid or visible flakes. This flocculent turbidity is known as precipitate as seen in Venereal Disease Research Laboratory (VDRL) test; they may form line of precipitation when they react with soluble antigens.
6. Agglutinins:
They bring about the clumping of particulate antigens (bacteria, red blood cells)—hence agglutination (Fig. 8%). Many diagnostic tests are based on bacterial agglutinins. For example, Widal test is used to diagnose typhoid fever.
Haemagglutinins are responsible for the agglutination of red blood cells or haemagglutination; cold haemagglutins; viral haemagglutinins; and isohaemagglutinins:
(a) Cold haemagglutinins:
Are substances that appear in the blood of person with certain respiratory diseases e.g. atypical pneumonia of unknown or viral origin and trypanosomiasis. The sera of the patient in dilution of 1:10,000 agglutinate their own erythrocytes (auto-agglutination) when cooled to 2°C, but not at 37°C. This agglutination at low temperature is known as cold haemagglutination.
(b) Viral haemagglutinins:
Agglutination of red cells is brought about by several respiratory virus (influenza virus), not by antibody. The virus itself attaches to the red cells of the chick at 5°C and agglutinate them. It can be separated (eluted) from them by mixing with saline solution followed by an incubation at 37°C.
The viral haemagglutination tests are useful in the study of viruses and the diagnosis of viral diseases. In some infections, antibodies may prevent the viral agglutination of erythrocytes. They are called as haemagglutination-inhibition (HI) antibodies.
i. Isohaemagglutinins:
These antibodies are formed in response to one or more antigens (A, B, Rh, M) found in human erythrocytes. They do not agglutinate one’s own red blood cells, but cause agglutination, at body temperature (37°C), of red blood cells of another person.
They play an important role in blood transfusion. If the serum of the recipient agglutinates the red blood cells of a donor, or vice versa, then there is possibility of reaction or death of the recipient. It is not necessarily that the sera of all donors should agglutinate the erythrocytes of all the recipients, and vice versa, because there are numerous classes or groups of blood with respect to isohaemagglutinins, called blood groups.
Blood Groups:
One major system of blood groupings consist of A, B, AB and O groups. It detects the presence or absence of the agglutinogens (antigens) in the erythrocytes of the persons to be identified and grouped. For example, any one person may have antigen A (Group A) or B (Group B) or both (Group AB) or neither (Group O).
A person’s own serum never contains agglutinins against his own erythrocytes, but only against the absent agglutinogens (Fig. 8.2).
A person’s blood group is a physiological constant determined by the basic Mendelian laws of inheritance. It is possible to avoid the haemagglutination, by properly selecting donors and recipients of compatible group. Blood banks collect citrated blood donations already grouped and store in the refrigerator until needed.
Blood Grouping:
A nurse should have the knowledge of grouping the blood. A drop of the patient’s (recipient) blood should be added to a small amount of physiological saline (0.9 per cent sodium chloride) on a microscopic slide and then mixed with rabbit sera containing pure anti-A and anti-B antibodies. A readily visible agglutination will appear (Fig. 8.2) of the patient (recipient).
If the cells of the patient (recipient) are agglutinated by both anti-A and anti-B rabbit sera, then the cells of the patients are of group-AB.
If the cells are agglutinated by neither sera, they are of Group O. If anti-A serum agglutinates them and anti-B serum does not, then the blood cells are of Group A; if anti-B serum agglutinates them and anti-A serum does not, the cells are of Group B.
Certain irregularities in reaction can be eliminated and the blood groupings can be ascertained by testing the serum of the recipient against the red cells of the donor.
ii. Haemagglutination in Blood Grouping:
Anti-B serum and anti-A serum have been mixed on a glass slide with erythrocytes of group O, A, B and AB. Anti-B serum agglutinates erythrocytes of persons of Groups B and AB; anti-A serum agglutinates erythrocytes of persons of Groups A and AB.
Universal Donor:
A person of Group O is sometimes called universal donor because his red blood cells are not usually agglutinated by serum of recipients of any other group; but he must receive only Group O blood because his serum agglutinates cells of all other groups in the A-B-O system.
Direct cross-matching between recipient and donor is necessary to have a “safe universal donor.” There are several other antigen antibody systems in human bloods—e.g. Rh factor, M- N-s, p types which may cause serious transfusion reactions irrespective of A, B, O or AB groups.
Rh Factor:
When erythrocytes of Rhesus (Rh) monkeys are injected into rabbits, the serum of the rabbits contained anti-Rh agglutinins which agglutinate erythrocytes of 85 per cent or more human beings. The antigen in the human red cells which reacted with anti-Rh rabbit serum was designated as the Rh agglutinogen.
The person having the Rh agglutinogen is called Rh positive (+). If the Rh agglutination is absent in red blood cells of the remaining 15 per cent whose red blood cells do not react with the anti-Rh rabbit serum, they are called Rh- negative. The Rh agglutinogen is complex and corresponds to several antibodies or blood factors—Hr, hr’, rh’, hr’, rh’. The most important is Rh factor.
iii. Rh Factor and Transmission:
If an Rh negative person receives a blood transfusion from a person who is Rh positive, the recipient develops in about two weeks antibodies that agglutinate the red cells of all Rh positive persons.
Now, if a subsequent transfusion is given from an Rh positive donor and the Rh antibody titre (concentration) will rise to maximum, thereby all the red cells of the donor are agglutinated and, ultimately, haemolysis leading to severe transfusion reaction and death. Therefore, it is essential, before each blood transfusion, to evaluate the Rh factor and blood group.
iv. Erythroblastosis Foetalis (Haemolytic Disease of the Newborn):
If Rh negative woman becomes pregnant by an Rh positive man, the child will most probably, inherit Rh antigen in its blood cells. During pregnancy, if the child is Rh-positive (i.e. Rh factor), foetal red blood cells with Rh antigen often cross the foetal barrier, pass into the mother’s blood circulation and the foetal Rh factor stimulates the maternal tissue to produce Rh antibodies.
These Rh antibodies diffuse back into the foetal blood circulation. In any initial pregnancy, since the mother’s Rh antibody titre is too low, there will be no detrimental effects in the foetus; when Rh antibody concentration increases with subsequent pregnancies, the red cells of the foetus are destroyed by Rh antibodies.
Hence, the child with Rh factor (i.e. Rh positive) and Rh antibodies may be born dead (uncommon) or with haemolytic disease of the newborn (erythroblastosis foetalis) which is common. This type of condition may prevail if Rh negative mother has received a blood transfusion from Rh positive donor. The periodic checkup of the maternal blood for rising titre of Rh agglutinins is of great importance as a precautionary measure.