Indian Journal of Ophthalmology

ARTICLES
Year
: 1972  |  Volume : 20  |  Issue : 2  |  Page : 45--48

Properties and uses of antibodies


SS Rao 
 Haffkine Institute, Bombay-12, India

Correspondence Address:
S S Rao
Haffkine Institute, Bombay-12
India




How to cite this article:
Rao S S. Properties and uses of antibodies.Indian J Ophthalmol 1972;20:45-48


How to cite this URL:
Rao S S. Properties and uses of antibodies. Indian J Ophthalmol [serial online] 1972 [cited 2020 Jun 6 ];20:45-48
Available from: http://www.ijo.in/text.asp?1972/20/2/45/34667


Full Text

When a pure protein say cry­stalline egg albumin is injected into an animal, a family of antibo­dies are produced which differ in their physical properties (e.g. molecular weight, electrophoretic biochemical properties (carbo hydrate content, amino acid com­position etc.), immunological pro­perties and specificity of reaction with the different determinant groups (i.e. part of the antigen molecule that reacts with antibody) in the egg albumin. Collectively they are called Immunoglobulins (Ig.) Immunoglobulins are defined as proteins of animal origin en­dowed with known aitibody activity and certain proteins related to them in chemical structure and hence of "antigenic specificity". The related proteins which show similar chemical structure are the proteins found in large amounts in multiple myeloma patients and also Bence­Jones Proteins found in the urine of certain myeloma patients.

In man five classes of immuno­globulins have been found and some of them have subclasses also These, in order of decreasing con­centration are : IgG (with sub­ classes IgG 1 . IgG 2 , IgG 3 , and IgG 4 ). IgA (subclasses, IgA 1 , and IgA 2 ,), IgM, IgD and IgE. When an antigen is injected all these im­munoglobulins are formed to each of the determinant groups of the antigen. Their physical proper­ties biochemical properties and immunological properties are given in the table opposite. They differ in their immunologi­cal properties also. For example, IgG can pass through placenta and the immunity of the new born child is due to this immunoglobu­lin. IgA is found in secretions such as milk, saliva, gastrointestinal fluid, lung secretion etc. IgM is the immunoglobulin that is formed ear­liest after injection of the antigen and is most powerful in opsoniza­tion (i.e. increasing phagocytosis) and in fixing complement. IgE is most active in causing allergic and anaphylactic reactions. The func­tion of IgD is not known as yet. The most interesting property of a sub­class of IgD namely IgG, is tumour enhancement. Contrary to the nor­inal cytotoxic activity of the anti­bodies to transplanted tumours. IgG., actually helps the growth of the tumour. It it likely that this immunoglobulin is responsible for the prolonged survival of organ homografts.

Structure of Immunoglobulins

The structure of tre human Im­munoglobulins has been worked out thanks to the disease multi­ple myeloma which is malignancy of the plasma cells. Normal plasma cells have a short half life of 3 days and they like myeloma cells do not have the ability to multip­ly. Since plasma cells secrete im­munoglobulin, the myeloma cells secrete into blood circulation large amounts of immunoglobulin of one class or subclass. These im­munoglobulin are very homogene­ous in their physical and bio­chemical properties. They are formed without stimulation with an antigen. However, some of these myeloma proteins did show antibody activity when tested against a large panel of antigens.

Electron microscopic pictures have shown that these immunoglo­bulins have the shape of the letter Y with a hinge in the middle and two movable arms which can come together or stretch upto an angle of 180°. In the case of IgA, they occur as dimers attached to each other at ends of their nonflexible arm. In secretions, IgA dimer is attached to a protein having a molecular weight of about 50,000. IgM occurs as pentamer with five Y shaped units attached at their nonflexible ends in a circle. In elec­tron microscopic pictures IgM molecule looks like a ring with a hollow centre and with arms stretching out from the ring.

Analysis of myeloma proteins have revealed that the immunoglo­bulin monomer molecule has two pairs of polypeptide chains. One pair of identical polypeptide chain called L chain is made up of about 212 amino acids and stretch from the ends of the flexible arms down upto the hinge region. The second pair of identical heavy chains (H) made up of about 450 amino acids all the way from the top of the flexible arms down to the end of the nonflexible arm. The light and heavy chains are bound to each other and to themselves with dis­ulphide bonds through the sulphur containing amino acid cystine. Further, it was found that the monomer antibody molecule has two identical antigen binding sites at the end of the two flexible arms. Thus the monomer antibody mole­cule has two binding sites with an­tigen. The IgG molecule could also be treated under mild conditions with the proteolytic enzyme papain which splits the molecule at the hinge region giving three pieces; two identical flexible arms called Fab still retain the property of combining with antigen, and the third nonflexible part from the hinge down called Fd which has no antigen binding pro­perty. The Bence-Jones proteins occuring in the urine of some my­eloma patients were found to be dimers of the two identical light chains. The light chains were found to be of two types called Kappa and Lambda, the former is about two thirds and the latter about one third. The heavy chains vary from one class of antibody to another, two thirds of each class of the immunoglobulins have Kappa light chains and one third the lamda light chains. When the Fab which contains the complete light chain and about half of the heavy chain (Fd) is treated to separate the chains, the individual chains lose the antigen binding property. The antigen binding site of antibody is therefore shared by both chains at their ends and the two chains between them fit over the determinant group like the glove fitting over the hand. A very interesting finding from the ana­lysis of different myeloma and Bence-Jones proteins is that the specificity of the binding between antibody and that of antigen is due to differences in the sequence of amino acids from the end of the flexible arm to about 110 amino acids, roughly half the length of light chains. The same is true of the Fd part of the heavy chain also. The remaining part of the light and heavy chains have in­variable or constant amino acid sequence. The ability of the anti­body molecule to distinguish the difierent determinant groups of antigens is due to this variation in the amino acid sequence of part of light and heavy chains. How the antibody producing mechanism can vary the sequence of the amino acids in the variable part of the molecule only is still not known. How this most sophisti­cated mechanism has evolved and how it gets activated when an antigen is introduced are very challenging problems being tackl­ed vigorously by scientists.

Use of Immunoglobulins in therapy

Antisera have been used in the treatment of diseases such as tetanus and diphtheria since 1890 when Behring and Kitasato dis­covered their therapeutic use. Later on, antisera prepared in hor­ses came to be used in other toxic conditions such as bites of snakes etc. Horse antihuman lympocyte sera has recently been used as a powerful immunosuppressor after transplantation when the chemical immunosuppressors like hydro­cortisone are not effective.

Human anti Rh sera has now been found to be the only effective remedy to prevent Rh isoimmuni­zation. Injection of such anti Rh sera soon after the delivery of an Rh + infant by Rh mother can suppress and prevent formation of Rh antibodies by the mother. The next child will not suffer from erythroblastosis foetalis.

Use as analytical reagent

The most important use of anti­bodies is as biological reagents. Two properties of antibodies make them eminently suitable for this purpose. Their specificity and sensitivity. Immuno assay techni­ques using antisera is therefore finding application in every branch of biomedical research. The assay of some hormones secre­ted by pituitary which occur in nano (10 -9 ) or pico (10 -12 ) gram quantities in serum would not have been possible without the use of sensitive immunoassay techniques.

The specific reaction between antibody and its antigen is follow­ed by a variety of secondary pheno­menon which can be detected by a number of techniques : I) If the antigen has biological activity such as toxicity (e.g. tetanus toxin, venom etc.), enzymic activity or hormone activity, the antibody neutralizes such activity. 2) If the antigen is particulate in nature (e. g. RBC, bacteria or when a soluble antigen is firmly bound to particulate matter) the antisera will cause these particles to agglu­tinate. 3) In sufficient concentration the antibody will show precipita­tion with its particular antigen (precipitation technique). 4)Anti­gen-antibody complexes can fix certain heat labile factors which occur in blood serum and called collectively complement (comple­ment fixation test). 5) Antigen-antibody reaction can also be de­tected by their ability to release pnarmacologically active sub­stances like histamine which give rise to cutaneous or systemic ana­phylactic reaction. 6) Antigen­antibody reaction can also increase clearence of antigen from circula­tion, increase phagocytosis or in­hibit migration of sensitized micro­phages. Immunoassay techniques have therefore a wide choice of methods. Some of the techniques can be made more sensitive of amplified by tagging radio active isotopes (Radioimmuno assay), fluorescent dyes (immunofluores­cence technique) or by the use of double antibody (i.e. antisera pre­pared in another species of animal to the specific antibody).

Antisera have been in use for identification of different strains of bacteria for over seventy years However, their use in other fields of medical research, especially endocrenology is more recent. It is now possible to study insulin secretion or the output of hormo­nes by pituitary and other endo­crenes accurately by the use of radioimmuno assay techniques. For example, it has been possible by analysis of sera to accurately det­ermine the ovulation time of women and detect the pregnancy four days before the expected menstrual period. Pregnancy can also be detected within a few days of missed period by analysis of urine by simple agglutination test. Anti­sera can also be produced against small steroid hormones after at­taching them to carrier antigens and such sera are being used to study hormonal balance in abnor­mal cases. Sensitivity to penicillin or other drugs can also be detected by immuno assay techniques. The immuno assay techniques are therefore very versatile, specific and sensitive.