[Skip to Content](https://www.sigmaaldrich.com#main-content) [](https://www.sigmaaldrich.com/US/en) Products Cart0 USEN Products ProductsApplicationsServicesResourcesSupport [Login / Register](https://www.sigmaaldrich.com/oidc-sign-in) [Order Lookup](https://www.sigmaaldrich.com/US/en/order-lookup) [Quick Order](https://www.sigmaaldrich.com/US/en/quick-order) Cart0 [Home](https://www.sigmaaldrich.com/US/en)[Western Blotting](https://www.sigmaaldrich.com/US/en/applications/protein-biology/western-blotting)Antibody Basics # Antibody Basics - [Basic Antibody Structure](https://www.sigmaaldrich.com#basic) - [Antibody Classes and Subclasses](https://www.sigmaaldrich.com#classes) - [Antibody Forms](https://www.sigmaaldrich.com#form) ## [](https://www.sigmaaldrich.com)Basic Antibody Structure Immunoglobulins (Igs) are produced by B lymphocytes and secreted into plasma. The Ig molecule in monomeric form is a glycoprotein with a molecular weight of approximately 150 kDa that is shaped more or less like a Y. Basic structure of the Ig monomer (__Figure 1__) consists of two identical halves connected by two disulfide bonds. Each half is made up of a heavy chain of approximately 50 kDa and a light chain of approximately 25 kDa, joined together by a disulfide bond near the carboxyl terminus of the light chain. The heavy chain is divided into an Fc portion, which is at the carboxyl terminal (the base of the Y), and a Fab portion, which is at the amino terminal (the arm of the Y). Carbohydrate chains are attached to the Fc portion of the molecule. The Fc portion of the Ig molecule is composed only of heavy chains. Fc regions of IgG and IgM can bind to receptors on the surface of immunomodulatory cells such as macrophages and stimulate the release of cytokines that regulate the immune response. The Fc region contains protein sequences common to all Igs as well as determinants unique to the individual classes. These regions are referred to as the constant regions because they do not vary significantly among different Ig molecules within the same class. The Fab portion of the Ig molecule contains both heavy and light chains joined together by a single disulfide bond. One heavy and one light chain pair combine to form the antigen binding site of the antibody. Each Ig monomer is capable of binding two antigen molecules.   __Figure 1.__Basic Immunoblobulin structure. ## [](https://www.sigmaaldrich.com)Antibody Classes and Subclasses The main immunoglobulin (Ig) classes (__Figure 2__) are IgA, IgD, IgE, IgG, and IgM. Birds also produce IgY in egg yolks. [Search Antibodies](https://www.sigmaaldrich.com/US/en/products/protein-biology/antibodies#explorer)  __Figure 2.__Main Immunoglobulin classes. Class identity is determined by class-specific sequences in the Fc region of the heavy chain which are designated by Greek letters corresponding to the Ig letter designation: alpha-IgA, delta-IgD, epsilon-IgE, gamma-IgG, mu-IgM. Light chains are universal among immunoglobulins and occur as two types — kappa or lambda. These are usually designated by the Greek letters kappa and lambda. Refer to __Table A__ and __Table B__ for Human and Mouse Immunoglobulin Properties, respectively. Human Immunoglobulin Properties __Property__I__gG__ I__gA__ __IgM__ __IgD__ __IgE__ H Chain class (heavy chain)γ α µδε H Chain Subclassesγ1γ2γ3γ4α1α2None NoneNone H Chain MW50 kDa 50 kDa60 kDa50 kDa55 kDa55 kDa70 kDa62 kDa70 kDa L Chain MW* (light chain k & λ )23 kDa 23 kDa23 kDa23 kDa23 kDa23 kDa23 kDa23 kDa23 kDa Total MW 150 kDa150 kDa170 kDa150 kDa160 kDa (serum) 600 kDa (secretory)160 kDa (serum) 600 kDa (secretory)970 kDa180 kDa190 kDa Ext. Coeff. 0.1% @280 nm1.41.41.41.41.321.321.181.71.53 Complement fixationweakweakstrongnononostrongnono Fc receptor bindingstrongweakstrongweakyesyesyesnoyes Mast cell/basophil degranulationnonononononononoyes Placental transferstrongweakstrongstrongnonononono Table A \* Light chains are present on all Immunoglobulin classes. In humans, k. chains are found 67% of the time, and λ chains are found 33% of the time. For ratios in other species see Immunoglobulin Light Chain Ratios table. Mouse Immunoglobulin Properties __Property__                         __IgG__ __IgA__ __IgM__ __IgD__ __IgE__ H Chain class γα µδε H Chain Subclassesγ1γ2aγ2bγ3NoneNoneNoneNone 0% of total LPG 4624272     Light chain Approximately 95% kappa (K LC) - 5% lambda (λ LC) Molecular weight (kDa)160 160 (monomer) 350-400900180190 Normal Serum Concentration (mg/mL)0.3-5.00.1-4.00.1-50.1-0.20.01-0.030.1-1.60.003-0.010.0001-0.001 Serum half-life (days)8-113-122.6-3.54-80.5-10.5-1<1<1 Electrophoretic Mobilityfastslowslowslow     Carbohydrate (%)2-37-119-1212-1512 Complement fixation \-+++++++/--+++n.kn.k Ext. Coeff. 0.1% @280 nm 1.41.351.18   Sedimentation Coeff.6.66.7196.88 Table B The ratio of kappa to lambda found in the Ig population varies by species. Refer to __Table C__. | | | |-----------------------------------|-------------| | Immunoglobulin Light Chain Ratios | | | __Species__ | __% k / λ__ | | Human | 67/33 | | Mouse | 99/1 | | Rat | 99/1 | | Rabbit | 90/10 | | Goat | 1/99 | | Sheep | 1/99 | | Pig | 50/50 | | Bovine | 1/99 | | Horse | 1/99 | | Chicken | N/A | Table C ## [](https://www.sigmaaldrich.com)Antibody Forms We offer antibodies as whole (intact) molecules and as F(ab)2 fragments of antibodies. Table D highlights some common forms of antibodies. | | | | |--------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------| | Antibody Form | Description | Uses | | Whole Antiserum | Whole antiserum contains specific antibody as well as all the other host serum proteins. | | | Fractionated Antiserum (Ig fraction) | Whole antiserum is fractionated to yield primarily the immunoglobulin fraction of antiserum. This fraction may contain small amounts of the other host serum proteins. | IgG fractions and fractionated antiserum may be considered useful in situations where very high affinity is required, most commonly when the antigen of interest is rare or present in low abundance. IgG fractions may have the benefit of containing very high affinity antibodies. Affinity isolation may remove some very high affinity antibodies because they bind so tightly to the affinity matrix that they are not eluted. | | IgG Fraction of Antiserum | Whole antiserum is fractionated and then further purified by ion exchange chromatography to provide the IgG fraction of antiserum. This fraction is essentially free of other host serum proteins. | IgG fractions and fractionated antiserum may be considered useful in situations where very high affinity is required, most commonly when the antigen of interest is rare or present in low abundance. IgG fractions may have the benefit of containing very high affinity antibodies. Affinity isolation may remove some very high affinity antibodies because they bind so tightly to the affinity matrix that they are not eluted. | | Affinity Isolated Antibody (AIA) | These products are the most purified, therefore, give the lowest amount of non-specific binding. Affinity isolated antigen specific antibody is obtained from antiserum by immunospecific purification using antigen-bound agarose, which removes essentially all the host serum proteins, including immunoglobulins, which do not specifically bind to the antigen. | Universal | | Ascites Fluid | Ascites fluid is intraperitoneal fluid extracted from mice that have had hybridoma cells expressing specific monoclonal antibodies injected into their peritoneal cavity, which serves as a growth chamber for the cells. The hybridoma cells grow to high densities secrete high-titered antibodies. Ascites fluid is clarified by centrifugation to remove the lipid layer and the cell pellet. It contains specific antibody as well as other host serum proteins including immunoglobulins. Specific antibody concentration range is 1-10 mg/mL. Total protein concentration is approximately 20 mg/mL | Universal | | Tissue Culture Supernatant | Supernatant is the fluid resulting from centrifugation of hybridomas in tissue culture that are secreting specific monoclonal antibodies. Supernatants therefore typically contain culture medium and 5-10% fetal calf serum. Specific antibody concentration is in the range of 50 5g/mL. | Universal | | Purified Immunoglobulin | Purified immunoglobulin typically refers to monoclonal antibodies that have been purified using protein A or protein G affinity chromatography. | Universal | | F(ab)2 Fragment | F(ab)2 fragments are produced by digestion of IgG with pepsin yielding a divalent molecule (containing two antibody binding sites) but lacking the Fc portion. | F(ab) 2 antibody fragments are used in assay systems where the presence of the Fc region may cause problems. Samples such as lymph nodes, spleen, and peripheral blood preparations contain cells with Fc receptors (macrophages, B lymphocytes, and natural killer cells) which could bind the Fc region of intact antibodies, causing high background staining. Use of F(ab)2 fragments ensures that any antibody binding observed is not due to Fc receptors.1 These fragments may also be desirable for staining cell preparations in the presence of plasma, because they are not able to bind complement, which could lyse the cells. F(ab)2 fragments, because of their smaller size, are able to localize antigen more precisely than intact IgG, particularly in staining tissue for electron microscopy. The divalency of F(ab)2 fragments enables cross-linking of antigens, allowing use of these reagents in precipitation assays, rosetting assays2, or for cellular aggregation via surface antigens.3 | Table D ## Products Sorry, an unexpected error has occurred Response not successful: Received status code 500 ### References 1\. K S, T A, T T. 1998. Decreased Fcgamma receptor III (CD16) expression on peripheral blood mononuclear cells in patients with Sjögren's syndrome. 25 4 689-96. J Rheumatol. 2\. DESAINTMARTIN J, SCHWARTZ J, MANNESSIER L, EYQUEM A. 1983. Idiotypic and anti-idiotypic determinants on lymphocytes during anti-Rh immunization. Revue Francaise de Transfusion et Immuno-hématologie. 26(6):573-583. [https://doi.org/10.1016/s0338-4535(83)80072-5](https://doi.org/10.1016/s0338-4535%2883%2980072-5) 3\. De Reys S, Hoylaerts M, De Ley M, Vermylen J, Deckmyn H. 1994. Fc-independent cross-linking of a novel platelet membrane protein by a monoclonal antibody causes platelet activation. 84(2):547-555. 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