Monoclonal Antibody Monoclonal Antibody ProductionProduction

Cathy Langford, Judy Knadler, Cathy Langford, Judy Knadler, Jamie WolfJamie Wolf

Monoclonal Antibody ProductionMonoclonal Antibody ProductionMethodMethod

Monoclonal Antibody Production technology was developed in 1975. Since Monoclonal Antibody Production technology was developed in 1975. Since its development it has been very important in the modern medical science its development it has been very important in the modern medical science with the diagnosis, therapy, research and even basic science today. It is with the diagnosis, therapy, research and even basic science today. It is still largely dependent upon animal testing however. Because it requires still largely dependent upon animal testing however. Because it requires immunization of mice in order for them to create the antibodies to be grown. immunization of mice in order for them to create the antibodies to be grown.

Monoclonal Antibody Production or mAb is produced by cell lines or clones Monoclonal Antibody Production or mAb is produced by cell lines or clones obtained from the immunized animals with the substance to be studied. Cell obtained from the immunized animals with the substance to be studied. Cell lines are produced by fusing B cells from the immunized animal with lines are produced by fusing B cells from the immunized animal with myeloma cells. To produce the desired mAb, the cells must be grown in myeloma cells. To produce the desired mAb, the cells must be grown in either of two ways: by injection into the peritoneal cavity of a suitably either of two ways: by injection into the peritoneal cavity of a suitably prepared mouse (the in vivo, or mouse ascites, method) or by in vitro tissue prepared mouse (the in vivo, or mouse ascites, method) or by in vitro tissue culture. culture.

The vitro tissue culture is the method used when the cells are places in The vitro tissue culture is the method used when the cells are places in culture outside the mouse's body in a flask. culture outside the mouse's body in a flask.

Explanation Of PictureExplanation Of Picture The previous picture is showing a mouse being immunized against a target The previous picture is showing a mouse being immunized against a target

cell “X”. This will allow the mouse to produce antibodies for that will target cell “X”. This will allow the mouse to produce antibodies for that will target against the “X” antigen. against the “X” antigen.

Once the mouse has formed antibodies to the “X” antigen the cells are then Once the mouse has formed antibodies to the “X” antigen the cells are then isolated in the mouse’s spleen. Monoclonal antibodies are produced by isolated in the mouse’s spleen. Monoclonal antibodies are produced by fusing single antibody-forming cells to tumor cells grown in culture. The fusing single antibody-forming cells to tumor cells grown in culture. The resulting cell is called a hybridoma. Hybridoma cells are continuously resulting cell is called a hybridoma. Hybridoma cells are continuously growing cell line generated by the fusion of a myeloma cell and a normal growing cell line generated by the fusion of a myeloma cell and a normal cell that are capable of producing antibodies. cell that are capable of producing antibodies.

Each hybridoma will produce relatively large quantities of identical antibody Each hybridoma will produce relatively large quantities of identical antibody molecules. Because the hybridoma is multiplying in culture, it is possible to molecules. Because the hybridoma is multiplying in culture, it is possible to produce a population of cells, each is producing identical antibody produce a population of cells, each is producing identical antibody molecules. These antibodies are called "monoclonal antibodies" because molecules. These antibodies are called "monoclonal antibodies" because they are produced by the identical offspring of a single, cloned antibody they are produced by the identical offspring of a single, cloned antibody producing cell.producing cell.

Why this method is used!!Why this method is used!! This method is used because antibodies must be formed from the This method is used because antibodies must be formed from the

immunization of the substance being studied. So antibodies must be immunization of the substance being studied. So antibodies must be produced. Once the antibodies are produced the animal aspect of the study produced. Once the antibodies are produced the animal aspect of the study can be eliminated and tissue culture can then be used. can be eliminated and tissue culture can then be used.

When using live mice researchers have found that it is the better option When using live mice researchers have found that it is the better option because in vitro doesn’t always produce adequate cell lines that are because in vitro doesn’t always produce adequate cell lines that are adaptive to tissue culture. Protein denaturation can occur from purification adaptive to tissue culture. Protein denaturation can occur from purification techniques and antibody activity is decreased with normal activity not techniques and antibody activity is decreased with normal activity not represented. Also cell lines could possibly become contaminated when represented. Also cell lines could possibly become contaminated when

using in vitro technique.using in vitro technique.

Monoclonal ELISA test system run. Labels across thetop of the plate indicate the capture antibody used in the designatedcolumn: P, polyclonal rabbit anti-Lp 1 through 9, the mousemonoclonal antibody indicated by the same number; M, polyclonalnormal mouse immunoglobulin. Labels on the side of the plateindicate the specimen tested in that row: A, PBS-BSA; B through D,positive lung homogenate extracts from cases in the Philadelphiaoutbreak; E, Philadelphia 1 culture extract; F, OLDA cultureextract; G, Knoxville 1 culture extract; H, negative lung homogenateextract.

ReferencesReferences http://www.accessexcellence.org/RC/VL/GG/monoclonal.htmlhttp://www.accessexcellence.org/RC/VL/GG/monoclonal.html http://www.med.nyu.edu/dlar/iacuc/monoclonal.htmlhttp://www.med.nyu.edu/dlar/iacuc/monoclonal.html http://orsted.nap.edu/openbook.php?record_id=9450&page=5http://orsted.nap.edu/openbook.php?record_id=9450&page=5 SUSAN L. BROWN,* WILLIAM F. BIBB, AND ROGER M. McKINNEY Use of Monoclonal Antibodies in an Epidemiological MarkerSystem:

a Retrospective Study of Lung Specimens from the 1976Outbreak of Legionnaires Disease in Philadelphia by IndirectFluorescent-Antibody and Enzyme-Linked Immunosorbent; AssayMethodsJOURNAL OF CLINICAL MICRODIOLOGY, Jan. 1985, p. 15-19