10-08-2012, 11:33 PM
(This post was last modified: 09-23-2014, 05:48 PM by Administrator.)
Our immune system acts as the protective shield of our body against various infections by bacteria and virus causing various diseases. White blood cells, the army of immune system is composed of neutrophils, eosinophil, basophil, lymphocyte and monocyte each carrying out its unique function in fighting against the foreigner entering the body. The B cells of the lymphocytes are the intelligent soldiers which recognize the type of foreign object (antigen) entering the body and releases a weapon called antibody to track the foreigner and destroy them. The two novel traits of an antibody are its specificity to the antigen and once induced its assurance to the body to provide continual resistance to the particular type of disease acquired. Baffled by these two unique features of an antibody, scientists decided to use them for the welfare of the human kind and developed techniques to produce antibodies in vitro. The result is the production of ‘Monoclonal Antibody’.
Monoclonal antibody is the term used for the antibody produced in vitro by multiplying a single hybrid cell, obtained by cloning selected cells from a single source. Monoclonal antibodies are known for its purity and specificity. The conventional method of monoclonal antibody production was done by injecting the test animal with a particular type of antigen. After few days of the dose of the antigen, blood is drawn from the test animal and the antibodies were extracted from the serum of the blood. This method was failure both qualitatively and quantitatively. The antibodies obtained were found to be impure (mixed variants) and the amount obtained was also significantly less. Hence adoption of cloning technique was identified as the optional method to produce antibodies in vitro.
In this method, scientists selected tumor cells for its ability to multiply intensively and the antibody producing mammalian cells and fused these two under in vitro conditions. On the onset of the production, the test animal usually a mice is injected with an antigen to stimulate the antibody production. The antibody producing cells are identified and extracted from the spleen of the mice and it is fused with the myeloma cells which were extracted from the mice earlier and cultured in vitro. The resulting hybrid cell or hybridoma is observed for the presence of the desired antibody and once satisfied, the hybrids are subjected to grow in culture to produce splendid quantity of monoclonal antibodies. Again, the extraction and purification of the monoclonal antibody from the hybridoma is done by sequence of processes like centrifugation, filtration, ultra filtration or dialysis and ion exchange chromatography. Later the ion exchange chromatography was replaced by size exclusion chromatography which was found to be more effective in purifying. Also a procedure called affinity purification was employed to obtain the maximum purity. After undergoing the steps of purification, the final product, the monoclonal antibody is checked for the level of purity by using either chromatogram or gel electrophoresis or capillary electrophoresis.
The first test animal used for the production of monoclonal antibody is mice and a consequence reaction like allergy was observed in humans when supplemented with the monoclonal antibodies produced from mouse cell. Also, humans responded only to the initial dose and developed resistance to further doses. This posed as a bigger problem in obtaining the benefits of the monoclonal antibody and as a result evolved the chimeric antibody. The chimeric antibody is developed by inserting some human amino acid sequence into the animal developed monoclonal antibody.
The novel idea of developing fully human monoclonal antibody is a major breakthrough in the production of monoclonal antibody. In this method, blood sample is collected from an individual (donor) recovered from a particular type of infection and the antibody specific cells are extracted and immortalized. These cells are then subjected to micro well assay technique and the antigen specific antibodies are identified by fluorescence method and isolated. These cells are expanded and characterized before passing to other cell types for large scale production. The difficulty in identifying a donor is eluded by extracting cell from a healthy person and activating the cell for specific antibody production in vitro. The advancement in genetic engineering technology serves the human monoclonal antibody production by using transgenic mice.
The wide therapeutic application of monoclonal antibodies states the significance of the production of the monoclonal antibody.
Monoclonal antibody is the term used for the antibody produced in vitro by multiplying a single hybrid cell, obtained by cloning selected cells from a single source. Monoclonal antibodies are known for its purity and specificity. The conventional method of monoclonal antibody production was done by injecting the test animal with a particular type of antigen. After few days of the dose of the antigen, blood is drawn from the test animal and the antibodies were extracted from the serum of the blood. This method was failure both qualitatively and quantitatively. The antibodies obtained were found to be impure (mixed variants) and the amount obtained was also significantly less. Hence adoption of cloning technique was identified as the optional method to produce antibodies in vitro.
In this method, scientists selected tumor cells for its ability to multiply intensively and the antibody producing mammalian cells and fused these two under in vitro conditions. On the onset of the production, the test animal usually a mice is injected with an antigen to stimulate the antibody production. The antibody producing cells are identified and extracted from the spleen of the mice and it is fused with the myeloma cells which were extracted from the mice earlier and cultured in vitro. The resulting hybrid cell or hybridoma is observed for the presence of the desired antibody and once satisfied, the hybrids are subjected to grow in culture to produce splendid quantity of monoclonal antibodies. Again, the extraction and purification of the monoclonal antibody from the hybridoma is done by sequence of processes like centrifugation, filtration, ultra filtration or dialysis and ion exchange chromatography. Later the ion exchange chromatography was replaced by size exclusion chromatography which was found to be more effective in purifying. Also a procedure called affinity purification was employed to obtain the maximum purity. After undergoing the steps of purification, the final product, the monoclonal antibody is checked for the level of purity by using either chromatogram or gel electrophoresis or capillary electrophoresis.
The first test animal used for the production of monoclonal antibody is mice and a consequence reaction like allergy was observed in humans when supplemented with the monoclonal antibodies produced from mouse cell. Also, humans responded only to the initial dose and developed resistance to further doses. This posed as a bigger problem in obtaining the benefits of the monoclonal antibody and as a result evolved the chimeric antibody. The chimeric antibody is developed by inserting some human amino acid sequence into the animal developed monoclonal antibody.
The novel idea of developing fully human monoclonal antibody is a major breakthrough in the production of monoclonal antibody. In this method, blood sample is collected from an individual (donor) recovered from a particular type of infection and the antibody specific cells are extracted and immortalized. These cells are then subjected to micro well assay technique and the antigen specific antibodies are identified by fluorescence method and isolated. These cells are expanded and characterized before passing to other cell types for large scale production. The difficulty in identifying a donor is eluded by extracting cell from a healthy person and activating the cell for specific antibody production in vitro. The advancement in genetic engineering technology serves the human monoclonal antibody production by using transgenic mice.
The wide therapeutic application of monoclonal antibodies states the significance of the production of the monoclonal antibody.
![[+]](https://www.biotechnologyforums.com/images/netpen-pro/collapse_collapsed.png)