Molecular Biology of the Cell
Bruce AlbertsMolecular Biology of the Cell Posts
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In principle, therefore, a monoclonal antibody can be made against any protein in a biological sample. Once an antibody has been made, it can be used to localize the protein in cells and tissues, to follow its movement, and to purify the protein to study its structure and function.
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Hybridomas
B lymphocytes normally have a limited life-span in culture, but individual antibody-producing B lymphocytes from an immunized mouse, when fused with cells derived from a transformed B lymphocyte cell line, can give rise to hybrids that have both the ability to make a particular antibody and the ability to multiply indefinitely in culture.
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Cell culture
Tissue culture began in 1907 with an experiment designed to settle a controversy in neurobiology. The hypothesis under examination was known as the neuronal doctrine, which states that each nerve fiber is the outgrowth of a single nerve cell and not the product of the fusion of many cells. To test this contention, small pieces of spinal cord were placed on clotted tissue fluid in a warm, moist chamber and observed at regular intervals under the microscope. After a day or so, individual nerve cells could be seen extending long, thin filaments (axons) into the clot. Thus, the neuronal doctrine received strong support, and the foundation was laid for the cell-culture revolution.
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Cell can be isolated from tissues
The first step in isolating individual cells is to disrupt the extracellular matrix and cell-cell junctions that hold the cells together. For this purpose, a tissue sample is typically treated with proteolytic enzymes (such as trypsin and collagenase) to digest proteins in the extracellular matrix and with agents (such as ethylenediaminetetraacetic acid, or EDTA) that bind, or chelate, the Ca+2 on which cell-cell adhesion depends. The tissue can then be teased apart into single cells by gentle agitation.
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Two such proteins expressed by T cells have attracted great attention because of their roles in suppressing the ability of T cells to inhibit cancer progression: CTLA4 and PD1 proteins inhibit T cell activity in different ways, and monoclonal antibodies against either or especially both can relieve the inhibition and allow T cells to dramatically destroy the tumors in some patients with metastatic cancer.
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When an accessory receptor has a direct role in activating the T cell by generating its own intracellular signals, it is called a "co-receptor". The most important and best understood of the co-receptors on T cells are the CD4 and CD8 proteins, both of which are Single-Pass transmembrane proteins with extracellular Ig-like domains. Like TCRs, they recognize MHC proteins, but, unlike TCRs, they bind to invariant parts of the MHC protein, far away from the peptide-binding groove.
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