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The concept of clonality - introduction to the concepts behind Southern Blots and PCR

Leukemia is a monoclonal disease, or simply "clonal". This means that all of the renegade white cells derive from a single errant cell. Scientists make use of this fact to diagnose leukemia: if they can show that all of the blast population is alike, then they can prove that the patient does indeed have leukemia. Southern blots and PCR assays were developed to show that a population of cells has a clonal faction. With refinements, these methods are now being utilized in the detection of MRD.

To understand how these methods work, you need to understand some basics of DNA, genes, and B-cell development.

B-cell immunoglobulin and T-cell receptor genes

Immunoglobulins (Ig) and T-cell receptors (TCR) are two types of proteins produced by B-cells. Both of these proteins, Ig and TCR, are involved in antigen recognition. Antigens can be just about anything that comes into the body, and if the body wants to get rid of them (or to what the antigen is attached to), the body first has to recognize them. Our immune system must therefore be able to recognize essentially an infinite number of antigens that might invade humans over the centuries.

B-cells (and T-cells) have a system for producing an almost infinite number of different proteins to recognize antigens. This system is composed of proteins such as Ig and TCR which are similar but not exact in structure in each B-cell. Each protein can be built by the cell in millions of different ways, based on a basic building pattern.

For those who have forgotten their basic biology, recall that cells build proteins by first building DNA, which is then transcribed into RNA, which is finally translated into protein.

Modification of Ig and TCR proteins takes place at the level of the DNA. Before transcription and translation, the DNA itself is rearranged, and this is why so many different Ig and TCR proteins can be made. In a normal process during B-cell development, DNA is snipped from several areas and then glued back together, thus encoding different Ig and TCR proteins. They have found that most B-cell leukemias have rearrangements of the DNA in one or more of these protein encoding regions. Evolutionary, this is a good process, since it results in different proteins to recognize the countless different antigens that humans will face throughout the millennia.

The DNA of these encoding regions is sometimes called "super genes" because they can be rearranged into so many combinations. This super gene consist of "variable" (V), "diverse" (D), and "joining" (J) segments juxtaposed to a "constant" (C) region. As a B-cell develops, it rearranges this DNA by randomly choosing different segments of the V, D, and J regions, cutting them out and pasting them back together in random combinations.

Each B-cell makes a different V-D-J segment. Thus, each normal B-cell in a person’s body has a different size and sequence of V-D-J DNA.

The principle underlying the use of the above-described feature of B-cells to diagnose leukemia is the fact that leukemias are clonal. This means that all leukemia cells derive from one rampant B-cell, and therefore all the leukemia cells in one patient will have the same V-D-J segment, same both in size and sequence. The same is not true for normal B-cell populations.

Both Southern blot and PCR makes use of the V-D-J segment to detect clonal B-cell populations, as explained in the linked sections.

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