Re: How do hospitals find the correct tissue type done for transplants?

Answered by T.J. Cradick
tjc@itsa.ucsf.edu
Submitted by Mimi S. Mong
*what is tissue typing?

*molecular methods for tissue typing

*molecular diagnostic

The immune system seeks to differentiate between self and non-self so that it can work to eliminate the non-self. Certain molecules called Major Histocompatability Complexes, or MHC, are used to present pieces of protein (peptides) to the cells of our immune system. These proteins were named MHC when it was discovered that they mattered in preventing tissue rejection. It was later learned that they are important in the functioning of our immune system. The cells of our immune system recognize pieces of proteins held by the MHC as being either self or non-self. Our immune system's T cells are activated to mount a defense against foreign, i.e. non-self, proteins presented by our MHC on our cells. Our immune cells are also activated to attack cells with different MHC. Because every person has many different MHC it is difficult to locate an unrelated individual with the same MHC on their cells.

The process of locating possible donors is called tissue typing. To determine if tissue can be transfered between person A to person B, a series of plastic wells (indentations) are coated with white blood cells from each person. For our example, person A's white blood cells are coated on wells A1, A2,A3, etc and person B's are coated on B1,B2,B3 etc.. Then a series of antibodies (1 to X ) are added to a column of wells so that each pair of wells (A1, B1) gets identical pools of antibodies. These pools of antibodies with identical specificity are called monoclonal antibodies. After the antibodies are added, the wells are incubated to allow time for binding. The wells are then washed to remove the non-binding antibodies. Complement can then be added. Complement is used in our bodies to kill cells that are bound by antibodies. Similarly, complement is used in this assay to destroys cell that have antibodies bound to them. Then dye is used to stain the dead cells. If both peoples cells are bound by antibody 1, the white blood cells in each well (A1 and B1) will have antibodies bound to them and therefore will be killed by complement and stained by dye.

Alternatively, a different readout can be used to detect binding antibodies. Secondary antibodies are added that don't bind the white blood cells, but do bind to the primary (first) antibody. These secondary antibodies will only remain in wells if the primary antibody is bound to the cells, otherwise they are washed out. This secondary antibody can be hooked up to an enzyme that provides the readout and is why this method is called an ELISA, which stands for enzyme- linked immunosorbent assay. Some of these enzymes can be detected because they cause a color change.
ELISA:
(white blood cell) >====primary ab >==== secondary ab-(enzyme) ~color change

primary antibody binds cell, secondary antibody binds primary, enzyme causes color change which is detected.

HLA typing is the total of binding or non- binding of these sets of antibodies specific for different HLA allels. Usually one person is not compared to only one other, but is compared to many other people's resuslts done previously. Once a person is tested, the databank is searched for other people that have the same antibodies bidn to their cells. HLA compatible donors would be bound by the same antibodies. Transplantation may still be possible even if they aren't bound by all the same antibodies. To see if this may be possible other tests allows quantifying the immune reaction to the other person's cells your body recognizes as different. Similarly, cells that come with the grafted tissue can recognize the host persons cells as different leading to a process called graft versus host rejection. For these reasons, modern doctors often use drugs to minimize rejection.