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Bone Marrow Tests

Bone marrow is sampled using a technique called bone marrow aspiration. During this procedure, a thin hollow needle with a syringe attachment is used to suction up (aspirate) a teaspoon-sized sample of liquid bone marrow from the back of the hip bone. A larger needle then is employed to obtain a bone marrow biopsy (core biopsy), which removes roughly a 1/16 inch cylindrical piece of bone marrow from the hip site. After the bone marrow samples are obtained, they are examined by many physician specialists, including a pathologist (disease diagnosis specialist, who examines samples under a microscope), hematologist (blood specialist), and oncologist (cancer specialist).

Microscopic examination is performed on samples of the bone marrow, as well as any samples of the blood, cerebrospinal fluid, or lymph node tissue. The bone marrow cells are evaluated according to their size, shape, and content of granules (cellular enzymes that help some leukocytes to destroy germs).

Then these cells are classified with respect to maturity:

1. Mature cells are normal cells of the circulating blood, which are functional infection-fighters that can no longer reproduce.
2. Immature cells are undeveloped blood cells that, although poor infection-fighters, are still able to reproduce.
3. Blast cells are the most immature form of bone marrow cells.

Bone marrow samples also are categorized according to their number of cells (cellularity), because abnormal tissue may contain uncommon proportions of blood-forming (hematopoietic) versus fat cells. Hypercellular marrow holds too many hematopoietic cells, whereas hypocellular marrow holds too few hematopoietic cells.

Cytochemistry involves the identification of cell chemical components to distinguish different types of leukemia. Cytochemical tests often use special colored dyes (stains) that are only visible under a microscope. For example, one stain turns the granules of most acute myelogenous leukemia (AML) cells black, although acute lymphocytic leukemia (ALL) cells are unaffected by this substance. In the past, leukocyte alkaline phosphate (LAP) or neutrophil alkaline phophatase (NAP) tests were used to distinguish CML from other types of leukemia and noncancerous blood disorders; however, these assays no longer are considered particularly helpful in diagnosis, except in the absence of cytogenetic (cell genetic material) or other studies.

Flow cytometry is a computer-assisted technique in which bone marrow or other cells are treated with special antibodies and then placed in front of a laser beam. Some types of leukemia cells contain binding molecules called receptors that cause the antibodies to "label" (stick to) them. Laser treatment makes the antibody-coated cells fluorescent.

Light that is given off by these cells undergoes computer measurement and analysis and leukemia cells are counted and categorized using this method.

Immunocytochemistry, like flow cytometry, uses antibodies to treat the bone marrow or biopsy samples. Unlike flow cytometry, computers and lasers are not needed for this procedure. Instead, the sample is prepared so that specific types of cells undergo a color change that can be identified under a microscope. Immunocytochemistry allows the pathologist to identify specific types of leukemia.

Cytogenetic studies employ a variety of techniques for cell culture, slide-making, and preparation of chromosomes (genetic material) for microscopic examination. Bone marrow aspirate is the preferred tissue for most blood disorders; however, if marrow is unavailable, blood samples may be used when there are enough circulating blast cells. In cases of chronic lymphocytic leukemia (CLL), blood samples are essential; in lymphoma cases, lymph node samples can provide more information.

Researchers have found that leukemia cells often contain genetic defects known as translocations, inversions, deletions, and additions. Translocations are genetic errors that result when parts of two chromosomes are exchanged. Inversions are produced when part of a chromosome becomes inverted (upside down) and the order of its genetic material is reversed. Deletions occur when part of a chromosome is missing, and additions are caused by duplications of all or part of a chromosome.

Fluorescent in situ hybridization (FISH) is a type of cytogenetic test that can be used to detect chromosome changes and diagnose certain types of leukemia (e.g., CLL). By identifying specific deletions (e.g., part of chromosome 11, 13, 17), the test also can be used to help determine the expected outlook (prognosis) for patients who have leukemia. For example, a chromosome 13 deletion usually is associated with slower disease progression and a better outlook than chromosome 11 or 17 deletions.

Chronic myelogenous leukemia (CML) was the first type of cancer to show a consistent cytogenetic abnormality. This abnormality—a translocation between chromosomes 9 and 22 (written as t [9;22])—is known as the Philadelphia chromosome (Ph1). The Philadelphia chromosome causes uncontrolled reproduction and proliferation of all types of white blood cells and platelets.

Immunophenotyping is the classification of cell types according to their immunologic characteristics. With the development of a form of testing known as monoclonal antibody (MAb) technology, types of leukemia cell lines are now better defined. Numerous antibody reagants (substances used to create chemical reactions) have been identified. Some reagants recognize specific "clusters of differentiation" (CD); for example, CD79 recognizes B-cells, CD3 recognizes T-cells, and antimyeloperoxidase recognizes myeloid cells. Other useful, but less specific reagants are CD19, CD22,CD5, CD7, CD13, CD33, glycophorin, and CD61.

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