Method Description

A cell count is performed to establish a plating volume. Based on the cell count, a corresponding volume of blood is added to 2 culture flasks containing culture medium and incubated for 24 to 48 hours at 37° C. In the harvest process, the cells are exposed to Colcemid and hypotonic solution and then fixed with glacial acetic acid and methanol. Metaphase cells are dropped onto microscope slides and are stained by G-banding. Other staining methods are employed as needed. Twenty metaphases are usually examined. If a clone is suspected, but not confirmed within 20 metaphases, 30 metaphases will be analyzed. Minimal evidence for the presence of an abnormal clone is defined as 2 or more metaphases with the same structural abnormality or chromosome gain (trisomy), or 3 or more metaphases lacking the same chromosome. All cells analyzed are captured using a computerized imaging system, and 1 or more karyograms from each clone are prepared to document the abnormality and to permit systematic interpretation of the abnormalities

When a specimen is received from a patient 30 years of age or older with a reason for testing of chronic lymphocytic leukemia, small lymphocytic lymphoma, lymphocytosis, or Waldenstrom macroglobulinemia, a CpG-stimulated culture will be added and 10 additional cells will be analyzed. Additional metaphases may be analyzed from the unstimulated or CpG-stimulated cell cultures if necessary to provide an accurate interpretation. All metaphases are captured using a computerized imaging system, and 1 or more karyograms from each clone are prepared to document the type of abnormality and to permit systematic interpretation of the abnormalities.(Arsham, Marilyn S., et al. eds. The AGT Cytogenetics Laboratory Manual. 4th ed. Wiley-Blackwell, 2017)