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Fig. 2. Different types of DNA FISH probe. (A) CEP on normal metaphase and interphase- spectrum orange labeled centromeric probe for chromosome 11; (B) WCP on normal metaphase spread-simultaneous visualization of whole chromosomes 1, 2, and 4 in spectrum-green, spectrumorange, and spectrum-aqua fluorophores, repectively; (C) subtelomeric probe for the p arm of chromosome 8 labeled with spectrum green. (D) LSI detection of bcr-abl using a design of a dual-color extra signal, which is a combination of spectrum-green LSI for bcr and spectrumorange LSI for the abl-ASS region. Thus, a normal cell shows two orange and two green signals, whereas upon tanslocation of abl to bcr leaving behind intact ASS, in the bone marrow of a CML patient, one traslocation-specific fusion signal is seen along with a large orange signal for intact abl-ASS, a large green signal for intact bcr, and, in addition, a small extra orange signal for intact ASS left behind after bcr-abl translocation. (E) LSI break- apart design for detecting IgH rearragement. Because of the labeling of the probe in dual color (orange and green) in the same probe design, the normal nucleus in a non-Hodgkin's lymphoma bone marrow shows two fusion signals, whereas in an abnormal nucleus, the rearragement results in the separation of two fluorophores into one orange and one green signal along with the normal allele showing fusion signal. Original magnification: X1000. (Parts a, b and c are the courtesy of the Quality Control department of Vysis, Inc., Downers Grove, IL; parts d and e are the courtesy of Dr. Wei-Tong Hsu, Rush Medical College, Rush University, Chicago, IL). (Figure appears in color in insert following p. 172.)

Normal

Fig. 2. Different types of DNA FISH probe. (A) CEP on normal metaphase and interphase- spectrum orange labeled centromeric probe for chromosome 11; (B) WCP on normal metaphase spread-simultaneous visualization of whole chromosomes 1, 2, and 4 in spectrum-green, spectrumorange, and spectrum-aqua fluorophores, repectively; (C) subtelomeric probe for the p arm of chromosome 8 labeled with spectrum green. (D) LSI detection of bcr-abl using a design of a dual-color extra signal, which is a combination of spectrum-green LSI for bcr and spectrumorange LSI for the abl-ASS region. Thus, a normal cell shows two orange and two green signals, whereas upon tanslocation of abl to bcr leaving behind intact ASS, in the bone marrow of a CML patient, one traslocation-specific fusion signal is seen along with a large orange signal for intact abl-ASS, a large green signal for intact bcr, and, in addition, a small extra orange signal for intact ASS left behind after bcr-abl translocation. (E) LSI break- apart design for detecting IgH rearragement. Because of the labeling of the probe in dual color (orange and green) in the same probe design, the normal nucleus in a non-Hodgkin's lymphoma bone marrow shows two fusion signals, whereas in an abnormal nucleus, the rearragement results in the separation of two fluorophores into one orange and one green signal along with the normal allele showing fusion signal. Original magnification: X1000. (Parts a, b and c are the courtesy of the Quality Control department of Vysis, Inc., Downers Grove, IL; parts d and e are the courtesy of Dr. Wei-Tong Hsu, Rush Medical College, Rush University, Chicago, IL). (Figure appears in color in insert following p. 172.)

development in the principles of M-FISH/SKY is called combinatorial binary ratio labeling (COBRA-FISH). In this method, three fluorophores are used pair-wise to label a set of 12 chromosomes, each with a unique fluorochrome ratio. A fourth fluorophore is then added as a binary label to paint the second set of 12 chromosomes (45). Further addition of fluo-rophores has been proposed to provide extended painting options. Using COBRA-FISH, a recent report has shown new cryptic balanced translocations, t(7;17) (q32-34;q23) and t(7;17) (p15;q23) in accelerated phase/blastic crisis chronic myelogenonous leukemia (CML) (46).This approach, however, needs further investigation for sensitivity and specificity.

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