Figure 2: Clone validation by FISH on to normal male human metaphase spreads and interphase cells. (A) In situ hybridization of a biotinylated probe prepared from BAC RP11-242E13 exhibited strong signals on the long arm the human Y chromosome. The insert shows an enlarged picture of the Y chromosome with the DAPI picture to the left, the FITC picture in the center and a superposition of both on the right. (B) A digoxigenin-labeled DNA probe prepared from BAC RP11- 348G24 resulted in specific signals on the X chromosome (arrow) in the presence of a significant amount of cross-hybridization to other chromosomes. Please note the high level of cross-hybridization in the interphase nucleus shown in the lower left corner. (C) In contrast to the result shown in (B), a probe prepared from BAC RP11-294C12 bound virtually exclusively to the centromeric heterochromatin of the human X chromosome (arrow). (D) Multiplex analysis of human gonosomes was achieved by combining differently labeled DNA probes in a single hybridization experiment. In this experiment, the Y chromosomal target appears in green, while the X- chromosomal centromeric repeat DNA is shown decorated in red.