![Figure 2. Chromosomal deletion of TNFAIP3 in cHL detected by interphase cytogenetics. FICTION analyses of representative cHL cases combining CD30 expression (red) and FISH probes for TNFAIP3 and chromosome 6 centromere (blue [b]). In the double-color assays in A–C and E and F, the TNFAIP3 probe is labeled in green (g); in the triple-color assay applied in D, the TNFAIP3 probe gives a g/orange colocalized (co) signal. Two different strategies are used to display double- and triple-color FISH assays in combination with CD30 immunofluorescence; i.e., double-color assays (A–C and E and F) are shown using a triple-color display, whereas a false multicolor display as obtained by Isis software is applied for the triple-color assay (D) to simultaneously show four colors (i.e., CD30 [r], TNFAIP3 [g] and orange, and chromosome 6 centromere [b]). In contrast to the triple-color assay, the quadruple-color assay is based on the overlay of two displays generated by Isis software. To this end, the different channels needed to be individually enhanced so that the final integrated image shows multicolor signals. Fluorescent signals are shown in a false-color display. Arrows point to CD30+ HRS cells. For each case, between 6 and 26 evaluable HRS cells (mean = 12) were considered. (A) HRS cell (case 41) with normal diploid signal pattern (2g + 2b). (B) HRS cell (case 36) exhibiting one copy of TNFAIP3 (1g) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (C) HRS cell (case 16) exhibiting one copy of TNFAIP3 (1g) and two copies of chromosome 6 centromere (2b), indicating the presence of a chromosomal deletion of TNFAIP3. (D) HRS cell (case 19) exhibiting one copy of TNFAIP3 (1co) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (E) HRS cell (case 35) exhibiting two copies of TNFAIP3 (2g) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (F) HRS cell (case 33) lacking signals for TNFAIP3 (0g) but showing two copies of chromosome 6 centromere (2b), indicating the presence of a chromosomal (homozygous?) deletion of TNFAIP3. Neighboring bystander CD30− cells mostly show normal signal patterns (2g or 2co + 2b), although truncation artifacts do occur (some signals are out of the focus plane). Bars, 10 µm.](https://cdn.rupress.org/rup/content_public/journal/jem/206/5/10.1084_jem.20090528/6/jem_20090528_rgb_fig2.jpeg?Expires=1773588327&Signature=XsGL0PhKgJ6oa84rI~ugfA-qoigmjG0Kv0AOEkZGff-00vL2-qOOqMbsiaDcOLAtUB21Sl5viURjVaIrLfY0gamVdp5QAgJUCk~L-AsEbraVznUWHmHBtJxiR8WjCjCyVvUWOy-O8-kWbSzEvJ1q7K7FCoFyZftHqt79-Usk63DwQgHD35GDvDfv2wkqf34FjztMrr4iupQEVI4K-qmvozUc1siL2cMuY9u3VGc-mGL9JKIDpMTzAMH2Vj9m0t3V3OmguBd9ygPFmpuU~g3R24SxIDQ~DXpd68RvfUFTfuIFCxayMH2eCvcdL0HC2f4nYXO-G68oTa7ky6zqQVqejg__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Chromosomal deletion of TNFAIP3 in cHL detected by interphase cytogenetics. FICTION analyses of representative cHL cases combining CD30 expression (red) and FISH probes for TNFAIP3 and chromosome 6 centromere (blue [b]). In the double-color assays in A–C and E and F, the TNFAIP3 probe is labeled in green (g); in the triple-color assay applied in D, the TNFAIP3 probe gives a g/orange colocalized (co) signal. Two different strategies are used to display double- and triple-color FISH assays in combination with CD30 immunofluorescence; i.e., double-color assays (A–C and E and F) are shown using a triple-color display, whereas a false multicolor display as obtained by Isis software is applied for the triple-color assay (D) to simultaneously show four colors (i.e., CD30 [r], TNFAIP3 [g] and orange, and chromosome 6 centromere [b]). In contrast to the triple-color assay, the quadruple-color assay is based on the overlay of two displays generated by Isis software. To this end, the different channels needed to be individually enhanced so that the final integrated image shows multicolor signals. Fluorescent signals are shown in a false-color display. Arrows point to CD30+ HRS cells. For each case, between 6 and 26 evaluable HRS cells (mean = 12) were considered. (A) HRS cell (case 41) with normal diploid signal pattern (2g + 2b). (B) HRS cell (case 36) exhibiting one copy of TNFAIP3 (1g) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (C) HRS cell (case 16) exhibiting one copy of TNFAIP3 (1g) and two copies of chromosome 6 centromere (2b), indicating the presence of a chromosomal deletion of TNFAIP3. (D) HRS cell (case 19) exhibiting one copy of TNFAIP3 (1co) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (E) HRS cell (case 35) exhibiting two copies of TNFAIP3 (2g) and three copies of chromosome 6 centromere (3b), indicating the presence of a chromosomal deletion of TNFAIP3. (F) HRS cell (case 33) lacking signals for TNFAIP3 (0g) but showing two copies of chromosome 6 centromere (2b), indicating the presence of a chromosomal (homozygous?) deletion of TNFAIP3. Neighboring bystander CD30− cells mostly show normal signal patterns (2g or 2co + 2b), although truncation artifacts do occur (some signals are out of the focus plane). Bars, 10 µm.
