Molecular Cytogenetics: FISH

Fluorescence in situ Hybridization (FISH) is a procedure whereby unique DNA fragments are labelled with a fluorochrome and then hybridized to complementary DNA in nuclei or on chromosomes. This unique property of DNA complementary hybridization makes it possible to identify the number of these unique sequences present in a cell. The fluorochrome signals are visualized under a microscope using fluorescent light and equated to the number of a particular chromosome or DNA segment present.

Kindly consult our Probe List for tests that are currently available at Unistel Medical Laboratories.

We are willing to investigate providing new FISH tests as required by customers. Please do not hesitate to contact us on 021 938 9214 should you wish to discuss new tests.

22q Deletion

Ten (10) metaphases from cultured lymphocytes are analyzed for 22q11.2 deletion syndrome.

FISH and the Vysis DiGeorge Region Probe TUPLE1 (HIRA)/LSI ARSA is used.

The probe set includes a probe for the TUPLE1 (HIRA) gene located within the 22q11.2 region (orange) as well as a control probe for the ARSA gene in 22q13.3 region (green).

Deletion of 22q11.2 is present in this metaphase spread

Acute Myeloid Leukemia

Bone marrow (130 cells) is analysed using FISH and Vysis locus specific DNA probes for the detection of t(8;21)(q22;q22), MLL rearrangement, inversion 16 and deletions of 5p15, 5q31, 5q33, 7q31 and 20q12.

The t(8;21) is found in 15% of childhood and 5% of adult AML, making it the most frequent translocation in this disease. Complex translocations involving 8q22 and/or 21q22 with an additional chromosomal region may also occur. Patients with t(8;21)-positive AML usually achieve complete remission after conventional chemotherapy and have been shown to respond particularly well to high-dose cytarabine treatment. The t(8;21) is therefore associated with a favorable prognosis.

Monosomy 5 and del(5q) are associated with resistance to chemotherapy and, hence, with an adverse prognosis. Cases with 5q loss are stratified as high-risk AML. Patients with monosomy7 and del(7q) display poor response to chemotherapy and, thus, have a dismal outcome.

The prognosis for patients with translocations involving MLL have been reported to have various outcomes, ranging from favourable to poor to dismal, depending on the translocation partner. This probe for MLL rearrangements does not identify the translocation partner.

The inv(16)(p13;q22) or t(16;16)(p13;q22), which both result in a CBFB-MYH11 fusion gene, is seen in 5% of paediatric and adult AML with the inversion being much more common. Almost all studies have identified a favourable prognosis for patients with inv(16)/t(16;16)-positive AML.

Deletions of the long arm of chromosome 20 are found in 1-2% of cytogenetically abnormal AML. The del(20q) is the sole anomaly in one-third of cytogenetically abnormal AML. The prognostic impact of del(20q) is somewhat unclear. It is definitely not a favourable abnormality, as it is in MDS, instead it has variously been reported to be associated with either an intermediate or an unfavourable outcome (1).


1.. Heim, S, Mitelman, F. Cancer Cytogenetics 4th edition. New Jersey: Wiley-Blackwell Publishers; 2015. p.64, 74, 77, 80, 83, 91, 95.

Alveolar Rhabdomyosarcoma

Paraffin embedded tissue (200 cells) is analysed using the Vysis FOXO1 (13q14) dual colour, break apart rearrangement probe. Chromosomal rearrangements involving the FOXO1 gene on chromosome 13q14 are associated with alveolar rhabdomyosarcoma (ARMS).

ARMS is characterized by two consistent chromosomal translocations, the common t(2;13)(q35.2;q14) and the variant t(1;13)(p36;q14) resulting in the formation of PAX3-FOXO1 and PAX7-FOXO1 fusion genes respectively (~80% of cases of ARMS).

Hybridization with the Vysis FOXO1 rearrangement probe will identify a chromosomal rearrangement at the FOXO1 gene but not a specific gene-fusion partner.

Amniotic Fluid / Chorionic Villi / Cord Blood FISH Testing
Prenatal Aneuploidy Detection

Uncultured amniotic fluid/chorionic villi/ cord blood (130 cells) is analysed for chromosomal numerical abnormalities using FISH and the Vysis AneuVysion DNA Probe Kit.

Interpretation of the analysis indicates the sex of the fetus as well as numerical abnormal copy numbers of 13, 18 and 21.

Maternal cell contamination cannot be excluded when samples are bloody or the fetus is thought to be female.

Triploidy is estimated to occur in 1% of all conceptions. Triploidy accounts for about 20% of chromosomally abnormal spontaneous abortions. Fetal loss may be related to hydatidiform degeneration of the placenta. Only a small percentage of triploid conceptions survive to term. Parental age does not appear to be a risk factor and the recurrence risk is low. Prenatal diagnosis for subsequent pregnancies is discretionary because triploid conceptions will spontaneously fail(1).

Trisomy 13 (Patau syndrome) has an incidence of 1/9500 live births. There is a strong maternal age effect and 90% are maternal in origin. The recurrence risk for trisomy 13 is low at 0.5%. The median survival is 7-10 days with 5-10% survival to >12 months (1).

Trisomy 18 (Edwards syndrome) has an incidence of 1/7900 livebirths. There is a strong maternal age effect and 85% are maternal in origin. The recurrence risk for trisomy 18 is 1/200 (0.55%)(1).

The risk of trisomy 21 (Down syndrome) increases strongly with maternal age. The estimated recurrence risk for women <39 years is approximately 0,8% with an age related risk thereafter. Some women who have had a previous trisomy 21 conception may have a small increased risk for other aneuploidies.

Please note that these tests do not provide information on whether the trisomy 13, 18 or 21 is as a result of non-disjunction or as a result of an unbalanced rearrangement.


1. Firth, HV et al. Oxford Desk Reference Clinical Genetics, p 524-6, p 534, p 556. Oxford University Press Inc., New York, 2005.

Anaplastic Lymphoma
Fish Analysis For 2p23 Alk Rearrangement

Paraffin wax embedded tissue (200 cells) is analysed using FISH and the Vysis ALK (Anaplastic Lymphoma Kinase) Break Apart FISH Probe kit designed to detect chromosome 2p23 rearrangements.

Rearrangement of the ALK locus has been implicated in the development of Non-Small Cell Lung Cancer (NSCLC), lymphoma and neuroblastoma.


Bone marrow (130 cells) is analysed using FISH and Vysis locus specific DNA probes for the detection of t(1;19)(q23;p13.3), t(9;22)(q34;q11.2), t(12;21)(p13;q22), chromosome 10 to determine hyperdiploidy and rearrangement of 11q23. These chromosome aberrations define a population of patients who are a high-risk prognostic group.

Studies have shown t(12;21) to be the most common translocation in childhood Acute Lymphoblastic Leukemia (ALL), occurring in 25-30% of pre-B/common ALL. The outlook for children with ALL and this abnormality is generally very good, although there are indications that it may be associated with a late relapse (1).

The t(1;19) is one of the most common translocations in ALL, occurring in approximately 6% of cases overall (1). The early association of t(1;19) with a poor outcome has been moderated by more aggressive therapy in modern protocols.

In ALL, the t(9;22) occurs at a higher incidence in adults (25-30%) than children (approximately 2%), increasing exponentially with age (1). In all age groups, the outcome is extremely poor despite high-dose intensified chemotherapy regiments. However, with the recent introduction of the TKI imatinib, the historically poor outcome of BCR-ABL1-positive ALL has improved (2).

  1. Heim, S, Mitelman, F. Cancer Cytogenetics 3rd edition. New Jersey: Wiley-Blackwell Publishers; 2009. p. 248, 260, 264.
  2. Heim, S, Mitelman, F. Cancer Cytogenetics 4th edition. New Jersey: Wiley-Blackwell Publishers; 2015. p. 198, 222.
Breast Cancer
Fish Diagnosis of Her2 Gene Amplification

The breast tissue specimen is analysed for HER2 amplification using the Dako HER2/CEN-17 IQISH probe kit. The amplification of the HER2 gene is a prognostic marker for breast cancer and correlates with poor prognosis and shorter survival rates. The HER2 status is mainly used to identify patients with breast cancer who may benefit from therapy with Herceptin.
Results on enumeration of interphase nuclei from tumour cells are reported as the ratio of average HER2 copy number (red signals) to that of CEP17 (green signals).

The 2013 ASCO/CAP Guidelines state the following:

  • FISH positive – ratio >= 2 or ratio < 2 & copy no. >= 6;
  • FISH equivocal – ratio < 2 & copy no. >= 4 & < 6;
  • FISH negative – ratio < 2 & copy no. < 4
Burkitt’s Lymphoma
Fish Analysis t(8;14)

Paraffin wax embedded tissue/Bone marrow (200 cells) is analysed using FISH and the Vysis LSI IGH/MYC, CEP 8 Tri-color, Dual Fusion Translocation Probe to detect juxtaposition of the IGH locus and the MYC gene region sequences.

The CEP 8 is an indicator for chromosome 8 which is important when MYC amplification and loss of the der(8) chromosome occurs.

t(8;14)(q24;q32) is the most frequently observed MYC region translocation. It is believed to disrupt the normal regulation of the MYC transcription factor by bringing the MYC gene under control of a regulatory element from one of the immunoglobulin loci. The t(8;14) is present in virtually all cases of Burkitt’s lymphoma and is also characteristic of the L3 subtype of acute lymphocytic leukemia (ALL). The translocation can also be found in a minority of diffuse large B-cell lymphomas (DLBCL).

Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma (CLL) FISH Panel

Bone marrow (130 cells) is analysed using FISH and locus specific DNA probes for 11q22.3, CEP12, 13q14.3, 13q34 and 17p13.1 (TP53). This panel is used to detect chromosomal deletions and copy number changes associated with chronic lymphocytic leukemia (CLL). According to the literature aberrations in these chromosomes are important factors in predicting survival (1).

Patients with 17p deletions have the worst prognosis (32 months), followed by patients with 11q22.3 deletions (79 months). Patients who have trisomy 12 have a median survival of 114 months, whereas patients with 13q deletions as the sole abnormality have the longest estimated survival time (133 months).


1. Heim, S, Mitelman, F. Cancer Cytogenetics 4th edition. New Jersey: Wiley-Blackwell Publishers; 2015. p. 269.

Chronic Myelogenous Leukemia
Fish Analysis for Philadelphia Chromosome

Fluorescence in situ hybridisation (FISH) is performed on interphase cells derived from the bone marrow aspirate/peripheral blood. The analysis is done with the Vysis LSI BCR/ABL1 Dual Color, Dual Fusion Translocation Probe Set for detection of the t(9;22)(q34;q11.2) and its variants, resulting in the BCR-ABL1 gene fusion.

The t(9;22)(q34;q11.2) produces the Philadelphia (Ph) chromosome which is the hallmark in 90% of chronic myeloid leukemia (CML) cases and can be used as a diagnostic indicator and to monitor therapy efficiency. The Ph chromosome also occurs in 6% of children and 17% of adults with acute lymphoblastic leukemia (ALL) and is associated with a poor prognosis. The Ph chromosome is rarely observed in other leukemias.

The percentage of the Ph positive clone is the base value against which all future analysis will be compared for disease monitoring.

Follow up FISH analysis is recommended to monitor disease progression.

Diffuse Large B-cell Lymphoma
Fish Analysis BCL2 Rearrangement

Paraffin wax embedded tissue (200 cells) is analysed using FISH and the MetaSystems XL BCL2 break-apart probe on chromosome 18q21.3.

The BCL2 rearrangement is commonly identified in B-cell lymphomas. In particular, the translocation t(14;18)(q32;q21) can be found in 50% of follicular lymphoma (FL), 23.3% of B-cell lymphoma, and about 15% of DLBCL. In FL this translocation results in the BCL2 gene being juxtaposed to the IGH locus at 14q32.33 which leads to overexpression of the anti-apoptotic protein BCL2, and finally to progression to lymphoma. In DLBCL, BCL2 gene overexpression has been implicated in conferring resistance to chemotherapy and has been associated with poor prognosis.

Fish Analysis BCL6 Rearrangement

Paraffin wax embedded tissue (200 cells) is analysed using FISH and the Vysis BCL6 break-apart probe on chromosome 3q27 to detect chromosome breaks associated with a number of different translocations that involve the BCL6 gene.

Rearrangement of the BCL6 gene has been shown to be associated with a favourable outcome in patients with diffuse large B-cell lymphomas.

Fish Analysis for MYC Rearrangements

Paraffin wax embedded tissue (200 cells) is analysed using the Vysis LSI MYC Dual Colour Break Apart Rearrangement probe to detect rearrangements of the MYC gene region on chromosome 8q24.

Translocations involving the MYC region have diagnostic and prognostic importance in B-cell malignancies. In Burkitt’s lymphoma approximately 75%-80% of cases carry t(8;14)IGH-MYC and the remainder are associated with t(8;22)IGL-MYC or t(2;8)IGK-MYC. In approximately 5-10% of diffuse large B-cell lymphoma (DLBCL) patients also have MYC region rearrangements, and detection of these rearrangements has been associated with a poor prognosis.

Diffuse Malignant Peritoneal Mesothelioma

Formalin fixed paraffin embedded tissue (200 cells) is analysed using FISH and the Vysis LSI CDKN2A/CEP 9 Probe to detect the deletion of CDKN2A (p16) located on 9p21.

This test is to detect homozygous (both signals) or heterozygous (one signal) deletions of p16 (9p21). Homozygous deletions of the 9p21 locus, which contains p16, was reported in cell lines derived from many types of human tumours, including lung, breast, brain, bladder and ovary.

This test can also differentiate between Diffuse Malignant Peritoneal Mesothelioma (DMPM) from Mesothelial Hyperplasia (RMH) and Epithelial Ovarian Cancer (EOC). Cut off values for homozygous deletion is set at 10% and for heterozygous deletions at 41% (1).

This test is not recommended for differentiation between Diffuse Malignant Peritoneal Mesothelioma (DMPM) from Pancreatic Ductal Adenocarcinoma (PDAC) and Cholangiocarcinoma (CCA) as homozygous deletions of p16 occur in similar values to that of DMPM.

Very small deletions may occur that do not delete the entire LSI p16 probe target and will not be detected (2).

  1. Ito, T., Hamasaki, M., Matsumoto, S., Hiroshima, K., Tsujimura, T., Kawai, T., Shimao, Y., Marutsuka, K., Moriguchi, S., Maruyama, R., Miyamoto, S. and Nabeshima, K. (2015). p16/CDKN2A FISH in Differentiation of Diffuse Malignant Peritoneal Mesothelioma From Mesothelial Hyperplasia and Epithelial Ovarian Cancer. American Journal of Clinical Pathology, 143(6), pp.830-838.
Ewing Sarcoma
Fish Analysis for Ewing Sarcoma

Formalin-fixed paraffin embedded tissue (200 cells) is analysed using FISH and the EWSR1 (22q12) Dual Colour, Break Apart Rearrangement Probe from Vysis.

Chromosome rearrangements involving the EWSR1 (Ewing sarcoma breakpoint region1) gene on chromosome 22q12 have been observed in several tumour types. Approximately 90% of the translocations involving the EWSR1 gene result in the t(11;22)(q24;q12), which juxtaposes the EWSR1 with the FL1 (Friend leukemia virus integration 1) gene on chromosome 11q24, thus creating a novel fusion gene with oncogenic properties.

Extranodal Marginal Zone B Cell Lymphoma (MALT lymphoma)
Fish Analysis For BIRC3/MALT1 t(11;18)

Paraffin embedded tissue (200 cells) is analysed using FISH and the Vysis LSI BIRC3/MALT1 Dual Color Dual Fusion Probe Kit to detect the t(11;18)(q21;q21) reciprocal translocation involving the BIRC3 (also known as API2) and MALT1 gene regions. The t(11;18)(q21;q21) translocation is the most common chromosomal translocation found in mucosa-associated lymphoid tissue (MALT) lymphoma and is the most common in gastric MALT lymphoma. The t(11;18)(q21;q21) translocation is associated with failure to respond to Helicobacter pylori eradication and is an aggressive disease.

Trisomy 18q21, including MALT1, may be associated with advanced tumour stage and may be a predictor of poor outcome in surgically resected primary gastrointestinal B cell lymphomas (1).


1. Krugmann J. Unfavorable prognosis of patients with trisomy 18q21 detected by fluorescence in situ hybridization in t(11;18) negative, surgically resected, gastrointestinal B cell lymphomas. Journal of Clinical Pathology. 2004;57(4):360-364.

Follicular Lymphoma
Fish Analysis for the Igh/Bcl2 Translocation t(14;18)

Paraffin wax embedded tissue/Bone marrow (200 cells) is analysed using FISH and the Vysis IGH/BCL2 translocation probe to detect the juxtaposition of immunoglobulin heavy chain (IGH) locus and BCL2 gene sequences.

The translocation involving IGH at 14q32 and BCL2 at 18q21, t(14;18)(q32;q21), is found in 80-90% of follicular lymphoma and in 20-30% of diffuse large B-cell lymphoma. Relocation of an IGH transcriptional enhancer next to the BCL2 gene as a result of the t(14;18) is thought to cause constitutive over-expression of the anti-apoptotic BCL2 protein.

Hypereosinophilic Syndrome
Fish for Rearrangement of PDGFRA, PDGFRB and FGFR1

The fluorescence in situ hybridisation (FISH) technique is performed on interphase cells derived from the bone marrow aspirate/peripheral blood received.

Early recognition of hematolymphoid neoplasms associated with rearrangements of PDGFRA, PDGFRB and FGFR1 is important because patients with PDGFRA or PDGFRB rearrangements often respond to tyrosine kinase inhibitor therapy, whereas patients with FGFR1 rearrangements usually do not respond.

The Vysis LSI 4q12 Tri-Color Rearrangement Probe Set is used to detect an approximately 800 kb deletion or rearrangement of the region containing LNX and CHIC2 and subsequent fusion of FIP1L1 and PDGFRA.

It appears that almost all patients with the FIP1L1-PDGFRA fusion are sensitive to tyrosine kinase inhibitors and therefore should be treated with imatinib. Primary or secondary resistance to imatinib is unusual. However, the sensitivity to tyrosine kinase inhibitor rearrangements of PDGFRA with other gene partners or other abnormalities is uncertain at this time. Similarly, additional adjuvant therapy may be needed when patients have disease in the blast phase. (1)

The Vysis PDGFRB Break Apart FISH Probe Kit is intended to detect chromosomal rearrangements involving the platelet derived growth factor receptor beta (PDGFRB) gene at chromosome region 5q32-33 using the fluorescence in situ hybridization (FISH) technique. A rearrangement of the PDGFRB gene can result from a gene fusion to one of as many as twenty different known partner genes. Most hematolymphoid neoplasms associated with translocations of PDGFRB are sensitive to tyrosine kinase inhibitors. Patients with these neoplasms respond well to imatinib therapy with excellent hematopoietic and molecular responses. Primary or secondary resistance to imatinib is very uncommon. Additional adjuvant therapy is needed when patients develop the blast phase of the disease.

The Meta XL FGFR1 Break Apart Probe is used to detect rearrangements involving the Fibroblast Growth Factor Receptor 1 (FGFR1) gene in chromosome region 8p11 using the fluorescence in situ hybridization (FISH) technique. Patients with hematolymphoid neoplasms associated with FGFR1 translocations have an aggressive disease course that is usually not responsive to first-generation tyrosine kinase inhibitor (imatinib) therapy. Prognosis is poor, and aggressive chemotherapy and often stem cell transplantations are needed. (1)

Rapid transformation to acute leukemia, mostly AML, occurs within 1 or 2 years from diagnosis. (2)

  1. Vega F, Medeiros L, Bueso-Ramos C, Arboleda P, Miranda R. Hematolymphoid Neoplasms Associated With Rearrangements of PDGFRA, PDGFRB, and FGFR1. American Journal of Clinical Pathology. 2015;144(3):377-392.
  2. Heim, S, Mitelman, F. Cancer Cytogenetics 4th edition. New Jersey: Wiley-Blackwell Publishers; 2015. p. 187.
Fish Analysis For Mdm2/Cep 12 Amplification

Paraffin wax embedded tissue (100 cells) is analysed using the Vysis MDM2/CEP 12 FISH probe kit. The chromosomal region 12q13-q15 is often affected by translocations and amplifications in soft tissue sarcoma and chronic lymphocytic leukemia. This region includes the mouse double minute 2 (MDM2) gene. MDM2 inhibits p53 transcriptional activity by binding to p53 and moving the protein into the cytoplasm. This results in inactivation of the tumour suppressor and the formation of tumours, which ultimately leads to cancer.

*Ratio >/= 2: Amplified;
Ratio < 2: Not amplified (1)


1. Weaver J., Downs-Kelly E., et al. FISH for MDM2 gene amplification as a diagnostic tool in lipomatous neoplasms. Modern Pathology 2008; 21:943-949.

Lung Cancer
Fish Analysis for EGFR Abnormalities

The paraffin embedded tissue sample (200 cells) is analysed using FISH and the Vysis EGFR/CEP7 Probe Kit. EGFR abnormalities including increased copy number and amplification have been correlated with the development of many solid tumors, including non-small cell lung cancer (NSCLC) which is the leading cause of cancer death worldwide. NSCLC has a 5-year survival rate of approximately 15%.

Inhibition of EGFR by agents that block its tyrosine kinase domain has been demonstrated to reduce proliferation of lung cancer cells, resulting in suppression of tumour growth. The EGFR copy number is considered to be abnormal when high polysomy or amplification is observed.(1)

*High polysomy positive: >/= 4 EGFR copies in >/= 40% of cells;
EGFR amplification positive: Ratio CEP7:EGFR >/= 2 OR >/= 15 EGFR copies in >/= 10%
of cells OR EGFR gene clusters


1. Hirsch et al., ‘EGFR FISH in NSCLC Patients Treated with Cetuximab+Chemotherapy’, J Clin Oncol, 2008, Vol 26(20): 3351-3357.

Fish Analysis for 2p23 Alk Rearrangement

Paraffin wax embedded tissue (200 cells) is analysed using FISH and the Vysis ALK (Anaplastic Lymphoma Kinase) Break Apart FISH Probe kit designed to detect chromosome 2p23 rearrangements.

Rearrangement of the ALK locus has been implicated in the development of Non-Small Cell Lung Cancer (NSCLC), lymphoma and neuroblastoma.

Fish Analysis Ros1 Rearrangements

The paraffin wax embedded tissue (200 cells) is analysed using FISH and the SureFish ROS1 Break-Apart Probe. This probe detects rearrangements of the ROS1 gene (located at chromosomal region 6q22) but not the translocation partner.

ROS1 encodes for a receptor tyrosine kinase and rearrangements of this gene results in constitutively activated downstream signalling of oncogenic pathways. Rearrangements are not common and are found in 0,6-1,8% of patients with non-small cell lung cancer (NSCLC) (1).

Samples are considered positive if more than 15% of cells showed split 5’ROS1 (Spec. Orange) and 3’ROS1 (Spec. Green) signals (typical rearranged pattern) or isolated 3’ROS1 signals (atypical rearranged pattern).

Gain is defined as a mean copy number greater than 3 fused signals in >40% of nuclei, and amplification as the presence of >15 copies of ROS1 per cell (high polysomy) or clusters, in a minimum of 15% of analyzed cells.


1. Clave S et al. ROS1 Copy number alterations are frequent in non-small cell lung cancer. Oncotarget. 2016:7(7): 8019-8028

Mantle Cell Lymphoma
Fish Analysis t(11;14)

Paraffin embedded formalin fixed tissue/ Bone marrow smear (200 cells) is analysed using FISH and locus specific DNA probes for the IGH locus on 14q32 and the cyclin D1 gene on 11q13. About 95% of patients with mantle cell lymphoma (MCL) have the t(11;14)(q13;q32), thus distinguishing mantle cell lymphoma from other small B cell lymphomas.

This translocation is thought to drive the overexpression of cyclin D1 by the juxtaposition of a transcriptional enhancer from the IGH locus (14q32) next to the CCND1 gene (11q13).

Multiple myeloma (MM) FISH Primary Panel
Fish Analysis for Multiple Myeloma

FISH and Metasystems XL locus-specific DNA probes were used for the detection of deletions of chromosome 17p13.1 (TP53) and chromosome 1p32.3, the amplification/gain of chromosome 1q21, and the rearrangement of the IgH locus on chromosomal region 14q32.3.

The deletion of chromosome 17p13.1 (TP53) define a population of -25% of Multiple Myeloma patients who are in a high-risk prognostic group.

Locus rearrangement of 14q32 IgH was associated with more patients in remission indicating its good impact on patients’ prognosis.(3)

The CDKN2C deletion of the gene in chromosome band 1p32.3, strongly affects cell-cycle regulation and MM pathogeneses. This is a rare event (<10%) and is considered as an adverse prognostic factor. Whereas amplification/gains of chromosome band 1q21 (Amp1q21) is one of the most recurrent chromosomal aberrations in MM and the amplification and overexpression of the CKS1B gene in chromosome band 1q21 has been associated with an aggressive clinical course in MM and is considered an adverse prognostic factor. (1) It has been suggested that gains of 1q21 is related to an advanced phenotype of MM and therefore may be associated with disease progression.(2)

If 14q32.3 (IgH) rearrangement is present then the sample must be probed for t(4;14), t(14;16) and/or t(14;20) to detect the chromosomal rearrangement partner.

  1. Metaprobe catalogue XL CDKN2C/CKS1B reference nr D-5099-100-OG
  2. Hanamura I, Steward JP, Huang Y, Zhan F, Santra M, Sawyer RJ, Hollmig K, Zangarri M, Pineda-Roman M, van Rhee F, Cavallo F, Burington B, Crowley J, Tricot G, Barlogie B, Shaughnessy JD Jr, et al. Frequent gain of chromosome band 1q21 in plasma-cell dyscrasias detected by fluorescence in situ hybridization: incidence increases from MGUS to relapsed myeloma and is related to prognosis and disease progression following tandem stem-cell transplantation. Blood 2006;108(5): 1724-32
  3. Moussa, M., Ali, R., Elswefy, D., Eman, N., Kassem, N. and Mogy, M. (2014). Prognostic value of 13q14 deletion and IgH 14q32 rearrangement by interphase fluorescence in situ hybridization in patients with multiple myeloma. Journal of Applied Hematology, 5(4), p.141.
Myelodysplastic Disorder
Fish Analysis for Mds

Bone marrow (130 cells) is analysed using FISH and Vysis locus specific DNA probes for CEP7, 7q31, CEP8 and 20q12 and MetaSystems locus specific DNA probes for 5p15, 5q31 and 5q33.

In de novo MDS, rearrangements resulting in the loss of 5q have been reported in 10-20% of patients. In therapy related MDS (t-MDS), this figure rises to 40%. Del(5q) in a complex karyotype implies a poor prognosis. However in the 5q- syndrome, which has 5q- as the sole karyotypic abnormality or with one additional abnormality, the prognosis is relatively good with a low risk of leukemic transformation.

Loss of chromosome 7 as well as deletions of 7q are common abnormalities of myeloid diseases. Del(7q) as the sole abnormality has an intermediate prognosis. The incidence of trisomy 8 in MDS is ~10%. Deletion of 20q12 is also a common abnormality in malignant myeloid disorders, noted in ~5% of MDS cases and 7% of t-MDS cases. Although deletion 20q is associated with a favorable diagnosis when noted as a sole abnormality, it is associated with a poor prognosis when observed in the setting of a complex karyotype (1).


1. Heim, S, Mitelman, F. Cancer Cytogenetics 4th edition. New Jersey: Wiley-Blackwell Publishers; 2015. p. 131-136.

Myxoid Liposarcoma
Fish Analysis for DDIT3 (Dna-Damage-Inducible Transcript 3)

Formalin fixed paraffin embedded tissue is analyzed using FISH and the Vysis LSI DDIT3 Dual Color Break Apart Probe designed to detect rearrangement of the LSI DDIT3 probe target located at chromosome 12q13.1-13.3.

This gene encodes a member of the CCAAT/enhancer-binding protein (C/EBP) family of transcription factors. The protein functions as a dominant-negative inhibitor by forming heterodimers with other C/EBP members, such as C/EBP and LAP (liver activator protein), and preventing their DNA binding activity. The protein is implicated in adipogenesis and erythropoiesis, is activated by endoplasmic reticulum stress, and promotes apoptosis. Fusion of this gene and FUS on chromosome 16 or EWSR1 on chromosome 22 induced by translocation generates chimeric proteins in myxoid liposarcomas or Ewing sarcoma.

N-MYC Fish Analysis

Paraffin-embedded tissue (200 cells) is analysed using FISH and the Vysis LSI N-MYC/CEP2 probe. This probe contains unique DNA sequences specific to the N-MYC oncogene located within the 2p24.1 region of chromosome 2 and may be used to detect the N-MYC oncogene copy number.

Frequency of N-MYC amplification in neuroblastoma (NBL) is high and is rarely observed in ganglioneuroblastoma (GNBL) and ganglioneuroma (GN). This suggests N-MYC gene plays an important role in neuroblastic tumor differentiation (1).

Amplification of N-MYC oncogene is an established marker indicating aggressive tumor progression of neuroblastoma (NBL). Amplification is correlated with decreased overall survival in NBL (1).

Patients with N-MYC gene gain had a significantly longer mean survival time than those with normal N-MYC gene copy number (1).

NBL cases with N-MYC gene gain as well as gain of CEP2, suggesting polyploidy, were associated with long-term disease-free survival when treated with cyclophosphamide-doxorubicin. While diploidy invariably predicted early treatment failure (1).

*N-MYC gene copy number alteration can be classified into four groups:
No Alterations: Cells with 2 copies of N-MYC and 2 copies of CEP2;
Gain: The number of N-MYC signals is 1-9 copies more than CEP2 signals;
Amplification: The number of N-MYC signals is at least 10 copies greater than signals for CEP2.
Loss/Imbalance: Presence of at least 2 N-MYC and increased CEP2 signals.


1. Wang et al.: Copy number gain of MYCN gene is a recurrent genetic aberration and favorable prognostic factor in Chinese pediatric neuroblastoma patients. Diagnostic Pathology 2013 8:5.

Oligodendroglial Tumours
Fish Analysis 1p36 19q13 Codeletion

Paraffin wax embedded tissue (200 cells) is analysed using FISH and the Vysis LSI 1p36/LSI 1q25 and LSI 19q13/19p13 Dual-Colour Probes. The codeletion of 1p 19q is an early genomic event in oligodendroglial tumours. It is associated with good prognosis and a superior response to therapy (1). Please note that this FISH assay only uses probes binding at two loci on each chromosome 1 and 19. Therefore, although detection of a codeletion is consistent with loss of both 1p and 19q, it may also represent interstitial deletions of target loci only (2). The cut-off for deletion-positive cells is 25%, additionally, a ratio of <0,8 can be included in the interpretation.


Normal signals – two (2) copies of the control probe and two (2) copies of the area of interest (in this case, either 1p36 or 19q13).
Absolute deletion – two (2) copies of the control probe and one (1) copy of the area of interest signifying a deletion of the area of interest.
Associated deletion – multiples of the copies signifying the absolute deletion, for example, four (4) copies of the control probe and two (2) copies of the area of interest.
Relative deletion – similar to the associated deletion but not representative of a true deletion since the absolute deletion is not present.
Imbalance – Copies that are not proportional, if the majority of cells show an imbalance this is an inconclusive result.

  1. Woehrer, A et al. FISH-based detection of 1p 19q codeletion in oligodendroglial tumors. Clinical Neuropathology 30:47-55.
  2. UK NEQAS Guidelines for the analysis and interpretation of 1p 19q codeletion testing.
Prader-Willi and Angelman Syndrome

Prader-Willi syndrome is a complex genetic condition that affects many parts of the body. In infancy, this condition is characterized by weak muscle tone (hypotonia), feeding difficulties, poor growth, and delayed development. Beginning in childhood, affected individuals develop an insatiable appetite, which leads to chronic overeating (hyperphagia) and obesity. Some people with Prader-Willi syndrome, particularly those with obesity, also develop type 2 diabetes. People with Prader-Willi syndrome typically have mild to moderate intellectual impairment and learning disabilities as well as behavioural problems.

Angelman syndrome is also a complex genetic disorder that primarily affects the nervous system. Characteristic features of this condition include delayed development, intellectual disability, severe speech impairment, and problems with movement and balance (ataxia).

PRADER-WILLI SYNDROME (PWS) and ANGELMAN SYNDROME (AS) are distinct neurogenetic disorders, however both usually caused by chromosomal deletions on chromosome 15q11 or by uniparental disomy (UPD). Normal individuals have one copy of the unmethylated paternal allele and one copy of the methylated maternal allele. Two methylated alleles from a maternal disomy will appear in the case of PWS or two unmethylated alleles from a paternal disomy will indicate AS.

FISH testing cannot detect methylation patterns but molecular tests using MLPA can be carried out.


Soft Tissue Sarcoma
Fish Analysis FUS Gene Rearrangement

Paraffin wax embedded tissue (200 cells) is analyzed using FISH and the Vysis FUS (16p11) Dual Color, Break-Apart Rearrangement Probe to identify chromosomal rearrangements in the FUS gene but not the specific gene partners.

Chromosomal rearrangements involving the FUS gene located on chromosome 16p11 have been observed in many tumor types including soft tissue sarcomas (STS). Different types of STS are characterized by specific chromosomal translocations including t(12;16)(q13;p11) FUS-DDIT3 (Myxoid liposarcoma), t(12;16) FUS-ATF1 (Angiomatoid-fibrous histio- cytoma) and t(7;16)(q32-34;p11) FUS CREB3L2 (Low grade fibromyxoid sarcoma). The resulting chimeric fusion proteins are mainly transactivators exerting deregulation of differentiation control on tumor-target cell.

Synovial Sarcoma
Fish Analysis for Ss18 Rearrangement

Paraffin wax embedded tissue (100 cells) is analyzed using FISH and the LSI SS18 Dual Color Break Apart probe to detect rearrangements involving the SS18 gene located in the breakpoint region of chromosome 18q11.2. These rearrangements are common among synovial sarcoma soft tissue tumors. This probe will identify translocations of 18q11.2, but not the specific translocation partner.

Fish Analysis for Williams Syndrome

Ten (10) metaphases from cultured lymphocytes are analyzed for Williams syndrome. FISH and the Vysis Williams Region Probe LSI ELN/LSI D7S486, D7S522 is used.

The probe set includes a probe for the Elastin (ELN) gene located within the 7q11.23 region as well as a control probe for D7S486 and D7S522 in region 7q31. Williams syndrome is the result of microdeletion of the 7q11.23 region which leads to the loss of many genes including the ELN gene.