Molecular Cytogenetic/FISH
WHAT IS FISH? FISH is an acronym for Fluorescent In Situ Hybridization. This test blends cytogenetic and DNA testing. This molecular cytogenetic test represents an accurate, rapid, and direct approach for identifying the presence of a DNA probe/gene/segment on a chromosome or the presence of a chromosome within interphase nuclei. This technique allows for the rapid identification of trisomies (2-6 hours) in peripheral blood, prenatal samples or bone marrow.
The Molecular Cytogenetic Laboratory of the Wake Forest University Health Sciences was established in 1991 through a North Carolina Baptist Hospital Technology grant and Brenner Children's Hospital. This facility offers state-of-the-art technology for both clinical and research activities. The Molecular Cytogenetic Laboratory has been involved in FDA clinical validation studies using various diagnostic FISH-DNA probes. The laboratory can perform a wide array of molecular cytogenetic techniques, both clinical and research, including single or multiple gene FISH, whole chromosome painting (WCP), multicolor FISH (M-FISH), subtelomeric FISH, chromosome microdissection with reverse PCR-FISH, and comparative genomic hybridization (CGH).
In most cases, the FISH probes used for clinical testing are FDA approved. Those not approved go through rigorous testing by both the manufacturer and this laboratory prior their use. The WFUHS Molecular Cytogenetic Laboratory does not use home brew probes for clinical testing because of validation issues.
The Molecular Cytogenetic Laboratory also performs research involving physical mapping of DNA gene probes using FISH which is the most accurate, rapid and direct approach for identifying the specific chromosome location of a segment of human DNA and/or determine the physical gene order of two or more loci on a chromosome. The Molecular Cytogenetic Laboratory is currently developing and validating the use of genomic DNA microarray analysis for molecular karyotyping. This potentially can identify deletions or duplications as small as 1Mb.
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Test: Prenatal Aneuploidy Detection - Amniotic Fluid/CVS/PUBS
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To rapidly identify one of the primary chromosome aneuploidies involving chromosomes 13, 18, 21, X, and/or Y. This analysis allows the identification of the following chromosomal syndromes: Down syndrome - trisomy 21; Edwards syndrome - trisomy 18; Klinefelter syndrome - 47,XXY; Patau syndrome - trisomy 13; Turner syndrome - 45,X; other sex chromosome aneuploidies including 47,XXX and 47,XYY; and to resolve cases of ambiguous genitalia.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
SUBMIT ONLY 1 OF THE FOLLOWING SPECIMENS:
Amniotic Fluid - Collection Information
Chorionic Villus Sample - Collection Information
PUBS
Reflex Testing: Routine Chromosome analysis
Confirmation will be performed on metaphase chromosomes thus requiring culturing of cells.
Turn Around Time for FISH: 6-18 hours
Probes: 13q14(Rb1), 18cen(D18Z1), 21q22.13-q22.2(D21S259,D21S341,D21S342), Xcen(DXZ1), Ycen(DYZ3)
CPT Code Information:
88271x5 DNA probe, each
88274x5 Interphase in situ hybridization, analyze 25-99 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Aneuploidy Detection - Peripheral Blood/Bone Marrow/Skin Biopsy/POC
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To rapidly identify one of the primary chromosome aneuploidies involving chromosome 13, 18, 21, X, and/or Y. This analysis allows the identification of the following chromosomal syndromes: Down syndrome - trisomy 21; Edwards syndrome - trisomy 18; Klinefelter syndrome - 47,XXY; Patau syndrome - trisomy 13; Turner syndrome - 45,X; other sex chromosome aneuploidies including 47,XXX and 47,XYY; and to resolve cases of ambiguous genitalia.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: In cases where metaphase cells are necessary for accurate analysis, cells will be cultured.
Peripheral Blood (PB) – Collection Information
Bone Marrow (BM) – Collection Information
Autopsy Tissue/Skin Biopsy – Collection Information
Products of Conception (POC) – Collection Information
Stored Pellets
Reflex Testing: Routine cytogenetic testing is strongly recommended.
Turn Around Time: 3-5 hours
Probes: 13q14(Rb1), 18cen(D18Z1), 21q22.13-q22.2(D21S259, D21S341, D21S342), Xcen(DXZ1), Ycen(DYZ3)
CPT Code Information:
88271x5 DNA probe, each
88274x5 Interphase in situ hybridization, analyze 25-99 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Microdeletion Syndromes - Amniotic Fluid / CVS / Peripheral Blood
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To identify chromosome anomalies such a microdeletions/ duplication not identifiable by routine molecular cytogenetic analysis.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: Routine cytogenetic testing is strongly recommended to identify the possibility of other clinically associated chromosome abnormalities. In addition, an accurate FISH analysis typically requires metaphase chromosomes thus requiring cell culturing. However, FISH analysis can be performed on interphase cells in special cases.
Amniotic Fluid (AF) - Collection Information
Chorionic Villus Sample (CVS) - Collection Information
Peripheral Blood (PB) - Collection Information
Stored Pellets - Previously studied samples for cytogenetic analysis are kept for 6 months. These samples may be used for FISH analysis. After that time, an additional sample from the patient will be required.
Reflex Testing: Routine cytogenetic analysis if requested
Probes:
| Disorder | Deleted Region | Disorder | Deleted Region |
| Wolf Hirschhorn | 4p | Miller-Dieker | 17p13 |
| Cri du Chat | 5p | Smith-Magenis | 17p11 |
| Williams | 7q22 | DiGeorge/VCF DiGeorgeII | 22q11 10p24 |
| Retinoblastoma | 13q14 | STS | Xp22.3 |
| Angelman | 15q12 | Kallmans | Xp22.3 |
| Prader-Wili | 15q12 | SRY | Xp22.3 |
Turn Around Time: 2-4 days
CPT Code Information:
88271x2 DNA probe, each
88273 Chromosomal in situ chromosomal hybridization analyze 10-30 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Metaphase Chromosome Painting – Amniotic Fluid/CVS/Blood/Bone Marrow
Multicolored Fluorescence In Situ Hybridization (M-FISH)
Purpose of Test: To identify chromosome abnormalities not otherwise identifiable with routine cytogenetics or single FISH probes.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: Routine chromosome analysis must be performed prior to M-FISH testing. If routine chromosome analysis has been previously performed at another reference laboratory, a copy of the karyotype(s) and the final report must accompany the specimen.
SUBMIT ONLY 1 OF THE FOLLOWING SPECIMENS:
Amniotic Fluid (AF) - Collection Information
Chorionic Villus Sample (CVS) - Collection Information
Peripheral Blood (PB) - Collection Information
Bone Marrow (BM) - Collection Information
Stored Pellets - Samples on previously studied samples for cytogenetic analysis are kept for 6 months. These samples may be used for FISH analysis. After that time, an additional sample from the patient will be required.
Reflex Testing: Routine cytogenetic analysis if requested
Probes: Individual whole chromosome DNA probe sets that depict each chromosome in a unique color.
Turn Around Time: 1 week
CPT Code Information:
88271x23 Molecular cytogenetics; DNA probe, each (eg FISH)
88273x23 Chromosomal in situ hybridize, analyze 10-30 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: SubTelomeric Region Anomalies – Amniotic Fluid/CVS/Blood
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To identify chromosome anomalies involving the very distal chromosome regions such as submicroscopic translocations, deletions, or duplications.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: Routine chromosome analysis must be performed prior to subtelomeric testing. If routine chromosome analysis has been previously performed at another reference laboratory, a copy of the karyotype(s) and the final report must accompany the specimen. In addition, an accurate FISH analysis typically requires metaphase chromosomes thus requiring cell culturing. However, FISH analysis can be performed on interphase cells in special cases.
SUBMIT ONLY 1 OF THE FOLLOWING SPECIMENS:
Amniotic Fluid (AF) - Collection Information
Chorionic Villus Sample (CVS) - Collection Information
Peripheral Blood (PB) - Collection Information
Stored Pellets - Samples on previously studied samples for cytogenetic analysis are kept for 6 months. These samples may be used for FISH analysis. After that time, an additional sample from the patient will be required.
Probes:
| 1p CEB108/T7 | 6p 6P TEL48 | 11p D11S2071 | 18p D18S552 |
| 1q D1S3738 | 6q VIJyRM2158 | 11q VIJyRM2072 | 18q VIJyRM2050 |
| 2p VIJyRM2052 | 7p VIJyRM2185 | 12p SAVH27 | 19p 129F16/SP |
| 2q D2S447 | 7q VYJyRM2000 | 12q VIJyRM2196 | 19q D19S238E |
| 3p D3S4559 | 8p D8S504 | 13q D13S327
STS X58399 | 20p D20S1157 |
| 3q D3S4560 | 8q VIJyRM2053 | 14q SHGC | 20q 20Q TEL14 |
| 4p D4S3359 | 9p 305J7 T7 | 15q D15S396 | 21q IJyRM2029 |
| 4q D4S2930 | 9q D9S325 | 16p D16S3400 | 22q MS607, ACR
DXYS129 |
| 5p C84C11/T3 | 10p Z96139 | 16q 16Q TEL013 | Xp/Yp DXYS129 |
| 5q D5S2907 | 10q D10S249 | 17p D17S2199 | Xq/Yq Cdy 16c07 |
| | | 17q AFMZ17yD10,D17S928 | |
Turn Around Time: 4-7 days
CPT Code Information:
88271x15 DNA probe, each
88275x15 Interphase in situ hybridization, analyze 100-300 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: X and Y (Sex) Aneuploidy Detection – Buccal Smear
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To rapidly detect the presence of a sex chromosome (X/Y) mosaicism in a tissue other than peripheral blood. Sex chromosome mosaicism is common among recognized syndromes. For example, individuals with Turner syndrome will have a single X chromosome in 55% of cases and the remaining 45% are either variants or mosaics. FISH using X and Y chromosome specific DNA probes allows for the rapid analysis of a large number of cells thus detecting sex chromosome aneuploidy as well as mosaicism. The limitation, however, is that structural chromosome abnormalities of X and Y will not be detected by this technique.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Buccal Smear
- Patient should rinse out mouth vigorously with water before obtaining specimen.
- Remove the Cyto-Pak Brush (can be supplied by lab) from the container, touch only the "stick" end.
- Using MEDIUM pressure, swab inside of cheek with a Cyto-Pak Brush by rapidly rotating bristles across cheek surface for 10 seconds.
- Place brush back into the original container, re-cap, seal with tape, and label container with patient’s name.
- Repeat collection of specimen from the other cheek.
Turn Around Time: 1–2 days
Probes: X and Y centromeric [ Xcen(DXZ1), Ycen(DYZ3] specific probes
Alternative: SRY – Sex Region on the Y chromosomes probe (microdeletion).
CPT Code Information:
88271x2 DNA probe, each
88275 Interphase in situ hybridization analyze 100-300 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Hematology (Leukemia) Rearrangements / Aneuploidy – Bone Marrow / BoneCore / Peripheral Blood
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To rapidly identify acquired chromosome aneuploidies, rearrangements, and/or deletions that are specifically associated with a particular leukemic subtype. This analysis uses DNA probes for either a chromosome(s) or genes involved in a specific chromosome rearrangement. The results can be used for diagnosis, prognosis, or monitoring the disease process or therapeutic response/relapse.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: Routine cytogenetic analysis is strongly recommended to identify possible additional hematological-associated chromosome abnormalities which would not be identified through FISH analysis.
SUBMIT ONLY 1 OF THE FOLLOWING SPECIMENS:
Blood – Collection Information
Done on interphase or metaphase cells requiring cytogenetic confirmation.
Bone Marrow/Core – Collection Information
Done on interphase or metaphase cells requiring cytogenetic confirmation.
Unstimulated Peripheral Blood – Collection Information
Done on interphase or metaphase cells requiring cytogenetic confirmation.
Solid Tumor / FNA – Collection Information
Done on interphase or metaphase cells requiring cytogenetic confirmation.
Pleural Fluid / CNS / Lymph Node – Collection Information
Done on interphase or metaphase cells requiring cytogenetic confirmation.
Turn Around Time: 1-2 days; stats 3-5 hours
Probe(s): See the List (PDF)
CPT Code Information:
88271x2 DNA probe, each
88275x2 Interphase in situ hybridization
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Breast Cancer HER-2/neu Amplification – Solid Tumor Tissue
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To detect amplification of the HER-2/neu gene which has been associated with breast cancer prognosis and treatment decisions.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Paraffin-Embedded Tissue Block - Formalin-fixed, paraffin-embedded breast cancer tissue block. Send at least 4 slides with tissue sections cut at ~4um thick and put on positively charged slides. Include 1 hematoxylin-and-eosin stained slide with abnormal region clearly outlined with a marker. If a tissue block is sent, the tissue will be sectioned and slides will be prepared at an additional cost.
Turn Around Time: 3-4 days
Probe(s): HER-2/neu gene and centromere for chromosome 17 (CEP 17)
CPT Code Information:
88271x2 DNA probe, each
88274 Molecular cytogenetics; 25-99 cells
88365 Interpretation and report
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Oncological (Lymphoma) Anomalies – Paraffin Embedded Tissue - Touch Prep
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To rapidly identify the presence of various chromosomal rearrangements or aneuploidy in interphase cells of tissue sample embedded in paraffin.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Note: When a fresh sterile sample is available, routine cytogenetic analysis is strongly recommended to identify possible additional oncological-associated chromosome abnormalities which would not be identified through FISH analysis.
Paraffin-Embedded Tissue Block - Formalin-fixed, paraffin-embedded cancer tissue block. Send at least 4 slides with tissue sections cut at ~4um thick and put on positively charged slides. If a tissue block is sent, the tissue will be sectioned and slides will be prepared at an additional cost.
Touch Prep - Analysis can be done on a simple touch prep on a positive charge slide can very quickly if possible.
Turn Around Time: 3-5 days
Probe(s): See the List (PDF). Not all probes can be used on paraffin embedded samples. Please contact the laboratory director for specific cases.
CPT Code Information:
88271x2 DNA probe, each (eg FISH)
88275 Interphase in situ hybridization, analyze 100-300 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]
Test: Oncological Urothelial – Urine or Uretheral/Bladder Wash
Fluorescence In Situ Hybridization (FISH)
Purpose of Test: To detect the presence of urothelial carcinoma or transitional cell carcinoma of the bladder by the analysis of cells from urine or urethral/bladder washes for aneuploidy for chromosomes 3, 7, 17, and/or loss of the 9p21 locus.
Required Information: Please provide indications (ICD-9) for testing on our referral form with each specimen. Accurate testing and interpretation may otherwise be compromised.
Urine of Uretheral/Bladder Wash
- Aseptically collect 33 to 50 mls of urine or urethral/bladder wash.
- Transfer to a 50ml centrifuge tube containing Carobwax (provided by our laboratory) or other tightly sealed sterile plastic container.
- Label tube(s) with patient name.
- IF SPECIMEN IS NOT SENT THE SAME DAY, REFRIGERATE OVERNIGHT AND SEND SPECIMEN REFRIGERATED WITHIN 24 HRS, DO NOT FREEZE.
Turn Around Time: 5 days
CPT Code Information:
88271x4 DNA probe
88275 Molecular cytogenetics; 100-300 cells
[Lab Referral Form] [Printable PDF File] [Back to Top]