Fluorescent in situ hybridization (FISH): A useful diagnostic tool for childhood conjunctival melanoma

2021 ◽  
pp. 112067212110307
Author(s):  
Raquel María Moral ◽  
Carlos Monteagudo ◽  
Javier Muriel ◽  
Lucía Moreno ◽  
Ana María Peiró
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Pediatric Population ◽  
Diagnostic Technique ◽  
Histopathological Diagnosis ◽  
Fish Analysis ◽  
Conjunctival Melanoma ◽  
Adequate Treatment ◽  
First Case

Introduction: Conjunctival melanoma is extremely rare in children and has low rates of resolution. Definitive histopathological diagnosis based exclusively on microscopic findings is sometimes difficult. Thus, early diagnosis and adequate treatment are essential to improve clinical outcomes. Clinical case: We present the first case in which the fluorescent in situ hybridization (FISH) diagnostic technique was applied to a 10-year-old boy initially suspected of having amelanotic nevi in his right eye. Based on the 65% of tumor cells with 11q13 (CCND1) copy number gain and 33% with 6p25 (RREB1) gain as measured by the FISH analysis, and on supporting histopathological findings, the diagnosis of conjunctival melanoma could be made. Following a larger re-excision, adjuvant therapy with Mitomycin C (MMC), cryotherapy and an amniotic membrane graft, the patient has remained disease-free during 9 years of long-term follow-up. Case discussion: Every ophthalmologist should remember to consider and not forget the possibility of using FISH analyses during the differential diagnosis of any suspicious conjunctival lesions. Genetic techniques, such as FISH, have led to great advances in the classification of ambiguous lesions. Evidence-based guidelines for diagnosing conjunctival melanoma in the pediatric population are needed to determine the most appropriate strategy for this age group.

Detection of t(4;14)(p16.3;q32) Chromosomal Translocation in Multiple Myeloma by Double-Color Fluorescent In Situ Hybridization

Blood ◽  
1999 ◽  
Vol 94 (2) ◽  
pp. 724-732 ◽  
Author(s):  
Palma Finelli ◽  
Sonia Fabris ◽  
Savina Zagano ◽  
Luca Baldini ◽  
Daniela Intini ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
In Situ Hybridization ◽  
Cell Lines ◽  
Fluorescent In Situ Hybridization ◽  
Chromosomal Translocation ◽  
Fish Analysis ◽  
Interphase Nuclei ◽  
Igh Locus ◽  
Normal Controls

Abstract Chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus at chromosome 14q32 represent a common mechanism of oncogene activation in lymphoid malignancies. In multiple myeloma (MM), variable chromosome partners have been identified by conventional cytogenetics, including the 11q13, 8q24, 18q21, and 6p21 loci. We and others have recently reported a novel, karyotypically undetectable chromosomal translocation t(4;14)(p16.3;q32) in MM-derived cell lines, as well as in primary tumors. The 4p16.3 breakpoints are relatively scattered and located less than 100 kb centromeric of the fibroblast growth factor receptor 3 (FGFR3) gene or within the recently identified WHSC1 gene, both of which are apparently deregulated by the translocation. To assess the frequency of the t(4;14)(p16.3;q32) translocation in MM, we performed a double-color fluorescent in situ hybridization (FISH) analysis of interphase nuclei with differently labeled probes specific for the IGH locus (a pool of plasmid clones specific for the IGH constant regions) or 4p16.3 (yeast artificial chromosome (YAC) 764-H1 spanning the region involved in breakpoints). Thirty MM patients, the MM-derived cell lines KMS-11 and OPM2, and six normal controls were examined. The identification of a t(4;14) translocation, evaluated as the presence of a der(14) chromosome, was based on the colocalization of signals specific for the two probes; a cutoff value of 15% (mean + 3 standard deviation [SD]) derived from the interphase FISH of the normal controls (range, 5% to 11%; mean ± SD, 8.16 ± 2.2) was used for the quantification analysis. In interphase FISH, five patients (one in clinical stage I, two in stage II, one in stage III, and a plasma cell leukemia) were found to be positive (≈15%). FISH metaphases with split or colocalized signals were detected in only two of the translocated cases and confirmed the pattern found in the interphase nuclei. Furthermore, in three of the five cases with the translocation, FISH analysis with the IGH joining probe (JH) showed the presence of the reciprocal product of the translocation [der(4) chromosome]. Overall, our study indicates that the t(4;14)(p16.3;q32) chromosomal translocation is a recurrent event in MM tumors and may contribute towards the detection of this lesion and our understanding of its pathogenetic and clinical implications in MM.

Detection of t(4;14)(p16.3;q32) Chromosomal Translocation in Multiple Myeloma by Double-Color Fluorescent In Situ Hybridization

Blood ◽  
1999 ◽  
Vol 94 (2) ◽  
pp. 724-732 ◽  
Author(s):  
Palma Finelli ◽  
Sonia Fabris ◽  
Savina Zagano ◽  
Luca Baldini ◽  
Daniela Intini ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
In Situ Hybridization ◽  
Cell Lines ◽  
Fluorescent In Situ Hybridization ◽  
Chromosomal Translocation ◽  
Fish Analysis ◽  
Interphase Nuclei ◽  
Igh Locus ◽  
Normal Controls

Chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus at chromosome 14q32 represent a common mechanism of oncogene activation in lymphoid malignancies. In multiple myeloma (MM), variable chromosome partners have been identified by conventional cytogenetics, including the 11q13, 8q24, 18q21, and 6p21 loci. We and others have recently reported a novel, karyotypically undetectable chromosomal translocation t(4;14)(p16.3;q32) in MM-derived cell lines, as well as in primary tumors. The 4p16.3 breakpoints are relatively scattered and located less than 100 kb centromeric of the fibroblast growth factor receptor 3 (FGFR3) gene or within the recently identified WHSC1 gene, both of which are apparently deregulated by the translocation. To assess the frequency of the t(4;14)(p16.3;q32) translocation in MM, we performed a double-color fluorescent in situ hybridization (FISH) analysis of interphase nuclei with differently labeled probes specific for the IGH locus (a pool of plasmid clones specific for the IGH constant regions) or 4p16.3 (yeast artificial chromosome (YAC) 764-H1 spanning the region involved in breakpoints). Thirty MM patients, the MM-derived cell lines KMS-11 and OPM2, and six normal controls were examined. The identification of a t(4;14) translocation, evaluated as the presence of a der(14) chromosome, was based on the colocalization of signals specific for the two probes; a cutoff value of 15% (mean + 3 standard deviation [SD]) derived from the interphase FISH of the normal controls (range, 5% to 11%; mean ± SD, 8.16 ± 2.2) was used for the quantification analysis. In interphase FISH, five patients (one in clinical stage I, two in stage II, one in stage III, and a plasma cell leukemia) were found to be positive (≈15%). FISH metaphases with split or colocalized signals were detected in only two of the translocated cases and confirmed the pattern found in the interphase nuclei. Furthermore, in three of the five cases with the translocation, FISH analysis with the IGH joining probe (JH) showed the presence of the reciprocal product of the translocation [der(4) chromosome]. Overall, our study indicates that the t(4;14)(p16.3;q32) chromosomal translocation is a recurrent event in MM tumors and may contribute towards the detection of this lesion and our understanding of its pathogenetic and clinical implications in MM.

scholarly journals Use of FISH technique in the diagnosis of chromosomal syndromes

1999 ◽  
Vol 5 (6) ◽  
pp. 1218-1224
Author(s):  
M. A. Iqbal ◽  
C. Ulmer ◽  
N. Sakati
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Chromosome Analysis ◽  
Laboratory Diagnosis ◽  
Fish Analysis ◽  
Prader Willi Syndrome ◽  
Microdeletion Syndrome ◽  
Microdeletion Syndromes ◽  
Di George Syndrome

Major chromosome abnormalities are present in 0.65% of all neonates. Fluorescent in situ hybridization [FISH]is useful in diagnosing microdeletion syndromes that would otherwise be difficult to diagnose using standard cytogenetics. In this study, we used FISH analysis in the laboratory diagnosis of 4 patients with Prader-Willi Syndrome [del[15][q11.2q12], 4 patients with Di George syndrome [del[22][q11.2q11.23] and 4 patients with Williams syndrome [del[7][q11.23q11.23]. High-resolution chromosome analysis in all these patients was either normal or inconclusive but all the syndromes were confirmed using FISH. We recommend cytogenetic analysis should always be supplemented with FISH to diagnose all cases suspected of a microdeletion syndrome

scholarly journals Automated fluorescent in situ hybridization (FISH) analysis of t(9;22)(q34;q11) in interphase nuclei

2006 ◽  
Vol 69A (6) ◽  
pp. 506-514 ◽  
Author(s):  
Béla Kajtár ◽  
Gábor Méhes ◽  
Thomas Lörch ◽  
Linda Deák ◽  
Marika Kneifné ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Fish Analysis ◽  
Interphase Nuclei

scholarly journals Molecular Delineation of 13q Deletion Boundaries in 20 Patients With Myeloid Malignancies

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 231-237 ◽  
Author(s):  
R. La Starza ◽  
I. Wlodarska ◽  
A. Aventin ◽  
D. Falzetti ◽  
B. Crescenzi ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Chromosome Morphology ◽  
Lymphocytic Leukemia ◽  
Fish Analysis ◽  
Genomic Segment ◽  
Myeloid Malignancies ◽  
Homozygous Deletions ◽  
13Q Deletion

Fluorescent in situ hybridization (FISH) analysis with a panel of DNA probes for 13q13.1-q14.3 was performed on 20 cases of myeloid malignancies, of which 17 showed a del(13)(q) and three had translocations affecting 13q. By chromosome morphology, deletions consistently involved bands q14 and q21. In addition to confirming the chromosome data, FISH allowed us to delineate a commonly deleted region that was flanked by YAC 833A2 and YAC 854D4. Three cases with 13q translocations unexpectedly showed accompanying cryptic microdeletions of 13q, and in one case the commonly deleted region could be narrowed to a genomic segment, which includes YAC 937C7, RB1, and YAC 745E3. Homozygous deletions were not detected. This region overlaps with the smallest deleted region of 13q14 in chronic lymphocytic leukemia.

367 FLUORESCENT IN SITU HYBRIDIZATION FOR SEX DETERMINATION AT PROGRESSIVE STAGES OF FERTILIZATION AND DEVELOPMENT IN MICE

2007 ◽  
Vol 19 (1) ◽  
pp. 299
Author(s):  
J. J. Whyte ◽  
E. D. Fountain ◽  
C. S. Rosenfeld
Keyword(s):  
In Situ Hybridization ◽  
Sex Determination ◽  
Fluorescent In Situ Hybridization ◽  
Fish Analysis ◽  
Epididymal Sperm ◽  
Pregnant Females ◽  
Y Chromosomes ◽  
Nih Swiss ◽  
Post Implantation

In mice, relative numbers of male and female pups in a litter can not only vary but may change over the course of pregnancy in response to a range of environmental and physiological factors. The ability to determine the temporal nature of changes in gender ratios in mice has extrapolative value for agriculture, endangered species, and human applications. We have developed a robust and accurate fluorescent in situ hybridization (FISH) suite for visualization of X and Y chromosomes in sperm, pre- and post-implantation conceptuses, and dead/partially cannibalized pups with minimal modification for various samples. Epididymal sperm was collected from NIH Swiss males (n = 5; 15 weeks old) and prepared for FISH by using a modified microwave decondensation–denaturation technique. Pre-implantation embryos (0.5 dpc) collected from pregnant females (n = 9; 20 weeks old) were cultured to the morula stage to permit the development of several blastomeres for FISH analysis. They were then acid-treated to remove the zona pellucida and allowed to adhere to a single slide. Two pre-hybridization procedures, microwave decondensation–denaturation and pepsin digestion (100 �g mL-1 in 0.01 N HCl), were compared for efficacy. Cranial tissue homogenates in phosphate-buffered saline from post-implantation conceptuses (8.5 dpc) collected from pregnant females (n = 8; 20 weeks old) were fixed to pre-etched slides. To determine whether this procedure could be used on partially cannibalized or stillborn pups, 2 such litters with n = 7 pups from each were retrieved from NIH Swiss females. As the dams had cannibalized the caudal half of the pups, the sex of these could not be determined based on anogenital distance. A small sample of tissue from the remaining cranial region from each was prepared as above for XY FISH analysis, allowing genotypic sexing in pups that would otherwise be of undetermined gender. For each stage of development, slides were hybridized overnight with identical commercially available fluorescence-labeled DNA probes for the X (FITC) and Y (Cy3) chromosomes. Epididymal sperm ratios met the expected value of 0.5. Pepsin treatment of pre implantation embryos provided superior results over microwave decondensation with distinct fluorescence of X and Y chromosomes in morula. Mean sex ratios of 0.5 were determined for both 0.5 and 8.5 dpc embryos (n = 11 � 1 and 10 � 1 embryos per female, respectively). Sex determination in post-implantation embryos was confirmed by using 2 separate PCR analyses. The procedure was also successful in recovered postmortem pups and yielded 8 females and 6 males between the 2 litters. This technique may thus be used to detect selective post-natal mortality or cannibalism of one gender over the other. FISH analysis of sperm and multiple pre- or post-implantational embryos or postmortem pups provides a cost-effective and more accurate alternative to PCR-based sex determination. This technique may have potential for use in studies where offspring sex ratios have been influenced by external factors, but wherein the mechanism and stage at which the skewing occurs is uncertain.

scholarly journals Determination of the metabolically active fraction of benthic foraminifera by means of Fluorescent in situ Hybridization (FISH)

2010 ◽  
Vol 7 (5) ◽  
pp. 7475-7503 ◽  
Author(s):  
C. Borrelli ◽  
A. Sabbatini ◽  
G. M. Luna ◽  
C. Morigi ◽  
R. Danovaro ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Rose Bengal ◽  
Benthic Foraminifera ◽  
Fluorescent In Situ Hybridization ◽  
Physiological Status ◽  
Hybridization Technique ◽  
Fish Analysis ◽  
Type I ◽  
Active Metabolism

Abstract. Benthic foraminifera are an important component of the marine living biota, but protocols for investigating their viability and metabolism are still extremely limited. Classical studies on benthic foraminifera have been based on direct counting under light microscopy. Typically these organisms are stained with Rose Bengal, which binds proteins and other macromolecules, but this approach does not allow discriminating between viable and recently dead organisms. The fluorescent in situ hybridization technique (FISH) represents a potentially useful approach identifying living cells with active metabolism cells. In this work, we tested for the first time the suitability of the FISH technique based on fluorescent probes targeting the 18S rRNA, to detect these live benthic protists. The protocol was applied on the genus Ammonia, on the Miliolidae group and an attempt was made also with agglutinated species (i.e., Leptohalysis scottii and Eggerella scabra). In addition microscopic analysis of the cytoplasm colour, presence of pigments and, sometimes, those of pseudopodial activity where conducted. The results of the present study indicate that FISH targeted only live and metabolically active foraminifera. These results allowed to identify as "live", cells improperly classified as "dead" by means of the classical technique (Type I error) and vice versa to identify as dead the foraminifera without rRNA, but stained using Rose Bengal (Type II error). In addition, the comparative FISH analysis of starved and actively growing cells demonstrated that individuals with active metabolism were stained more intensively than starved cells. This finding supports the hypothesis that the physiological status of cells can be directly related with the intensity of the fluorescent signal emitted by the fluorescent probe. We conclude that the use of molecular approaches could represent a key tool for acquiring crucial information on living foraminifera specimens and for investigating their ecological role in marine sediments.

scholarly journals Molecular Delineation of 13q Deletion Boundaries in 20 Patients With Myeloid Malignancies

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 231-237 ◽  
Author(s):  
R. La Starza ◽  
I. Wlodarska ◽  
A. Aventin ◽  
D. Falzetti ◽  
B. Crescenzi ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Chromosome Morphology ◽  
Lymphocytic Leukemia ◽  
Fish Analysis ◽  
Genomic Segment ◽  
Myeloid Malignancies ◽  
Homozygous Deletions ◽  
13Q Deletion

Abstract Fluorescent in situ hybridization (FISH) analysis with a panel of DNA probes for 13q13.1-q14.3 was performed on 20 cases of myeloid malignancies, of which 17 showed a del(13)(q) and three had translocations affecting 13q. By chromosome morphology, deletions consistently involved bands q14 and q21. In addition to confirming the chromosome data, FISH allowed us to delineate a commonly deleted region that was flanked by YAC 833A2 and YAC 854D4. Three cases with 13q translocations unexpectedly showed accompanying cryptic microdeletions of 13q, and in one case the commonly deleted region could be narrowed to a genomic segment, which includes YAC 937C7, RB1, and YAC 745E3. Homozygous deletions were not detected. This region overlaps with the smallest deleted region of 13q14 in chronic lymphocytic leukemia.

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