Statistical Treatment of Fluorescence in Situ Hybridization Validation Data to Generate Normal Reference Ranges Using Excel Functions

Ciolino, Allison; Tang, Mary E.; Bryant, Ron

doi:10.2353/jmoldx.2009.080101

Cited by 22 publications

(18 citation statements)

References 8 publications

(14 reference statements)

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“…The beta inverse formula does not allow a cutoff of zero. 19 Ten specimens, including normal cases and cases with MLL rearrangement confirmed by another method, were then examined in a blinded fashion for detection of different MLL abnormalities in patient samples. Specimens known to have 11q23 rearrangement by cytogenetics were selected as a gold standard to validate the cutoff values.…”

Section: Reference Range (Normal Cutoff)mentioning

confidence: 99%

Validation of Fluorescence In Situ Hybridization Using an Analyte-Specific Reagent for Detection of Abnormalities Involving the Mixed Lineage Leukemia Gene

Saxe¹,

Persons²,

Wolff³

et al. 2012

Archives of Pathology &Amp; Laboratory Medicine

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Objective.-To provide an example of how a FISH assay, using an analyte-specific reagent probe, may be validated in a clinical laboratory.Design.-We describe the approach used by an individual laboratory for validation of a FISH assay for mixed lineage leukemia (MLL) gene.Results.-Specific validation data are provided illustrating how initial assay performance characteristics in a FISH assay for MLL may be established.Conclusions.-Protocols for initial validation of FISH assays may vary between laboratories. However, all laboratories must establish several defined performance specifications prior to implementation of FISH assays for clinical use. We describe an approach used for assessing performance specifications and validation of an analytespecific reagent FISH assay using probes for MLL rearrangement in interphase nuclei.

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Section: Reference Range (Normal Cutoff)mentioning

confidence: 99%

Validation of Fluorescence In Situ Hybridization Using an Analyte-Specific Reagent for Detection of Abnormalities Involving the Mixed Lineage Leukemia Gene

Saxe¹,

Persons²,

Wolff³

et al. 2012

Archives of Pathology &Amp; Laboratory Medicine

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“…The definition of false positive signal pattern was less than 2 red signals for TP53 /CEP17, any yellow fusion signal for t(4;14) IGH / FGFR3 or t(14;16) IGH / MAF , and more than 2 red signals for chromosome 1q CKS1B / CDKN2C . The critical binomial function of the Microsoft Excel spreadsheet was used to determine the 95 % confidence limit of normal cutoff value [ 13 ]. The normal cutoff value for TP53 /CEP17 was 3.4 %, t(4;14) IGH / FGFR3 was 6.8 %, t(14;16) IGH / MAF was 5.6 % and chromosome 1q CKS1B / CDKN2C was 5.7 %.…”

Section: Resultsmentioning

confidence: 99%

Target fluorescence in-situ hybridization (Target FISH) for plasma cell enrichment in myeloma

et al. 2016

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BackgroundCytogenetic abnormalities are important prognostic markers in plasma cell myeloma (PCM) and detection is routinely performed by interphase fluorescence in-situ hybridization (FISH) with a panel of probes after enrichment of the plasma cells in the bone marrow specimen. Cell sorting by immunomagnetic beads and concurrent labeling of the cytoplasmic immunoglobulin are the usual enrichment methods. We present an alternative method of plasma cell enrichment termed Target FISH, which is an automated system that combines the images of May-Grünwald- Giemsa (MGG) staining and FISH study on the same plasma cell for analysis.ResultsOur experience of Target FISH on 40 PCM patients was described. Briefly, plasma cells were MGG stained, image captured, de-stained, FISH probe hybridized and finally relocated for simultaneous analysis of morphology and FISH signal pattern. The FISH probe panel was TP53/CEP17, t(4;14) IGH/FGFR3, t(14;16) IGH/MAF and CKS1B(1q21)/CDKN2C(P18). Gain of 1q21 was the most common abnormality detected in 18 patients (45 %), to be followed by t(4;14) IGH/FGFR3 detected in 11 patients (27.5 %). Of note, 10 patients showed coexistence of both t(4;14) and 1q21 gain. Two patients showed del(17p)/TP53, one in association with t(4;14) and 1q gain while the other was stand alone. None of this patient cohort showed t(14;16) IGH/MAF. Using the critical binomial function, the normal cutoff FISH positive value for del(17p)/TP53 was 3.4 %, t(4;14) IGH/FGFR3 was 6.8 %, t(14;16) IGH/MAF was 5.6 % and +1q21 was 5.7 %.ConclusionsThe equipment cost notwithstanding, when compared with cell sorting, the total reagent cost was around 10 % lower in Target FISH. The total processing time was longer for Target FISH but manual fluorescence microscopy was no longer necessary. The main advantage of Target FISH was the complete certainty that the cytogenetic abnormality was detected in the cells of interest, and hence a more stringent analytical cutoff value might be considered. Optimization of the cell collection and slide preparation process upfront was required to accrue adequate target cells on each slide for analysis. Our experience suggested that Target FISH was applicable as a routine method of plasma cell enrichment in clinical diagnostic laboratories.

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“…Based on FISH analysis of 18 normal pancreatic tissue samples in a preliminary study, cutoff values for the normal range for FISH analysis were calculated using the Excel 2013 (Microsoft Corp., Redmond, WA, USA) statistical function CRITBINOM (n, p, α) with a confidence level of 95% (Table 1). 24 When the percentage of cells containing >2 or <2 FISH signals exceeded the cutoff value, cases were interpreted as positive for polysomy (gain) or monosomy (deletion), respectively.…”

Section: Fluorescence In Situ Hybridizationmentioning

confidence: 99%

The Implication of Cytogenetic Alterations in Pancreatic Ductal Adenocarcinoma and Intraductal Papillary Mucinous Neoplasm Identified by Fluorescence In Situ Hybridization and Their Potential Diagnostic Utility

Lim

Lee

et al. 2020

Gut and Liver

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Background/Aims: We investigated chromosomal aberrations in patients with pancreatic ductal adenocarcinoma (PDAC) and intraductal papillary mucinous neoplasm (IPMN) by fluorescence in situ hybridization (FISH) to identify cytogenetic changes and molecular markers that may be useful for preoperative diagnosis. Methods: Tissue samples from 48 PDAC and 17 IPMN patients were investigated by FISH analysis using probes targeting chromosomes 7q, 17p, 18q, 20q, and 21q and the pericentromeric region of chromosome 18 (CEP18). Results: The PDAC samples harbored 17p deletion (95.8%), 18q deletion (83.3%), CEP18 deletion (81.2%), 20q gain (81.2%), 21q deletion (77.1%), and 7q gain (70.8%). The IPMN samples had 17p deletion (94.1%), CEP18 deletion (94.1%), 21q deletion (70.6%), 18q deletion (58.8%), 20q gain (58.8%), and 7q gain (58.8%). A significant difference in CEP18 gain was identified between the PDAC and IPMN groups (p=0.029). Detection of 17p or 18q deletion had the highest diagnostic accuracy (80.0%) for PDAC. Conclusions: Chromosomal alterations were frequently identified in both PDAC and IPMN with similar patterns. CEP18 gain and 17p and 18q deletions might be involved in the later stages of PDAC tumorigenesis. Chromosome 17p and 18q deletions might be excellent diagnostic markers.

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Statistical Treatment of Fluorescence in Situ Hybridization Validation Data to Generate Normal Reference Ranges Using Excel Functions

Cited by 22 publications

References 8 publications

Validation of Fluorescence In Situ Hybridization Using an Analyte-Specific Reagent for Detection of Abnormalities Involving the Mixed Lineage Leukemia Gene

Validation of Fluorescence In Situ Hybridization Using an Analyte-Specific Reagent for Detection of Abnormalities Involving the Mixed Lineage Leukemia Gene

Target fluorescence in-situ hybridization (Target FISH) for plasma cell enrichment in myeloma

The Implication of Cytogenetic Alterations in Pancreatic Ductal Adenocarcinoma and Intraductal Papillary Mucinous Neoplasm Identified by Fluorescence In Situ Hybridization and Their Potential Diagnostic Utility

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