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A method of analyzing chromosomal translocations and a system therefore

A technique for chromosomal translocation and sample analysis, applied in the field of systems for analyzing chromosomal translocation

Active Publication Date: 2014-01-22
VENTANA MEDICAL SYST INC
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, none of these systems have been applied to address the problems associated with false positive results in split probe systems

Method used

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  • A method of analyzing chromosomal translocations and a system therefore
  • A method of analyzing chromosomal translocations and a system therefore
  • A method of analyzing chromosomal translocations and a system therefore

Examples

Experimental program
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Embodiment 1

[0139] Materials and Method

[0140] ALK triple probe design

[0141] The dissociation in situ hybridization (ba-ISH) assay is designed to assess the rearrangement of the ALK gene locus (ALK-EML4 fusion). Generate 3 kinds of probes to hybridize with the adjacent centromeric region (770kb) and telomere region (683kb) of ALK gene and ALK gene region (728Kb) ( figure 1 ). The ALK gene probe is labeled with NP hapten, the 5'ALK probe is labeled with DIG hapten, and the 3'ALK probe is labeled with DNP hapten.

[0142] Automated brightfield separation in situ hybridization protocol

[0143] All optimization and performance evaluation of the ALK gene ba-ISH assay for brightfield in situ hybridization are used XT automated slide processing system (Ventana Medical Systems, Inc., Tucson, AZ). Create ba-ISH instrument software, so that all steps from baking to counterstaining can be performed without interruption. Bake the slides on the instrument at 65°C for 20 minutes to melt the paraffin,...

Embodiment 2

[0145] The following example describes the method used to analyze ALK rearrangements using a three-color separation probe system. The slides were baked on the instrument at 65°C for 20 minutes to melt the paraffin, followed by the EZ preparation (Ventana Medical Systems, Inc.) deparaffinization step initiated by the liquid coverslip (Ventana Medical Systems, Inc.). The DNA target was recovered by a combination of heat treatment with a citrate buffer-based target recovery solution CC2 (Ventana Medical Systems, Inc.) and tissue digestion with ISH protease 3 (Ventana Medical Systems, Inc.). Due to the different tissue fixation and processing conditions of clinical samples, the appropriate protease digestion time is determined for each tissue sample. A mixture of 3 types of ALK probes (DNP-labeled 5'ALK probe, fluorescein-labeled internal ALK probe, and DIG-labeled 3'ALK probe, each 12μg / ml) and Ventana hybridization buffer The fish DNA is prepared together. The probe and target ...

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Abstract

The present disclosure relates to systems and methods for analyzing chromosomal translocations, and in particular to analysis of chromosomal translocation by in situ hybridization. The method employs a 5 '-probe, a 3 '-probe and a probe binding adjacent to, optionally overlapping, the breakpoint. The probes are differently labelled. In particular, the use in the detection of translocations involving the ALK gene are described.

Description

Invention field [0001] The present disclosure relates to systems and methods for analyzing chromosomal translocations, and in particular to the analysis of chromosomal translocations by in situ hybridization. Background of the invention [0002] The diagnosis, prognosis, and treatment of diseases based on the interpretation of tissue or cell samples taken from diseased organisms have expanded dramatically in the past few years. In addition to traditional histological staining techniques and immunohistochemical assays, in situ techniques such as in situ hybridization and in situ polymerase chain reaction are now used to help diagnose human disease states. As such, there are a variety of technologies that can assess not only cell morphology but also the presence of specific macromolecules in cells and tissues. [0003] Molecular cytogenetic techniques such as chromophore in situ hybridization (CISH) combine visual assessment of chromosomes (karyotype analysis) with molecular techniq...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6841C12Q2565/102C12Q2539/115C12Q1/6827C12Q2525/185C12Q2543/101
Inventor M.法雷尔T.M.格罗甘H.尼塔张文军
Owner VENTANA MEDICAL SYST INC