FISH Automation 소개 2007.09.06 진단검사의학과 강지연

Fluorescence in situ Hybridization 검사하고자 하는 표본에 찾고자 하는 probe를 반응시킨 후 특정부위의 유전자나 염색체의 결실, 추가, 증폭, 재배열을 형광현미경으로 관찰하는 방법 Specimen treatment Harvest / Slide preparation 및 경검 Front End FISH 1st day 2nd day (암실에서 진행) Pretreatment Dehydration 암실에서 진행 DNA-Probe 첨가 Denaturation Hybridization Cover slip 제거 Washing (Wash1,Wash2,Wash3) Counter stain: DAPI 부착

Back End FISH Analysis by fluorescence microscopy dual color, dual fusion break apart

FISH Automation - Overview FISH Automation - Overview Front End Automation Automates slide/specimen pretreatment and assay Can be used for FISH, G-Banding, IHC & others Two Configurations (1) VP2000 and ThermoBrite™ (2) Xmatrx Back End Automation Automated slide scanning & enumeration MetaSystems Other Applications in process We sell and support FISH however the instruments can be used for other slide-based techniques. VP2000 and ThermoBrite can be used separately and do not have to be used in succession

Automation Choices Front-End Back-End VP2000 ThermoBrite metasystems Denaturation & Hybridization ThermoBrite Automation Choices Slide Pre-treatment & Washing VP2000 Slide Scanning & Enumeration Slide preparation Pre-treatment, Washing, Denaturation, Hybridization & Final Cover slip Xmatrx* metasystems * In Development

Front End FISH VP2000 Update Automates FISH Assay Steps: FISH Pretreatment Specimen Staining Up to 50 slides simultaneously 3 heated reagent basins Ventilated closed system General purpose system accommodates most slide-based assays: histology, G-banding, etc Increase FISH throughput Increase consistency of results Decrease labor costs

Front-End FISH ThermoBrite™ Front-End FISH ThermoBrite™ Slide Hybridization & Denaturation System Eliminates separate denaturation 40 definable programs – LCD screen 12 slide capacity – guides hold slides Humidity cards Superior temperature uniformity Rapid temperature ramp

Front-End FISH XMATRX Automation of ALL FISH assay steps Throughput Front-End FISH XMATRX Automation of ALL FISH assay steps Pre-treatment, Probe, Coverslip application, Washing, Denaturation, Hybridization, Cover Slip Removal, Wash, Counter stain & Final Cover slip Throughput Up to 40 slides per run Up to 80 assays per run Random access, up to 8 different assays in one run Continuous access, slides can be added or removed during the run Full reagent management capability Applies, removes & reapplies cover slips PC with easy to use software

Front-End FISH A Fully Automated FISH System

Back-End FISH Carl Zeiss Microscope A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Back-End FISH Carl Zeiss Microscope Axio Imager.M1/Z1 Motorized stand Automatic Focussing Maximum variability Control by PC or touch screen The best fluorescence with higher contrast 1

Metafer - Slide Scanning System A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Back-End FISH Metafer - Slide Scanning System 1

real-time image analysis and microscope control A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer Concept CCD Camera Metafer PC Win 9x 2000 / NT Motorized stage (x,y) real-time image analysis and microscope control Motorized microscope (z) 1

A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2 A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer Concept • Microscope slide is scanned field by field at lowest possible magnification without gaps • Objects of interest are automatically identified ; gallery images, object features and positions are recorded 1

Metafer Advantages of Metafer High speed Slide Scanning (70mm/s) A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer Advantages of Metafer High speed Slide Scanning (70mm/s) High quality image capture during the scan High throughput and capacity multi-fluorochrome capability from UV to IR - very few objects available for analysis - statistical analysis of many objects 1

Metafer Applications MSearch : metaphase finder A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer Applications MSearch : metaphase finder - Micronucleus analysis - Automatic analysis of dicentrics MetaCyte : automated FISH analysis / spot counting - spot counting and translocation analysis - amplification analysis in tissue section - Comet scan - High content screening RCDetect : rare cell detection - minimum residual disease detection 1

Metafer - MSearch : metaphase finder A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer - MSearch : metaphase finder 1

Metafer – Msearch : metaphase finder A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer – Msearch : metaphase finder Fast metaphase finding in transmitted and fluorescent light 1

Metafer - MetaCyte : automated FISH analysis A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer - MetaCyte : automated FISH analysis 1

A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2 A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Metafer – Slide Feeder 1

Isis - Fluoresc. Imaging System A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Back-End FISH Isis - Fluoresc. Imaging System 1

Isis Fluoresc. Imaging System – features and benefits Fast and easy capture Powerful image enhancement Various color display mode, measurement function Database manegement as Ikaros Additional Isis modules Color karyotyping module mFISH and mBAND module CGH(Comparative Genome Hybridization) module Telomere measurement module

Isis Fluoresc. Imaging System – Capture A Team Meeting Marketintroduction LSM 5 PASCAL, LSM 510 SW Rel. 2.5, ConfoCor 2 Stand Alone Isis Fluoresc. Imaging System – Capture Focus adjustment live on screen Automatic exposure control If with Carl Zeiss Axio Imager.M1 or Z1 microscope - automatic filter change - focus stack capture 1

Summary Improve workflow efficiency Increase throughput; enable labs to do more with the same resources Reduce “hands-on” time; free up resources and simplifies workflow Increase assay quality and reproducibility Multiple solutions; different levels of automation Applications to our laboratory(metasystems) Rapid finding of metaphase & FISH interphase Good assay quality of FISH analysis Pannel FISH Urovision