CytoVision offers a multi-application platform to support investigations from routine cytogenetic analysis of blood and bone marrow, through to karyotyping, FISH and MFISH in stem cell research.

Case studies Case studies

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Automating scanning and capture followed by on-screen analysis — measuring the impact on laboratory services

West of Scotland Regional Cytogenetics Service
“Implementation of automated scanning and on-screen analysis has led to a significant improvement in reporting times, quality and success rate and has also decreased the poor quality rate for both Routine and Urgent blood samples”
Administrator, Yorkhill Hospital, Glasgow
  • Purpose: find, rank and capture metaphase cells in post natal blood samples
  • Criteria: scan up to maximum 300 metaphases per slide, capture 50 best cells
  • Typical throughput: 2800 samples per year

West of Scotland Regional Cytogenetics Service

Data courtesy of L. Monkman, J. Colgan, L. Crawford, M. Campbell, G.Lowther
Cytogenetics Laboratory, Duncan Guthrie Institute of Medical Genetics
Yorkhill Hospital, Glasgow, UK

Process

GSL-120 and CytoVision unattended overnight runs

  • 40 slides scanned and cells captured
    • 7 min and 10X scan per slide
    • 15-20 min. to capture 50 metaphases per slide

On-screen analysis next morning

  • Case checked for number and adequate quality for referral reason
  • Satisfactory cases pass to scientist for analysis
    • Karyotype cells
    • Examine chromosome pairs
    • Manipulate chromosome size and enhance contrast of banding
    • Fully analyze each homolog twice (follows UK guidelines)
    • Records completed analysis

Results of implementation

Increase in quality

  • Statistically significant increase in quality of Urgent and Routine samples after automation
Quality evaluated using G-banding Evaluation Score table from UK Professional Guidelines

Decrease in reporting times

  • Increase in % of cases reported within guideline times
UK Professional Guidelines for post-natal samples: 95% of Urgent samples to be reported within 10 days; 95% of Routine samples to be reported within 28 days.
Pre-automation
Post-automation
Routine
28.4%
99.6%
Urgent
69.7%
88.1%
% samples reported within guideline reporting times

Increase in success rate, decrease in poor quality rate

Urgent cases
Routine cases

Reduced cases backlog since installation

CytoVision contrast enhancement

“Slide-making and banding workload of technical staff has been reduced by around 50%”
Poster, 7th European Cytogeneticists Conference 2009, Stockholm
Before CytoVision enhancement
After CytoVision enhancement

Conclusions

Implementation of automated scanning and on-screen analysis has led to a significant improvement in reporting times, quality and success rate and has also decreased the poor quality rate for both Routine and Urgent blood samples.

Evaluation of efficiency gains using automated scanning and capture technology

Sonora Quest Laboratories
“The autoscanner, or “Auto” as it is affectionately called, has become a valuable member of our laboratory family.”

Evaluation of efficiency gains using automated scanning and capture technology

Data courtesy of Leah Weems ,BS,CLSp(CG) Angela Fylak,BA
Sonora Quest Laboratories, Tempe, Arizona, USA 

Process: Six Sigma productivity project

  • Collected time requirements for completion of 220 cases
    • Documented scanning, capture and karyotyping of neoplastic cases
    • Compared to time from 152 cases run using CytoVision/GSL
      • No capture time (done by GSL)
      • Combine analysis and karyotyping times - no longer done by different people
  • Each technologist provided with own review station for on-screen analysis and a microscope to review metaphases by conventional microscopy

Results of implementation

  • Reduced time for case completion by 31.06 mins (20.45%)
  • Decreased failure rate to 1.5%
    • Human inattention removed from the equation
    • Locate metaphases on slides a technologist may assume to be empty
Process
Time for completion
Before GSL-120

Scan and analyze 20 cells: 126 min.

Capture min. 5 pictures: 12.8 min.

Karyotype full case (~2-5 karyotypes per case):
13.1 min.

151.9 min. per case
(n=152)
After GSL-120

Scan, analyze 20 cells capture and cut.

120.8 min. per case
(n=220)

Conclusions

  • Technicians employed to ‘capture/cut’ free for other duties
  • QA benchmarking - abnormality rate within 1% of pre-GSL-120 implementation
  • Non-quantifiable time saving
    • If technologist forgets to write down metaphase co-ordinates – simply click back on-screen to recall
    • To find a picture where 2qs are not crossed – simply flip through 60 captured cells to find the best picture

Implementation into a diagnostics lab

Medical Genetics Laboratory Service, Liverpool Women’s NHS Foundation Unit
  • Purpose:
    • Karyotyping (G-banded analysis)
    • QF-PCR-aneuploidy screen (with karyotype)
    • FISH - micro-deletion studies and chromosome characterization
    • MLPA (sub-telomere and centromere) and aCGH
  • Typical throughput:
    • Amniotic fluid (n-1086), chorion (n=943), products of conception (n=228), bloods (n=2407)
Overall sample numbers

Implementation into a diagnostics lab

Data courtesy of Angela Douglas, Director Medical Genetics Laboratory Service
Liverpool Women’s NHS Foundation Unit, Cheshire and Merseyside, UK

Challenges

  • Overall sample numbers increasing, staff numbers decreasing
  • Increasing and ongoing salaries a limiting cost factor
  • NPAAC guidelines: 250 – 350 cases / scientist / yr, or 6 – 7 cases / week

Decision made — invest in automation

  • Installed CytoVision with GSL-120 Sept 2008
  • Monitored performance parameters pre- and post-installation
    • Analysis time per case
    • Overall reporting time
    • Weekly case load per scientist
    • Paid overtime hours

Results of implementation

  • Analysis time reduced by 35 min (44%) per case
    • GSL-120/CytoVision included karyotyping of all 5 cells analyzed
NPAAC guidelines for routine blood analysis: 5 cells analyzed band-for-band (550bphs), 10 cells counted, 2 representative cells captured and karyotyped.
  • Reporting times reduced, sample throughput increased
  • Overtime decreased, cases per analyst increased

Conclusions

  • Easy to use after minimal training
  • Reporting efficiencies
    • Increase sample referrals with the same staff level
    • Increase the number of samples reported/week
    • Reduce reporting times
  • Analysis efficiencies
    • Permanent record of cells/cases
    • Review large number of cells quickly
    • England finder cell co-ordinates facilitated manual cell finding

Automation in Cytogenomics

Washington University in St.Louis, School of Medicine
  • Purpose: full service laboratory
    • pre-natal
    • pediatric (constitutional)
    • chromosomal microarray
    • cancer and molecular cytogenetics
  • Approx. 30 technologists and staff
  • Challenge: to reduce workload

Automation in Cytogenomics

Data courtesy of Shashikant Kulkarni M.S (Medicine)., PhD., FACMG
Director of Cytogenomics and Molecular Pathology
Washington University in St.Louis, School of Medicine

Strategy to reduce workload – increase automation

  • Moved from CytoVision with few capture stations to one station per technologist, 2005
  • Introduced automated harvesting systems, 2006
  • Introduced GSL-120 system, 2008

Implementation

  • Implemented for GTG banded slide scanning
    • Peripheral blood and bone marrows
  • All workstations networked
  • CITRIX review stations used for
    • Karyotyping scanned images
    • Teaching fellows and new technologist

Results of implementation in practice

  • Substantial automation in workflow
    • CITRIX, paperless process, remote access
    • Possible to consult with colleagues outside of institute
  • Scanning times reduced ~35%
    • Replaced time-consuming manual scanning – especially for low mitotic cases
  • Analysis times reduced ~15%
    • Now able to select best quality cells from a gallery of metaphases

Conclusions

  • Reduction in turn-around-time ~20%
  • On-screen analysis reduces equipment costs
    • 2010 expansion plans do not include microscope capture stations
    • More CITRIX- based expansion helpful and cost-effective when space limited
  • Quality and efficiency of training significantly improved
  • Efficient reviewing of karyograms by inexperienced trainees

Business plan for new way of working

Medical Genetics Laboratory Service, Liverpool Women’s NHS Foundation Unit
  • Purpose: provide a comprehensive service for pre-natal, post-natal and oncology
    • Conventional cytogenetics
    • FISH
    • aCGH
  • Typical throughput: over 11000 samples per year
    • Incl. amniotic fluid, blood, CVS, marrow, solid tissue, solid tumor, microarrays

Business plan for new way of working

Data courtesy of Angela Douglas, Director Medical Genetics Laboratory Service
Liverpool Women’s NHS Foundation Unit, Cheshire and Merseyside, UK

Objectives of business plan (set Nov 2006)

  • Meet efficiency savings
  • Reduce costs - potential revenue saving through natural wastage of established staff posts, reducing reference costs
  • Reduce turnaround times for reporting
  • Decrease patient waiting time
  • Potential to increase capacity to fit increasing testing repertoire
  • Improve accuracy and quality of analysis
  • Electronic documentation, audit trail

Results of implementation

GSL-120 with 10 CytoVision review stations installed May 2008*
Enlarge
Since February 2009*, 90% of Routine blood cases reported within 28 days – UK BPG Standard
Enlarge
Blood weekly figures

Conclusions

Measurable outcome

  • High sample throughput
  • Reduced laboratory test time
  • Decreased patient waiting time
  • Cost effective operation
  • Increased efficiency
  • Electronic documentation of analysis results
  • Built into Pandemic Flu plan

Non-tangible outcome

  • Improved staff morale
  • Improved working lives / culture
  • Improved quality of service
  • Satisfied users

PublicationsPublications

Molecular Cytogenetics of the California Condor: Evolutionary and Conservation Implications

Cytogenet Genome Res. 2009 Dec 29. [Epub ahead of print] DOI: 10.1159/000272458
Modi WS, Romanov M, Green ED, Ryder O.
Evolutionary cytogenetic comparisons involved 5 species of birds (California condor, chicken, zebra finch, collared flycatcher and black stork) belonging to divergent taxonomic orders. Seventy-four clones from a condor BAC library containing 80 genes were mapped to condor chromosomes using FISH, and 15 clones containing 16 genes were mapped to the stork Z chromosome. Maps for chicken and finch were derived from genome sequence databases, and that for flycatcher from the published literature. Gene content and gene order were highly conserved when individual condor, chicken, and zebra finch autosomes were compared, confirming that these species largely retain karyotypes close to the ancestral condition for neognathous birds. However, several differences were noted: zebra finch chromosomes 1 and 1A are homologous to condor and chicken chromosomes 1, the CHUNK1 gene appears to have transposed on condor chromosome 1, condor chromosomes 4 and 9 and zebra finch chromosomes 4 and 4A are homologous to chicken chromosome arms 4q and 4p, and novel inversions on chromosomes 4, 12 and 13 were found. Condor and stork Z chromosome gene orders are collinear and differentiated by a series of inversions/transpositions when compared to chicken, zebra finch, or flycatcher;

Cribriform-Morular Variant of Papillary Thyroid Carcinoma: Molecular Characterization of a Case With Neuroendocrine Differentiation and Aggressive Behavior

Am J Clin Pathol, Jan 2009; 131: 134 - 142
José Cameselle-Teijeiro, Lia P. Menasce, Beng K. Yap, Rovel J. Colaco, Patricia Castro, Ricardo Celestino, Clara Ruíz-Ponte, Paula Soares, and Manuel Sobrinho-Simões.
We describe an especially aggressive case of cribriform-morular variant (C-MV) of papillary thyroid carcinoma (PTC) in a 42-year-old man with familial adenomatous polyposis who died with lung and brain metastases 17 months after thyroidectomy. The angioinvasive neoplasm combined a mixture of trabecular, solid, cribriform, and follicular patterns of growth with CD10+ morules. Follicles were devoid of colloid, and the nuclear features typical of PTC were present in some areas and missing in others. Tumor cells were positive for thyroid transcription factor-1 and, in 40% of the tumoral mass, also were positive for chromogranin and synaptophysin and were negative for thyroglobulin and calcitonin. Strong nuclear staining for beta-catenin was found in all tumor cells, as was positivity for p53 and cyclin D1. In addition to the germline heterozygous APC Ex 2-3 duplication mutation, a somatic homozygous silent p. Thr1493Thr gene variant was found in the neoplastic cells along with RET/PTC rearrangement. This tumor represents the first case of C-MV of PTC showing neuroendocrine differentiation.

MET increased gene copy number and primary resistance to gefitinib therapy in non-small-cell lung cancer patients

Annals of Oncology 2009 20(2):298-304
F. Cappuzzo 1,*, P. A. Jänne2, M. Skokan3, G. Finocchiaro1, E. Rossi4, C. Ligorio1, P. A. Zucali1, L. Terracciano5, L. Toschi2, M. Roncalli6, A. Destro1, M. Incarbone1, M. Alloisio1, A. Santoro1 and M. Varella-Garcia3
1 Department of Oncology-Hematology, Istituto Clinico Humanitas IRCCS, Rozzano, Italy
2 Department of Medical Oncology, Dana-Farber Cancer Institute, Boston
3 Department of Medicine/Medical Oncology, University of Colorado Cancer Center, Aurora, USA
4 CINECA-Interuniversity Consortium, Bologna, Italy
5 Division of Molecular Pathology, University Hospital, Basel, Switzerland
6 Department of Pathology, Milan University, Rozzano, Italy

Background: MET amplification has been detected in 20% of non-small-cell lung cancer patients (NSCLC) with epidermal growth factor receptor (EGFR) mutations progressing after an initial response to tyrosine kinase inhibitor (TKI) therapy.


Patients and methods: We analyzed MET gene copy number using FISH in two related NSCLC cell lines, one sensitive (HCC827) and one resistant (HCC827 GR6) to gefitinib therapy and in two different NSCLC patient populations: 24 never smokers or EGFR FISH-positive patients treated with gefitinib (ONCOBELL cohort) and 182 surgically resected NSCLC not exposed to anti-EGFR agents.


Results: HCC827 GR6-resistant cell line displayed MET amplification, with a mean MET copy number >12, while sensitive HCC827 cell line had a mean MET copy number of 4. In the ONCOBELL cohort, no patient had gene amplification and MET gene copy number was not associated with outcome to gefitinib therapy. Among the surgically resected patients, MET was amplified in 12 cases (7.3%) and only four (2.4%) had a higher MET copy number than the resistant HCC827 GR6 cell line.


Conclusions: MET gene amplification is a rare event in patients with advanced NSCLC. The development of anti-MET therapeutic strategies should be focused on patients with acquired EGFR-TKI resistance.

A rosette-type, self-renewing human ES cell-derived neural stem cell with potential for in vitro instruction and synaptic integration

PNAS, Mar 2009; 106: 3225 - 3230.
Philipp Koch, Thoralf Opitz, Julius A. Steinbeck, Julia Ladewig and Oliver Brüstle.

Abstract:

An intriguing question in human embryonic stem cell (hESC) biology is whether these pluripotent cells can give rise to stably expandable somatic stem cells, which are still amenable to extrinsic fate instruction. Here, we present a pure population of long-term self-renewing rosette-type hESC-derived neural stem cells (lt-hESNSCs), which exhibit extensive self-renewal, clonogenicity, and stable neurogenesis. Although lt-hESNSCs show a restricted expression of regional transcription factors, they retain responsiveness to instructive cues promoting the induction of distinct subpopulations, such as ventral midbrain and spinal cord fates. Using lt-hESNSCs as a donor source for neural transplantation, we provide direct evidence that hESC-derived neurons can establish synaptic connectivity with the mammalian nervous system. Combining long-term stability, maintenance of rosette-properties and phenotypic plasticity, lt-hESNSCs may serve as useful tool to study mechanisms of human NSC self-renewal, lineage segregation, and functional in vivo integration.

Identification of CD133-Positive Radioresistant Cells in Atypical Teratoid/ Rhabdoid Tumor

PLoS ONE. 2008; 3(5): e2090. Published online 2008 May 7
Shih-Hwa Chiou, Chung-Lan Kao, Yi-Wei Chen, Chien-Shu Chien, Shih-Chieh Hung, Jeng-Fan Lo, Yann-Jang Chen, Hung-Hai Ku, Ming-Ta Hsu, and Tai-Tong Wong
Atypical teratoid/rhabdoid tumor (AT/RT) is an extremely malignant neoplasm in the central nervous system (CNS) which occurs in infancy and childhood. Recent studies suggested that CD133 could be considered a marker for brain cancer stem-like cells (CSCs). However, the role of CD133 in AT/RT has never been investigated. Herein we report the isolation of CD133-positive cells (CD133+), found to have the potential to differentiate into three germ layer tissues, from tissues of nine AT/RT patients. The migration/invasion/malignancy and radioresistant capabilities of CD133+ were significantly augmented when compared to CD133−. The clinical data showed that the amount of CD133+ in AT/RTs correlated positively with the degree of resistance to radiation therapy. Using cDNA microarray analysis, the genotoxic–response profiles of CD133+ and CD133− irradiated with 10 Gy ionizing radiation (IR) were analyzed 0.5, 2, 6, 12 and 24 h post-IR. We then validated these microarray data and showed increased phosphorylation after IR of p-ATM, p-RAD17, and p-CHX2 as well as increased expression of BCL-2 protein in CD133+ compared to CD133−. Furthermore, we found that CD133+ can effectively resist IR with cisplatin- and/or TRAIL-induced apoptosis. Immunohistochemical analysis confirmed the up-regulated expression of p-ATM and BCL-2 proteins in IR-treated CD133+ xenotransgrafts in SCID mice but not in IR-treated CD133−. Importantly, the effect of IR in CD133+ transplanted mice can be significantly improved by a combination of BCL-2 siRNA with debromohymenialdisine, an inhibitor of checkpoint kinases. In sum, this is the first report indicating that CD133+ AT/RT cells demonstrate the characteristics of CSCs. The IR-resistant and anti-apoptotic properties in CD133+ may reflect the clinical refractory malignancy of AT/RTs and thus the activated p-ATM pathway and BCL-2 expression in CD133+ could be possible targets to improve future treatment of deadly diseases like AT/RT.

Gene targeting in adult rhesus macaque fibroblasts

BMC Biotechnol. 2008; 8: 31.
Daniel T Meehan, Mary Ann Zink, Melissa Mahlen, Marilu Nelson, Warren G Sanger, Shoukhrat M Mitalipov, Don P Wolf, Michel M Ouellette, and Robert B Norgren, Jr
Background
Gene targeting in nonhuman primates has the potential to produce critical animal models for translational studies related to human diseases. Successful gene targeting in fibroblasts followed by somatic cell nuclear transfer (SCNT) has been achieved in several species of large mammals but not yet in primates. Our goal was to establish the protocols necessary to achieve gene targeting in primary culture of adult rhesus macaque fibroblasts as a first step in creating nonhuman primate models of genetic disease using nuclear transfer technology.

Results
A primary culture of adult male fibroblasts was transfected with hTERT to overcome senescence and allow long term in vitro manipulations. Successful gene targeting of the HPRT locus in rhesus macaques was achieved by electroporating S-phase synchronized cells with a construct containing a SV40 enhancer.

Conclusion
The cell lines reported here could be used for the production of null mutant rhesus macaque models of human genetic disease using SCNT technology. In addition, given the close evolutionary relationship and biological similarity between rhesus macaques and humans, the protocols described here may prove useful in the genetic engineering of human somatic cells.

Intended Use (US)

The Genetix CytoVision® system is an automated scanning microscope and image analysis system.

In the US, CytoVision Karyotyper and CEP XY are For In-Vitro Diagnostic Use.

CytoVision Karyotyper is a rapid metaphase finder and computer aided chromosome analysis system which assists the cytogeneticist in viewing the chromosome displays and looking for cellular anomalies. The Karyotyper enables the qualified cytogeneticist to rapidly and accurately analyze the chromosome banding pattern. All diagnostic decisions are made by the qualified clinician.

CytoVision CEPXY_ENG is an accessory to the CEP® X Spectrum Orange™/CEP® Y Spectrum Green™ DNA Probe kit (Abbott Laboratories. Abbott Park, Illinois, U.S.A) and is limited to the analysis of CEP XY probes via high magnification capture and analysis of interphase nuclei. CEP XY is indicated for use to assess the effectiveness of bone marrow transplantation in opposite-sex transplants. All diagnostic decisions are made by the qualified clinician.

CytoVision Fluorescence In-Situ Hybridization (FISH), Comparative Genomic Hybridization (CGH), SPOT AX, RxFISH Color Chromosome Analysis, and M-FISH - For Research Use Only*. Not for Use in Diagnostic Procedures.

The Flexible Karyotyper System is For Research Use Only*. Not for Use in Diagnostic Procedures

Intended Use (EU and CANADA)

In the EU and Canada, ONLY CytoVision Karyotyper is For In-Vitro Diagnostic Use. All other applications are For Research Use Only*. Not for Use in Diagnostic Procedures.

CytoVision Fluorescence In-Situ Hybridization (FISH), Comparative Genomic Hybridization (CGH), SPOT AX, RxFISH Color Chromosome Analysis, and M-FISH - For Research Use Only*. Not for Use in Diagnostic Procedures.

The Flexible Karyotyper System is For Research Use Only*. Not for Use in Diagnostic Procedures

Note* : A Research Application is not intended for in vitro diagnostic or clinical use, but is intended solely for use in the research setting, for example university or pharmaceutical development. These applications are described as Research Applications or Research Use Only.

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