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With contributions and assistance from Scheme Organisers of the National External Quality Assessment Schemes.
Women's CentreAs part of the activities of Unit 1
The views expressed in this document are those of the meeting participants and do not necessarily reflect the policies of the institutions or companies they are affiliated to.
| Participant | Institution | Country |
| Karsten Held | Scheme Organiser German Constitutional EQA, Fertility Centre, Hamburg, | Germany |
| Oliver Bartsch | Scheme Organiser German Constitutional FISH EQA, Institut fuer Humangenetik, Mainz, | Germany |
| Harald Rieder | Scheme Organiser German Haematology EQA,Dusseldorf, | Germany |
| Reiner Siebert | Scheme Organiser German Haematology EQA, Kiel, | Germany |
| Clemens Müller-Reible | EuroGentest WP1.3 co-leader
Professional Association of German Human Geneticists (BVDH) |
Germany |
| Ros Hastings | EuroGentest Unit 1 co-leader and WP1.4 leader,
Scheme Organiser for UK NEQAS for Clinical Cytogenetics, Oxford, |
UK |
| Rod Howell | EuroGentest project officer, Oxford | UK |
| Bettina Quellhorst-Pawley | Quality Manager, Oxford | UK |
| Els Dequeker | EuroGentest unit 1 leader
Cystic Fibrosis Network, Leuven |
Belgium |
| Joris Vermeesch | WP1.4 co-leader, Leuven | Belgium |
| Francisco Sole | Haematology Scheme Organiser, Barcelona | Spain |
| Carmen Ramos | Constitutional Scheme Organiser, Fundacion Jimenez Diaz, Madrid | Spain |
| Marta Rodriguez de Alba | Deputy Constitutional Scheme Organiser, Fundacion Jimenez Diaz, Madrid | Spain |
| Zuzanna Zemanova | Deputy Haematology Scheme Organiser, Centre of Onco Cytogenetics, Prague | Czech Republic |
| Domenica Taruscio | Scheme Organiser, ISS | Italy |
| Giovanna Floridia | Deputy Scheme Organiser , ISS | Italy |
| Nicole Dastugue | Haematology Scheme Organiser, Toulouse | France |
| Martine Doco-Fenzy | Constitutional Scheme Organiser ,Genetic Department, CHU- Reims | France |
| Kalle Simola | Constitutional Scheme Organiser, Tampere University Hospital | Finland |
| Brigitte Faas | Scheme Organiser, Nijmegen | The Netherlands |
| Philippos Patsalis | The Cyprus Institute of Neurology and Genetics, Nicosia | Cyprus |
The purpose of the EuroGentest network is to improve levels of service of genetic laboratory testing by achieving consensus on appropriate standards throughout Europe. Work package 1.4 (WP1.4) is the branch of the network concerned with cytogenetic external quality assessment (EQA). Participation in EQA is an important aspect of effective and accurate laboratory work in particular as it provides a consistent standard or bench mark to which services should aspire.
Diagnostic cytogenetics is well-established throughout most of Europe, and a high standard of service has been provided in many countries for a number of years. Until recently, however, accepted methods and standards have varied as the discipline has evolved without a significant level of contact between cytogeneticists in different countries. A number of national external quality assessment schemes now operate, with a variety of approaches to assessment. Some rationalisation between national schemes, and the availability of a pan-European scheme, will be valuable in harmonization of standards and strategies, in assessment of emerging technologies and specialised methods, and also in provision of support and education particularly to laboratories in those countries where the national infrastructure is deficient.
Like any pathology laboratory discipline, cytogenetics involves a variety of technical and interpretive skills. Preparation of material suitable for analysis requires specialised technical knowledge in culture of various tissues, staining by various methods for bright-field or fluorescence microscopy, and computerised image acquisition and manipulation; interpretive skills in pattern recognition include chromosome analysis and karyotyping using conventional light microscopy, and analysis of fluorescent colour patterns. Microscopic, electronic and hard-copy images are all subject to analysis. In most instances, interpretation depends on visual recognition of what is normal and what is not, rather than assessing a range of numerical values.
In addition to providing adequate technical and interpretive skills, any effective cytogenetics service must adhere to appropriate turnaround times, and be able to provide interpretative reports written in clear and unambiguous language that can be understood by clinicians, non-specialist health care professionals and patients alike.
Cytogenetics services can be divided into three main categories: prenatal diagnosis, in which chromosome abnormalities (particularly trisomies) are identified by analysis of amniotic fluid, chorionic villi (CVS) and fetal blood samples; postnatal diagnosis of constitutional chromosome abnormalities mostly from peripheral blood samples; and cancer cytogenetics (diagnosis of acquired cytogenetic abnormality), the main area here being detection of abnormalities in bone marrow and peripheral bloods from patients with leukaemia or other haemopoietic disorder.
Research in genetics brings new technologies to the diagnostic arena: these are now having a significant impact on methodology in diagnostic cytogenetics. Increasingly, laboratories find the need to move from conventional microscopic analysis towards more powerful, molecular-based approaches to diagnosis, in particular, micro-arrays, MLPA and QF-PCR. EQA will have to embrace these new technologies, adapting and developing novel methods of assessment through pilot schemes.
There are currently known to be approximately 700 laboratories practising diagnostic cytogenetics across Europe. Not all laboratories provide a comprehensive service, some specialising (for example) in cancer studies or prenatal diagnosis. The approach to organisation of diagnostic cytogenetics varies in different countries. Centralisation of services into larger units dealing with multiple types of referral is common in, for example, the Netherlands and United Kingdom, while small specialist laboratories are more often found in other countries such as Italy. The funding and organisation underpinning services is also variable, with central government funded centres, private laboratories, and units closely associated with academic University departments occurring in different countries.
Table 1 summarises the approximate activity expected in a community where a fully developed comprehensive diagnostic cytogenetics service exists, to which the whole population has access. It is recognised that the figures given are, to a certain extent, dependant on local variation: for example, differences in birth rate and local statutes governing prenatal diagnosis. In general, a laboratory requires one cytogenetic analyst for every 150-300 samples depending on the type of referral. These figures are based on surveys undertaken by the UK Association of Clinical Cytogeneticists Professional Standards Committee and Manpower Group over the last 5 years [see ACC website].
TABLE 1. Samples per one million population
| Overall | Postnatal | Prenatal | Oncology |
| 2200 | 1000 | 700 | 500 |
The EuroGentest network will provide more information on the number of Cytogenetic laboratories and their activity in Europe through the WP1.2. database. This information will in turn then impact on the EQA provision by National EQA schemes.
External quality assessment (EQA) comprises of a system in which laboratories are required to analyse material distributed through a scheme operated by appropriately qualified professionals. Peer-reviewed assessment of the results includes a system of marking designed to identify those participating laboratories which reach, or fail to reach, those standards. An effective EQA scheme will
Agreement on appropriate standards in cytogenetics, and how to measure them, will assist in harmonising the quality of service across the European Union. The basic requirements of an external quality assessment scheme include the following:
Broadly, assessment takes one of two forms: it may be
retrospective, where diagnostic material already handled by the laboratory is submitted to the scheme; or
prospective, where reference material is distributed by the scheme to participants for processing, analysis and reporting. In either case, the scheme may
assess technical, analytical and interpretive parameters. Some schemes include both retrospective and prospective components. Retrospective EQA has the
advantage that the scheme assesses the standard that a laboratory routinely achieves but as the individual cases are unique, inter-laboratory comparison is limited.
Prospective EQA measures what a laboratory is capable of achieving and inter- laboratory comparison is practical as laboratories all receive the same material.
The first Forum Meeting of EuroGentest WP1.4 held in Prague in April 2005 sought to identify the existing scope of cytogenetics services and level of participation in external quality assessment in Europe. Organisers from seven EQA schemes known to be in operation were invited to present details of their schemes and to discuss ways in which EQA might be developed and harmonized. From this meeting it became clear that approximately 400 laboratories participate in external quality assessment, and a further three EQA schemes were identified, see Appendix B.
At the second Forum Meeting, in London in October 2005, agreement was reached to set up a European EQA scheme based on distribution of material through the internet. A document containing details concerning the scope of EQA in Europe was prepared and subsequently posted on the EuroGentest website.
At the third Forum Meeting, in Amsterdam April 2006, details of the structure and standards of the European EQA scheme, CEQA, were agreed (Appendix B), and a system of marking criteria was drawn up (Appendix C). It is anticipated that these standards and marking criteria adopted by CEQA would evolve and in time be incorporated into the current National EQA schemes.
Unlike chemistry or haematology, cytogenetic results do not generally consist of ranges of numerical values, and the lack of measurable parameters can make the design of an effective marking system in EQA more difficult. As a result, the cytogeneticists who have set up the existing National schemes have devised systems of assessment in various different ways, and none is comprehensive, so that different schemes currently measure different parameters. The scope of the eight existing National schemes are given in more detail overleaf.
A slide exchange system for haematological samples began in 2006.
External QA has been in existence since 1994 for routine postnatal cytogenetics, with the introduction of FISH assessment in 2002. The scheme is operated by Labquality Inc. (LQ), a non-profit organisation with dedicated staff and an expert voluntary group who report back to the Scheme Manager. A maximum of 25 laboratories can participate and participants are predominantly from Finland and neighbouring countries (Denmark, Sweden, Norway, Baltic countries). The administrative costs are covered by a €300 fee for the routine postnatal assessment, and €200 for FISH. No prenatal or haematological EQA is offered.
The scheme provides a prospective assessment by distribution of fresh blood samples, and fixed cell suspensions for FISH. Five cases are provided each year. Technical, analytical and interpretive aspects of the work are assessed and subjected to a marking system. The results are tabulated so that participants can compare their performance with the anonymised marks of the other laboratories. There is no contingency for identifying and dealing with poor performance.
This scheme, in its early stages of development, has been set up by ACLF (Association des Cytogénéticiens de Langue Française) in 2006 currently operates by retrospective assessment of images and laboratory clinical reports for amniotic fluid and blood samples, three of each sample type (one normal, one with numerical abnormality one with structural abnormality). Sixty laboratories were invited to participate, and 50% responded for the first round. For the immediate future, the scheme will operate on a voluntary basis. The results of the EQA will be given annually in September at the ACLF conference.
This scheme, in operation since 2005 and run by GFCH (Groupe Francophone de Cytogenetique Haematologique), is entirely separate from the French constitutional EQA scheme. Fixed cell suspension from a single case is distributed to 40 participating laboratories. Results are returned by means of images and interpretive reports. 80% of eligible laboratories currently participate. A web based EQA was introduced for viewing the images this year (2006). Assessors comment on technical (banding) quality, interpretation and reporting. The scheme also collects activity data and produces an annual report which is discussed at the GFCH meeting. Registration is €5 as the Scheme Organiser is voluntary.
In existence since 1989, this scheme has both retrospective and prospective components, with a yearly assessment of two prenatal and two postnatal cases from each component. Since 2005, images have been made available online, and reports submitted online. There is also a rapid testing scheme assessing either FISH or QF-PCR based on distribution of a single amniotic fluid sample once a year.
The voluntary Scheme Organiser reports to the board of the professional association of German human geneticists (BVDH), and the scientific association (GfH), and results are publicised by means of workshops and annual written reports.
Under German law, medical physicians are required to participate in EQA, and there are currently 107 participants, including laboratories from Austria, Switzerland and The Netherlands as well as Germany.
There is a €160.00 handling fee for participation in the whole scheme, or €80.00 to contribute to the retrospective component only. The rapid testing scheme costs €40.00.
This prospective scheme available since 1998, involves distribution of two constitutional samples per year, with assessment of the result and clinical report at a cost of €40.00. Up to 70 laboratories participate, with submissions from Austria and Switzerland as well as Germany. Testing for either microdeletion syndromes or constitutional marker chromosomes may be involved, and EBV transformed cell lines are used, with suspension or slides distributed to participants. The EQA scheme has a voluntary Scheme Organiser but technicians are paid to produce the EQA material.
The German Haematology scheme piloted the distribution of one haematology suspension in 2005. No other information is available on EQA in haematological samples.
The FISH haematological scheme (Quality Control in Tumour Cytogenetics Interphase FISH, QuiTZ-FISH) became available in 2003 as a pilot EQA. This was preceded by a questionnaire based evaluation of current routine applications for interphase FISH in tumour Cytogenetics in Germany. Two prospective QuiTZ-FISH pilot trials, using cell suspensions, have been run for BCR-ABL and one for XY sex-mismatched transplantation. Laboratories from the following countries participate: Germany, Austria, Switzerland, Belgium and Denmark. There is an evaluation panel of up to 10 assessors from three participating countries as well as an independent expert. The results are reported at the annual meeting of Tumour Cytogeneticists and Human Geneticists. Except for participants from Switzerland, the scheme is entirely voluntary and, so far, no administration fee is charged.
Schemes for which no information is available include assessment of RT-PCR in haematological samples.
The Italian Ministry of Health has, since 2001, funded EQA which is free to participants and run by a Scheme Organiser and panel of experts on a voluntary basis.
The assessment is retrospective, with images and laboratory reports submitted to the scheme, and embraces prenatal (amniotic fluid and CVS), postnatal blood, and oncology material. A single case from each area of prenatal, postnatal and haematology cytogenetics is submitted per year. In 2006 cases were submitted by the 51 participating laboratories. Submission to the scheme is not universal in Italy, with approximately 100 centres not participating.
Criteria for standards are based on guidelines provided by the Italian Society of Genetics, Istituto Superiore di Sanita (ISS) and ACC (UK)
All eight Dutch laboratories contribute to the constitutional scheme which has been operating since 2003. This is a retrospective scheme assessing submitted images and laboratory reports to the National Quality Co-ordinator. Assessment of the submissions is undertaken by a panel of representatives from all the participating laboratories. Amniotic fluid, CVS, blood and solid tissue samples and haematological samples are assessed. The scheme is currently free of charge as the Scheme Organiser is voluntary.
A scheme run on the same lines as the constitutional scheme (i.e. submission of images and reports) is available to haematology cytogenetics laboratories
This scheme is organised by the Spanish Association for Prenatal Diagnosis (AEDP), and is run by a committee of five experts. One assessment takes place alternate years, including amniotic fluid and blood samples, with both retrospective and prospective components. Technical, analytical and interpretive parameters are assessed using slides, images and clinical reports. The scheme also undertakes audit of laboratory staffing levels, reporting times, success rates and abnormality rates. The data is presented biennially.
Submissions are assessed against local guidelines drawn up in 1998 (revised 2004) and marked using a locally devised scoring system. Laboratories complying with the standards by achieving a satisfactory score are issued with a certificate of compliance. FISH assessment is planned for 2007.
Approximately half of Spanish laboratories participate at a cost of €150.00 per year, and the scheme is open to Portuguese laboratories also. The Scheme Organiser is voluntary.
The Spanish haematological scheme, in existence since 2000, operates separately from the constitutional scheme, and is organised by the Asociation Espanola de Hematologia y Hemoterapia. The Steering Group undertakes retrospective assessment of submitted images and reports from three to four cases per year, and prospective assessment of FISH (fixed cell suspension) and banded material (slides).
Currently this scheme is voluntary and free of charge, with 15-20 participants from Spain and Argentina. About 30 laboratories are eligible, and some do not yet participate. The Scheme Organiser is voluntary and presents the EQA data annually at the AEHH meeting.
External quality assessment has been in existence since 1982, and the EQA scheme has been a member of the UK NEQAS consortium since 1983. The scheme now has a full-time Scheme Organiser, who is a Consultant-Grade Clinical Scientist funded from fees paid by participating laboratories. The scheme has panels of expert assessors for constitutional and oncology samples and is overseen by a Steering Committee and National Quality Assurance Advisory Panel (NQAAP).
The UK NEQAS EQA scheme accepts participants worldwide and presently includes laboratories from Belgium, Netherlands, Spain, France, Portugal, Malta, Czech Republic, Switzerland, Denmark, Sweden and Hungary. There are currently 83 participants. Nine different EQA and three pilot EQAs are offered; pilot EQAs are free of charge while the established EQA costs on average £200. UK laboratories participate in an average of 10 EQAs while non-UK laboratories participate in an average of five EQAs. The current strategy includes assessment of EQA within the following categories:- prenatal, postnatal (constitutional), oncology, and FISH samples.
For some assessments, slides and reports are submitted by participating laboratories to be distributed to, and assessed by other laboratories. Thus the assessment comprises both a retrospective component for the submitting laboratory and a prospective component for the receiving laboratory. Discrepancies between the results from the submitting and assessing laboratories are judged by a panel of senior cytogeneticists, with incorrect analysis and interpretation subjected to a system of penalty marks.
Prospective assessments are also carried out by analysis of images posted on a secure participants’ website, again with a system of penalty marks for errors. Prospective assessments involve the reporting of Case Scenarios also form part of the EQA process. Audit data on success rates, report times and abnormality rates are collected by UK NEQAS for UK labs (on behalf of the ACC Professional Standards Committee) and non-UK labs.
Laboratories showing persistently poor performance are referred to the NQAAP. EQA returns are examined as part of the laboratory accreditation process undertaken by CPA(UK), and a laboratory with unsatisfactory EQA returns is unable to achieve accreditation status. EQA schemes themselves are subject to accreditation by CPA(UK) and the Clinical Cytogenetics Scheme is currently undergoing this process.
Under the umbrella of EuroGentest Work Package WP1.4, Forum Meetings have been organised to give existing National Scheme Organisers the opportunity to contribute ideas on how best to design an EQA scheme suitable for participants throughout Europe and to contribute to the improvement and harmonization of their National Schemes. In the short term it is not intended that this European scheme, CEQA should replace existing schemes but rather should provide an opening for laboratories in countries without access to an appropriate National EQA scheme, and also address emerging technologies and rare tests.
To date, four Forum Meetings have been convened: Prague April 2005; London, October 2005; Amsterdam May 2006; and Berlin October 2006. Consensus has been reached that the scheme, known as CEQA (Cytogenetics European Quality Assessment), should consist of a web-based prospective system on the lines of the existing UK NEQAS scheme with submission permissible in the native language of any assessor. The languages currently available are Czech, English, French, Finnish, German, Italian and Spanish. A pilot assessment, consisting of one prenatal and one postnatal constitutional case, was made available to invited participants in August 2006. Twenty-four laboratories signed up to participate in the pilot EQA, 15 laboratories submitted a report on line. Some laboratories had never participated in EQA before. The returns were discussed and marked according to an agreed provisional marking system by the assessment panel, consisting of the existing National Scheme Organisers, in an assessors meeting held after the fourth Forum Meeting (Berlin, October 2006).
Possibly as many as 300 European laboratories do not participate in external quality assessment, and for many of these there is no easy access to a EQA scheme. In order to publicise to all practising Cytogenetics that European EQA scheme is being developed, it will firstly be necessary to draw up a register of laboratories in Europe. This will enable feedback from laboratories with regard to their views on, and requirements for, EQA. Eventually, the register will permit CEQA to monitor non-participation.
WP1.4 Forum members have agreed that an effective way to undertake EQA will be to develop an internet-based distribution of images and case scenarios, along the lines of the systems already in place in the UK, France and Germany. It is agreed that existing national schemes should continue to operate, and countries with no EQA scheme would not be discouraged from setting up their own EQA scheme. For reasons of local statutes and policies, different countries inevitably have special needs that can only be addressed by a national scheme. However, it is envisaged that CEQA will supplement local schemes by providing assessment of rare disorders or novel tests not addressed by other schemes. So, as well as providing EQA for laboratories without access to a local scheme, CEQA will supplement the national schemes.
Development of national EQA schemes will be encouraged particularly where language is a barrier to participation in CEQA. However, WP1.4 Forum members agree that, for a scheme to be viable, a significant number of participants must contribute in order to make comparative assessment meaningful. It is recommended that a National scheme should consist of no fewer than eight participants per EQA. Consequently, for some smaller countries a national scheme may not be possible but CEQA will be an alternative.
The ability to agree and harmonize appropriate standards of service is an important product of participation in EQA, and the educational role of EQA schemes must not be underestimated. Over the coming years it is recommended that CEQA use information gathered from its participants and contributors, and other EQA schemes, to oversee the composition of consensus-based generic standard operating procedures.
A mechanism that permits action to be taken in the event of unsatisfactory or poor performance is an important aspect of external quality assessment. An effective way of ensuring that participants make the effort to comply with the standards of the scheme is to have a system of laboratory accreditation: unsatisfactory performance in external QA leads to failure to achieve (or loss of) accredited status. In many countries there is no requirement for accreditation, and a key recommendation must be that the efforts made by the WP1.4 Forum members in devising suitable EQA are also underpinned by development of robust laboratory accreditation systems (see Unit 1 objectives). As well as laboratory accreditation, EQA schemes themselves will eventually require accreditation by a suitable external body (see WP 1.9). WP1.4 and CEQA will be in a position to assist in this regard.
External assessment of novel technology as it moves from research to the diagnostic arena is important, and agreement on best practice for standards and strategies can be achieved by consensus. The traditional boundaries between Cytogenetics and Molecular Genetics are becoming less defined. Currently, cytogenetics is embracing a number of novel methods to supplement its existing use of karyotyping and FISH: these include, for example, array-CGH, pre-implantation genetic diagnosis (PGD), and QF-PCR for prenatal diagnosis. There is an urgent need to agree best practice for such technologies. It is hoped that WP1.4 and CEQA in collaboration with our Molecular Genetic colleagues and cytogenetic organisations such as ECA (European Cytogeneticists Association) will have a significant impact by taking the lead in identifying the needs and organising international meetings with contributions invited from laboratories involved in validating the technology.
In conclusion, it is recommended that CEQA, overseen by a committee drawn from executives of national EQA schemes and representatives of CEQA participants from an appropriate selection of countries, should continue in its development. Its aim will be to provide oversight and advice in external quality assessment throughout Europe as well as practical EQA to those who require it.
The authors would like to thank all members of the Forum for their comments and contributions to this document.
Guidelines for External Quality Schemes (EQA) in Cytogenetics
Background- Retrospective EQA assess the laboratories actual performance by looking at reports sent out by the lab while prospective EQA assesses what a laboratory is capable of achieving. Retrospective EQA has the disadvantage of not comparing like with like i.e. the case is unique to that laboratory. If one laboratory submits a t(13;14) Robertsonian translocation case and another laboratory a complex unbalanced translocation, the latter laboratory is more likely to have errors in the reporting of the case. Consequently a laboratory submitting a complex case is more likely to lose marks than one submitting a simple abnormal case. Prospective EQA overcomes this disparity as all laboratories see the same case (therefore equal weighting) but is the submitted report a true reflection of what is done in the laboratory routinely?
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