Immunocytochemistry. Results - Summary Graphs - Pass Rates Best Methods - Selected Images. Run 98. Assessment Dates: 25th June - 20th July 2012

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1 Run 98 Short Report IN THIS ISSUE UK NEQAS ICC & ISH User Survey 212 Page 4-9 Immunocytochemistry Modules Immunocytochemistry Results - Summary Graphs - Pass Rates Best Methods - Selected Images Assessment Dates: 25th June - 2th July 212 General Pathology: CD31/CD34 & TTF-1 Breast pathology: PR Breast pathology: HER2 IHC Gastric: HER2 IHC Lymphoid pathology: CD3 & BCl Molecular Pathology: Focussing on Patients October 12th 14th 212 A joint meeting provided by UK NEQAS for ICC & ISH and UK NEQAS for Molecular Genetics at The Beaumont Estate Hotel & Conference Centre Old Windsor, UK Neuropathology: Synaptophysin & Tau 5-57 Cytology: CD45 & Calretenin Alimentary Tract: GIST: CD117 & DOG In-situ Hybridisation Modules Breast: HER2 ISH Interpretive Breast HER2 ISH Technical Commercial sponsors in this Issue See Pages 2-3 for Programme and Registration Details UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent

2 General Information Data shown in this article is collated from UK NEQAS ICC & ISH assessments and is presented and described as collected, and does not ether endorse nor denounce any particular product or method and is provided as a guide to highlight optimal and suboptimal staining methodologies. UK NEQAS ICC & ISH does not endorse any of the products featured by the commercial sponsors and are placed at the discretion of UK NEQAS ICC & ISH. Furthermore, commercial companies featured do not have any input or influence over the content, including results that are shown. For further information of the UK NEQAS ICC & ISH scheme, general EQA enquiries, slide returns and advertising opportunities please contact: Dr Merdol Ibrahim, Scheme Manager UK NEQAS ICC & ISH Suite 3/2 Hamilton House Mabledon Place London WC1H 9BB, UK Tel: (+44) merdol.ibrahim@ucl.ac.uk For enquiries concerning training issues, meetings, or courses, please contact: Mr Keith Miller, Scheme Director UK NEQAS ICC & ISH UCL-AD, 21 University Street, University College London London WC1E 6JJ, UK Tel: (+44) k.miller@ucl.ac.uk Director Mr Keith Miller (k.miller@ucl.ac.uk) Manager Dr Merdol Ibrahim (merdol.ibrahim@ucl.ac.uk) Deputy Director Mr Andrew Dodson (a.r.dodson@liverpool.ac.uk) Assistant Manager Ms Suzanne Parry (s.parry@ucl.ac.uk) Office Manager Mrs Ailin Rhodes (a.rhodes@ucl.ac.uk) Quality Manager Mr Neil Bilbe (n.bilbe@ucl.ac.uk) Clerical Assistant Mrs Clara Lynch (clara.lynch@ucl.ac.uk) ASSESSORS United Kingdom Mr C Abbott, Bristol Dr N Atkey, Southampton Dr M Arends, Cambridge Dr M Ashton-Key, Southampton Mrs J Bell, Nottingham Mr N Bilbe, London Mr D Blythe, Leeds Mr J Brown, London Dr L Carson, Aberdeen Ms E Clark, Surrey Mrs A Clayton, Preston Mrs A Cramer, Manchester Dr S Di Palma, Surrey Mr A Dodson, Liverpool Mrs G Donald, Maidstone Dr D Faratian, Edinburgh Mr R Fincham, Cambridge Mr D Fish, Reading Mrs S Forrest, Liverpool Dr I Frayling, Cardiff, Wales Ms J Freeman, London Dr C Gillett, London Ms J Gorst, Bucks Mr J Gregory, Birmingham Prof A Hanby, Leeds Mr N Hand, Nottingham Ms L Happerfield, Cambridge Dr R Hunt, Stockport Dr M Ibrahim, London Mr P Jackson, Leeds Prof B Jasani, Cardiff Mrs N Johnson, Cambridge Ms S Jordan, London Dr J Joseph, Preston Mrs M Judd, Southampton Mrs J MacMillan, Glasgow Mr C Marsh, Newcastle Dr P Maxwell, Belfast Dr G King, Aberdeen Mrs H McBride, Belfast Mr J McGloin, London Dr S McQuaid, Belfast Mr K Miller, London Ms J Moorhead, London Dr M Morgan, Cardiff Ms P Jones, London Ms A Newman, London Mrs L Necus, Kettering Dr G Orchard, London Ms S Parry, London Dr S Pinder, London Dr M Pitt, Preston Mrs F Rae, Edinburgh Dr A Riley, Stirling Mr G Rock, Birmingham Mr J Ronan, Nottingham Dr J Starczynski, Birmingham Mrs C Thomas, Preston Mr P Thompson, Leeds Mr A Watson, Newcastle Mr P Wencyk, Nottingham Mrs H White, Maidstone Mrs J Williams, Portsmouth Ms S Wozniak, Cardiff Australia Mrs J Brincat, Victoria Canada Mrs J Tunnicliffe, Vancouver, Prof. J Bartlett, Toronto Denmark Mr J Askaa, Copenhagen Dr E Baslev, Herlev Dr B Rasmussen, Roskilde Germany Dr Iris Nagelmeier, Kassel Hungary Dr T Krenacs, Szeged Ireland Prof E Kaye, Dublin Mr K McAllister, Dublin Dr T O Grady, Dublin Ms Yvonne Connolly, Dublin Dr Hilary Magee, Dublin Portugal Dr J Cabecadas, Lisbon Dr M Franco, Lisbon Dr F Schmitt, Porto Mr A Ferrero, Lisbon Mrs T Periera, Lisbon Mr R Roque, Lisbon Mr J Matos, Lisbon Slovenia Dr M Flezar, Ljubljana Mrs I Kirbis, Ljubljana South Africa Mrs R Van Wijk, Cape town Sweden Dr G Elmberger, Stockholm Switzerland Prof. Pierre-Andre Diener, St Gallen Journal layout and design prepared by UK NEQAS ICC & ISH UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent

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4 Meeting Announcement Molecular Pathology: Focussing on Patients October 12th 14th 212 A joint meeting provided by UK NEQAS for ICC & ISH and UK NEQAS for Molecular Genetics at The Beaumont Estate Hotel & Conference Centre Old Windsor, UK Friday 12 th October 12. Registration and buffet lunch Lung Cancer: Pathology, Personalised Therapy & Related Biomarkers 14. Introduction by chairman Professor Adrienne Flanagan (UCL Cancer Institute) Histopathology of lung cancer Dr Mary Falzon (University College London Hospitals) 15.3 Refreshments and trade 15.5 Personalized Therapy for Advanced Non-small Cell Lung Cancer Professor Roy Hebst (Yale Comprehensive Cancer Centre) 16.3 ALK Rearrangements in Non-Small Cell Lung Cancer Dr Jens Dhein (Abbott Molecular) 17.2 Summary and discussion Saturday 13 th October Novel Slide Based Testing 9. Introduction by chairman Professor Bharat Jasani (University of Wales College of Medicine) 9.5 Modern slide based predictive testing Dr Kenneth Bloom (GE Health Care- Clarient) 9.5 Current status of IHC4 in breast cancer Professor Mitch Dowsett (ICR Royal Marsden Hospital) 1.35 Refreshments and trade Breast Cancer 11. Usual and unusual molecular Her-2/neu signal patterns in breast carcinoma Dr Matteo Brunelli (Verona, Italy) 11.5 HER2 IQFISH pharmdx. Get your In Situ Hybridization Results in 3½ hours. Performance Evaluation Results from Intra- Laboratory Comparison Study Dr Iris Nagelmeier (Targos) Clinical Perspectives How will this new Technology Change the Use of ISH in Cancer Diagnostics and the value for Oncologist and Patients Dr Kenneth Bloom (GE Health Care- Clarient) Lunch and trade 14. Introduction by chairman Andrew Dodson (Liverpool Royal Infirmary) 14.5 The Roche oncology pipeline Dr Julian Cole (Roche Products Ltd) 14.5 The Roche biomarking pipeline Dr Eric Walk, (Ventana Tucson) Refreshments and trade GI Tract: Pathology, Personalised Medicine & Targeted Therapy 16. Pathology of the GI tract Professor Marco Novelli ( UCLH) The impact of cetuximab therapy on colorectal cancer TBC 17.3 Summary and discussion UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 2

5 Meeting Announcement Sunday 14 th October Session 3 Selected Topics 9. Introduction by chairman Dr Matteo Brunelli (Verona, Italy) 9.5 Aspects of Quality Assessment (Part I) Dr Merdol Ibrahim (UK NEQAS ICC & ISH, UCL) 9.4 UK NEQAS ICC & ISH scheme updates: Technical modules for breast ISH and Gastric ICC HER2 Suzanne Parry (UK NEQAS ICC & ISH, UCL) 1.15 Refreshments and trade 1.45 Oncotype DX Dr Christer Svedman (Genomic Health) 11.3 The role of NICE in biomarker approval Professor Ian Cree (Warwick University Hospital) 12.15: Summary, discussion and presentation of poster prizes Lunch and depart 16. Introduction by chairman Dr Philippe Taniere (Birmingham) 16.5 Metastatic Melanoma The disease and current treatment TBC Metastatic Melanoma The laboratory s perspective Dr David Gonzalez de Castro (Royal Marsden Hospital) UK NEQAS Mock Assessments UKNEQAS ICC & ISH (Meeting Leader Dr Merdol Ibrahim, UKNEQAS ICC & ISH) ALK Workshop Workshop for ALK in Non-Small Cell Lung Cancer (Workshop Leader - Michael Gandy, UCL-Advanced Diagnostics) Saturday 13 th October Satellite Meetings Conference dinner and entertainment from 19. Session 1 Molecular Pathology Meeting Leader Dr Sandi Deans 9.25 Introduction by chairman Dr Sandi Deans (UK NEQAS for Molecular Genetics) 9.3 Aspects of personalised medicine Dr Hendrik-Tobias Arkenau (Sarah Cannon Research UK) TBC 1.3: Refreshments and Trade Sunday 14 th October UK NEQAS - CPT (Meeting Leader Garry Thompson (UKNEQAS-CPT) Workshop on Gastric HER-2 (Workshop Leader - Dr Iris Nagelmeier, Targos) POSTERS Abstracts are invited for posters closing date 31 st August Crizotinib: a wonder drug? Dr Sanjay Popatt (Royal Marsden Hospital) 11.5 Molecular diagnostics in lymphoma Professor Ming Du (Addenbrooke s Hospital) 12.3 Lunch and Trade Session Introduction by chairman Dr Rachel Butler (Cardiff) Royal College and IBMS CPD points to be confirmed For Further Details & Registration Click HERE Or Visit meetings.shtml 14.5 Molecular markers in ovarian cancer Dr James Brenton (CR-UK) TBC 14.5 CRUK Stratified Medicine Dr James Peach (CR-UK) TBC Refreshments and Trade UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 3

6 UK NEQAS ICC & ISH User Survey 212: Participant Feedback Neil Bilbe, Merdol Ibrahim and Keith Miller Introduction As part of the annual requirements of an EQA scheme to assess user satisfaction, a survey was sent to all participants in May 212. Method In May 212, approximately 528 active participants were registered with UK NEQAS ICC & ISH. An , with a link to the online survey, was sent to each of them, along with instructions. Participants were given 1 month to respond the closing date was June 8 th 212. At the start of the survey, there were 5 mandatory fields to complete: Participant Number Region (UK or non-uk) Person completing survey Yearly workloads Modules participated in for year 211/212 (Runs: 94 97) There were also a number of optional fields including How long have you been using our service There were 26 main questions. For the first 16 questions respondents were asked to give a rating of: Very Satisfied Satisfied Neutral Dissatisfied or Very Dissatisfied Question 17 asked how likely participants were to continue using the service; questions 18, 19, 2, 21, 23 and 24 required a Yes or No type response. Questions 22 A, B, & C asked participants to rate the quality of the EQA material. The final two questions (Q. 25 & 26) asked for the overall rating of our service; firstly by satisfaction level, and secondly, with a score out of 1. At the end of the main survey there were 4 additional questions (optional) relating to the use of in-house controls. Return Details 167 single replies were received, (any duplicate, incomplete or multiple entries were deleted), approximately 32% of participants. 67 (4%) of replies were from UK participants 31% of all registered UK participants 1 (6%) of replies were from overseas (OS) participants 32% of all registered non-uk participants. Survey questions and topics Q1. Packaging of samples Q2. Labelling of slides, module identification Q3. Time to stain/return samples Q4. Details in cover letter and online Q5. Procedure for returning samples Q6. Web based format of the results Q7. Assessor comments and feedback Q8. Technical help and advice Q9. Communication with UK NEQAS ICC & ISH Q1. Dealing with complaints Q11. Ease of contacting UK NEQAS ICC & ISH Q12. The UK NEQAS ICC & ISH E-Journal Q13. UK NEQAS ICC & ISH meetings generally Q14. Location of UK NEQAS ICC & ISH meetings Q15. Content or type of meetings Q16. Number of meetings Q17. How likely to continue using our service Q18. Would you recommend service to others Q19. Ever had cause to complain to UK NEQAS ICC & ISH office Q2. Do you treat UK NEQAS ICC & ISH samples differently to your in-house controls/slides Q21. Do you find the assessment of your in-house controls useful Q22. How do you rate the quality of: EQA Tissue sections EQA Cytospins EQA Cell Lines Q23. Do you use your EQA results to try and improve the quality of your in-house staining Q24. Have you ever requested a reassessment Q25. Overall how satisfied with our service Q26. Rating of UK NEQAS ICC & ISH out of 1 UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 4

7 UK NEQAS ICC & ISH User Survey 212: Participant Feedback At the end of each section, or following some individual questions, there were comments sections allowing participants to express their views, or make any comments they felt would be useful to UK NEQAS ICC & ISH management. Overall analysis of satisfaction Results from the response to Q. 25 & Q. 26 relating to overall satisfaction levels. Part 1: Sample and results section (Q. 1 6): (S) Slides & module labelling/identification: 58% (D) Web based format of results/time to stain and return slides (2 topics): 3.5% Part 2: Participant feedback and communication section (Q. 7 12) % of Labs 8% 7% 6% 5% 4% 3% 2% User Survey 212: Level of Satisfaction by Percentage 24.7% 65.66% (S) Ease of contacting UK NEQAS: 3.5% (D) Assessor comments and feedback: 12.5% Part 3: UK NEQAS ICC & ISH meetings section (Q ) (S) UK NEQAS ICC & ISH meetings: 1.1% (D) Location of meetings: 6.5% 1% % 9.64% Very Satisfied (41) 24.7% Satisfied (19) 65.7% Neutral (17) 9.6% Very/Dissatisfied %.%.% V. Satisfied Satisfied Neutral Dissatisfied V. Dissatisfied Part 4: Specific questions about the service (Q ) These required answers not related to satisfaction or dissatisfaction. 17. How likely are you to continue to use our service? Effectively, over 9% of users were either Very Satisfied or Satisfied with the service, 9.6% responded to being Neutral. There were no Dissatisfied responses this year (1 in 211). Response by region: UK (67) OS (1) Very Satisfied 21 (31.3%) 2 (2%) Satisfied 43 (64.1%) 66 (66%) Neutral 3 (4.4%) 14 (14%) Very/Dissatisfied (%) (%) 18. Would you recommend this service to others? N.B. In general, more UK labs expressed higher percentages of satisfaction than OS labs. Replies to individual questions The response to individual questions was not mandatory, therefore the number of participants leaving feedback varied from question to question, and topic to topic. The highest levels of very satisfied (S) and highest level of dissatisfaction (D) for each of the sections is summarised below. 19. Have you ever had cause to complain to UK NEQAS ICC & ISH in last year? UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 5

8 UK NEQAS ICC & ISH User Survey 212: Participant Feedback 2. Do you treat UK NEQAS ICC & ISH samples differently to your in-house sections/samples? Question 23 asked whether labs used their EQA results to improve in-house staining 21. Do you find the assessment of your in-house controls useful? Question 24 asked whether labs had ever requested a reassessment of their EQA result Questions 22 A, B & C related to the quality of the EQA material a. Tissue sections Module Related Data The average number of modules that labs participate in is 3.5; the combined number of modules for all labs responding was 583, the number of labs for each module was: b. Cytology preparations When this is compared to the actual number of labs registered for each module: c. Cell lines On average 35.6% of labs registered for a particular module responded to the survey, with a median of 33.6%. Lowest response 29.4% (Her2 both ICC & ISH), highest 48.1% (Cytology). UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 6

9 UK NEQAS ICC & ISH User Survey 212: Participant Feedback Overall rating of UK NEQAS ICC & ISH out of 1 by module (Q. 26) There is little difference in the two sets of figures, with over 9% of participants satisfied in both surveys. Using the same method employed for the 21 and 211 surveys, the percentage of non-satisfied responses derived from totalling up all the relevant individual questions (Q. 1-15/16) is 3.1% (21), 3.3% (211) and 3.% (212) so again static. Levels of dissatisfaction 211 v 212 The lowest levels of satisfaction in 211 are compared with the results for 212: The average score was 7.8 (same as last year), with a low of 7.7 (cytology and neuropathology) and a high of 8 (HER2 ISH). The median was also 7.8. Overall satisfaction levels by module (Q. 25) Using the combined Very Satisfied and Satisfied percentages returned for each module the results are: Location of meetings (same level) Assessor comments (improved) Online report format/data input (improved) Time given to stain samples (worse) Handling of complaints (improved) Receipt of samples (same) Most of these changes are small +/- 1-2% Topic 211 % (Dissatisfied) 212 % (Dissatisfied) Location of meetings Assessor comments Online reports/data input Handling of complaints Receipt of samples Time given to stain samples Comments, suggestions, and feedback about the service: With a low of 84.2% (cytology) and a high of 96% (HER2 ISH) this mirrors the scores out of 1 (see graph above). Comparisons with data from 211 Survey The levels of satisfaction remain relatively unchanged, but with a slight decrease in the number of Very Satisfied ( 6.8%) responses and an increase in the Satisfied ( 7.1%) responses Responses 211 (%) 212 (%) Very Satisfied Satisfied All other responses Comments were returned throughout the survey; these were collated and categorized into 6 main areas: Sample and results section Participant feedback and communication section UK NEQAS ICC & ISH meetings section Quality of UK NEQAS ICC & ISH EQA material General comments and feedback about the service Comments relating to the in-house controls mini survey (this will form part of a separate article and therefore will not be discussed here) It is not possible to list all the comments, many were not relevant to the section or questions they appeared in or responded to, and several participants made the same or a similar comment. UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 7

10 UK NEQAS ICC & ISH User Survey 212: Participant Feedback Suggestions and Comments with a response from UK NEQAS ICC & ISH Cover letters and s should mention both ICC and the ISH run numbers and references The HER2 ISH run number, along with the ICC run numbers, will appear on future participant communications (letter, etc) Late receipt of slides and hence return of samples (Overseas labs) UK NEQAS ICC & ISH appreciates that slides sent to some countries/regions takes longer than we would like, any labs experiencing particular difficulties are advised to contact the scheme. There is an option for slides to be sent by courier, but the labs have to pay for this. Labs are not penalised for late submissions, but we do audit the receipt times. Data entry still complicated and time consuming In order for us to give the level of feedback we desire (best methods etc) input of data relating to methodologies is extremely important. In order to ensure that labs re-enter important data every time these fields are not saved for each run and are mandatory More information on UK NEQAS ICC & ISH samples (pathology, fixation and processing) The amount of information we are able to provide is dependent on the details we receive from the EQA material supplier, but we will endeavour to obtain as much as we can Variable and conflicting assessor comments particularly for breast modules (ER/HER2) The breast modules are amongst the most difficult to assess, and like any staining method are very subjective; the comments are those of the individual assessors, UK NEQAS ICC & ISH does not audit these prior to reports being generated. Participants are welcome to contact the scheme concerning any report related issues. Cell lines require different (pre) treatments to paraffin sections making staining difficult or tricky This should only apply to certain modules or tissue types, but UK NEQAS ICC & ISH is aware that the preparation (fixation etc) of cell lines may differ from tissue sections Publication of the ISH reference centre methodologies Best methods are available as part of the standard feedback, both online, and in the E-Journal. UK NEQAS ICC & ISH will look at the feasibility of collating the reference centre methods The forthcoming meeting at Windsor will have ICC and ISH workshops. UK NEQAS ICC & ISH is also planning to run some local technical meetings for all modules, as well as possibly introducing webcasts to its educational portfolio Cell blocks more common than cytospins now UK NEQAS ICC & ISH currently uses effusions to create a mixed cellular population. Cytospins are an accepted preparation method in most labs and are a cytological sample. The use of cell blocks may be trialled in the future, but in view of the range of techniques in cytology, cytospins are considered the best compromise at present UK NEQAS ICC & ISH samples do not reflect laboratory day-to-day specimens Samples used by UK NEQAS ICC & ISH are taken from routine surgical cases and normally treated the same as for the diagnostic samples on the same cases. Given the quantity of material required for some of the modules may mean that on occasion, we are unable to obtain sufficient amounts of material resulting in samples being treated differently. The tissue, fixation and section details are provided as part of the online data More antigen specific staining criteria UK NEQAS ICC & ISH provides an overview of the staining requirements and characteristics of the antigens it requests. The tissue type and pathology (where appropriate) should allow labs to glean the expected staining of our EQA samples Website often out-of-date and lacks information The UK NEQAS ICC & ISH website is currently listed as a future scheme enhancement, but with the recent launch of several new modules and other upgrades, is now temporarily on hold. The new UK NEQAS ICC & ISH E-Journal contains the most up-to-date technical feedback and news Variety of antibodies needs to be expanded (cytology) We carried out a user antibody repertoire survey earlier this year, we will be contacting our suppliers with the details of any markers not currently being used by UK NEQAS ICC & ISH so that they may be able to provide suitable material in order for us to do so in the future Slides received broken more than previously The final handling and carrying of the packages sent out to participants is not overseen by UK NEQAS ICC & ISH. This will be discussed to see if arrangements can be made to minimise this as much as possible More technically based meetings and workshops (ISH) with some done online UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 8

11 UK NEQAS ICC & ISH User Survey 212: Participant Feedback Data entry only remembers details for 1 st stain not 2 nd The online entry system should store both sets of data. It is important to remember that until both sets of details have been added, to use the Save button so that you can return at a later date Control material (tumours etc) very precious so are reluctant to use for EQA purposes It is understandable that labs are reluctant to use valuable material for EQA purposes, but EQA samples should ideally be treated/stained alongside routine material demonstrating the same antigen. Admittedly, this creates a problem concerning the use and availability of the control slide(s). If this information is relayed to UK NEQAS ICC & ISH we will be aware of the reason for the use of the particular control material/slide submitted Summary Levels of participant satisfaction remain high at c. 9%. The actual level of Very Satisfied respondents showed a decrease from 31.5% (211) to 24.7% (212). The reason for this appears to be the reduction in Very Satisfied UK participants (down 9.7%). The main areas where there were any significant differences in dissatisfaction between UK and OS labs are for time taken to return slides and the procedure for returning samples. This possibly reflects the increase in OS participants and decrease in UK labs, so is perhaps not unexpected. The percentage of UK labs returning adverse comments concerning: Assessor comments and feedback and The number of UK NEQAS ICC & ISH meetings was higher than for OS labs; all other areas were equal. The overall number of dissatisfied response to all questions was exactly 5:5 between UK and OS labs (32/64 each). The survey results will be reviewed and discussed further, at UK NEQAS ICC & ISH Quality Management meetings, and the Annual Management Review, October 212. UKNEQAS ICC & ISH staff and management would like to thanks all the users and participants for their time and comments whilst completing this survey. If you require any further information, or wish to contact the office concerning this article, please write, call, or n.bilbe@ucl.ac.uk When comparing the score out of 1 results from last year, the overall response is the same at 7.8/1. Only three modules showed any significant difference in scores: 1. General Path (up) 7.6 to HNPCC (down) 8.4 to HER2 ISH (up) 7.7 to 8. The feedback received via the assessor comments, all aspects of the online data input, and the results format, are still the main cause of adverse comments and dissatisfaction, although this is closely followed by the comments from non- UK labs concerning the time taken to receive/stain/return EQA samples. The levels of very satisfied responses for specific questions are relatively static, with two noticeable exceptions: Time to stain samples ( 17%) and Assessor feedback and comments ( 3.5%) The new E-Journal had 72% Very/Satisfied with the rest Neutral. Section/cytospin/cell line quality is acceptable, without being spectacular, with between 6-1% returning an excellent response; the majority (8-9%) responded either Very Good or Good/Acceptable. 4-6% responded Poor and only the EQA cell lines got a response of Very Poor.9% (1 lab). There were no significant differences in the UK or OS lab responses. UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 9

12 The General Pathology Module Run 98 Perry Maxwell and Merdol Ibrahim Gold Standard Second Antibody Antigens Assessed: CD31/CD34 TTF-1 Tissue Sections circulated: Normal Appendix & GIST Normal Lung & Lung Tumour Number of Registered Participants: 357 Introduction Endothelial markers - Gold Antibody CD31: also called Platelet Endothelial Cell Molecule-1 (PECAM- 1) is a transmembrane glycoprotein found at endothelial cell junctions which is thought to play a role in angiogenesis, wound healing and thrombosis. It is expressed on endothelial cells in a wide range of tissues including sinusoidal cells of liver, lymph nodes and spleen. It is also found on megakaryocytes, platelets and other haematopoietic cells including B-lymphocytes particularly in the follicular mantle zone. Its main diagnostic use is in the identification of endothelial tumours as it is said to be a more sensitive marker than either CD34 or Von Willebrand Factor (DeYoung et al). It has been shown to be a good marker of endothelial differentiation. Staining is predominately in the cell membrane but some weaker staining of the cytoplasm can be seen. CD34: is a transmembrane glycoprotein which is expressed on immature haematopoietic stem/progenitor cells, capillary endothelial cells and embryonic fibroblasts. It can also be found in splenic marginal zones, dendritic interstitial cells around vessels, nerves, hair follicles, muscle cells and sweat glands in various tissues. CD34 labels capillaries in most tissues but may be absent in large veins and arteries and is negative in the sinus endothelium of placenta and spleen. CD34 is an excellent indicator of vascular differentiation, regardless of the tumour grade, therefore it is a good marker for vascular tumours (Cerilli et al, Leong). Due to the comparatively wide range of cell types stained by CD34 it is recommended that this antibody is used with a panel of endothelial makers including CD31 and Von Willebrand Factor. There are a number of clones available for CD34, of which, QBEnd/1 is the most popular. Staining with CD34 is restricted to the cell membrane. Features of optimal immunostaining CD31 Strong staining of the endothelial cells in the blood vessels and lymphatic vessels throughout the appendix Strong staining of the T-cells in the base of the lamina propria and weaker staining of the B-cells in the follicular mantle with HIER pretreatment. Minimal background staining. CD34 Strong staining of the endothelial cells in the blood vessels and lymphatic vessels throughout the appendix Strong staining on the dendritic interstitial cells in the lamina propria with HIER pre-treatment Minimal background staining. References 1. DeYoung BR, Wick MR, Fitzgibbon JF et al. CD31: an immunospecific marker for endothelial differentiation in human neoplasms. Applied Immunohistochem. 1993; 1: Cerilli LA and Wick MR. Immunohistology of soft tissue and osseous neoplasms. In: Diagnostic Immunohistochemistry. Dabbs DJ. (Editor). Churchill Livingstone, Philadelphia 22; Leong AS-Y, Cooper K, Leong FJW-M. CD34. In: Manual of Diagnostic Antibodies for Immunohistology (1st Ed.). Oxford University Press, Oxford 1999; Leong AS-Y, Cooper K, Leong FJW-M. Factor VIII RA (Von Willebrand factor). In: Manual of Diagnostic Antibodies for Immunohistology (1st Ed.). Oxford University Press, Oxford 1999; 167. Thyroid Transcription Factor (TTF1)- Secondary Antibody Thyroid transcription factor (TTF1) is part of the family of homeodomain (protein structural domain that binds DNA or RNA) transcription factors and is a gene regulatory factor expressed in the follicular epithelial and parafollicular c-cells of the thyroid, epithelial cells of the lung, brain and pituitary gland. TTF1 is essential for the normal development of both the thyroid and lung, and increased TTF1 immunohistochemical expression if seen in both pulmonary adenocarcinomas and thyroid neoplasms of follicular origin (follicular adenoma and follicular and papillary carcinoma). Not all pulmonary adenocarcinomas have been shown to be positive for TTF1, with a positivity range of between 65-8% (Comperat et al 25), depending on the antibody clone used. TTF1 in small lung carcinomas have been shown in about 96% of cases (Ordonez et al., 2) and in 75% of non-small cell pulmonary carcinomas, but generally not thought to be expressed in typical pulmonary carcinoids, although the literature does have some contrasting views. TTF1 can also be used to differentiate between tumours of primary and metastatic origin e.g. primary lung adenocarcinoma (TTF1+/cdx2-) vs metastastatic gastrointestinal adenocarcinomas (TTF1+/cdx2-). Furthermore, TTF1 alongside other antibodies such as CK 2 can help to further identify tumour types including Merkel cell carcinoma (TTF1-/CK2+) and metastatic small cell lung carcinoma ( TTF1+/CK2-). Features of optimal immunostaining Normal Lung Strong, nuclear staining of the basal epithelial cells lining the bronchial ducts Weak to moderate nuclear staining of bronchial luminal epithelial cells Clean background Lung adenocarcinoma Strong, nuclear staining virtually all neoplastic cells Strong, nuclear staining of the basal epithelial cells lining the bronchial ducts Weak to moderate nuclear staining of bronchial luminal epithelial cells Features of sub-optimal immunostaining CD31 and CD34 Weak or negative staining of the endothelial cells and other elements in the appendix. Non-specific nuclear staining possibly caused by over-pretreatment. Inappropriate staining of, for example, epithelium and lymphocytes, probably due to over-pretreatment Features of suboptimal immunostaining Weak staining of the basal epithelial cells lining the bronchial ducts Weak staining of neoplastic cells of the tumour Non-specific nuclear staining Excessive background staining References 1. Comperat E, Zhang F, Perrotin C, Molina T, Magdeleinat P, Marmey B, Regnard JF, Audouin J, Camilleri-Broet S. Variable sensitivity and specificity of TTF1 antibodies in lung metastatic adenocarcinoma of colorectal origin. Mod Pathol (1): Ordonez NG. Value of thyroid transcription factor-1 immunostaining in distinguishing small cell lung UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 1

13 ΡΥΝ 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε Σελεχτεδ Ιµαγεσ σηοωινγ Οπτιµαλ ανδ Συβ οπτιµαλ Ιµµυνοσταινινγ Φιγ 1. Οπτιµαλ δεµονστρατιον οφ Χ 34 ιν τηε ΥΚ ΝΕΘΑΣ διστριβυτεδ αππενδιξ σεχτιον. Εϖεν ατ λοω ποωερ τηε στρονγ διστινχτ σταινινγ οφ τηε ϖεσσελσ ανδ ενδοτηελιαλ χελλσ ισ χλεαρλψ ϖισιβλε. Σεχτιον σταινεδ ωιτη τηε ακο ΘΒΕνδ αντιβοδψ, 1:5, ακο ΠΤ Λινκ, αυτοσταινερ ανδ ΦΛΕΞ κιτ. Φιγ 2. Γοοδ δεµονστρατιον Χ 34 ιν ΥΚ ΝΕΘΑΣ ΓΙΣΤ σεχτιον, σηοωινγ ιντενσε ωελλ λοχαλισεδ πρεδοµιναντλψ µεµβρανουσ ανδ χψτοπλασµιχ σταινινγ. Σεχτιον σταινεδ ωιτη τηε ακο ΘΒΕνδ αντιβοδψ, 1:5, υσινγ τηε ακο ΠΤ Λινκ, αυτοσταινερ ανδ ΦΛΕΞ κιτ. Φιγ 3. Συβ οπτιµαλ δεµονστρατιον οφ Χ 34 ιν τηε ΥΚ ΝΕΘΑΣ αππενδιξ σεχτιον. Ονλψ α φεω ενδοτηελιαλ χελλσ ανδ ϖεσσελσ αρε δεµονστρατεδ ανδ τηε σταινινγ ισ ϖερψ ωεακ (χοµπαρε το Φιγ1). Σεχτιον σταινεδ ωιτη ακο ΘΒΕνδ αντιβοδψ, 1:, ον τηε Λειχα ΒονδΜαξ, ΕΡ1 ανδ Ρεφινε δετεχτιον κιτ. Φιγ 4. Συβ οπτιµαλ δεµονστρατιον οφ Χ 34 ιν ΥΚ ΝΕΘΑΣ ΓΙΣΤ σεχτιον. Τηε σταινινγ ισ ϖερψ ωεακ ανδ ιν σοµε αρεασ τηε αντιγεν ισ νοτ δεµονστρατεδ (χοµπαρε το Φιγ2). Σεχτιον σταινεδ ωιτη ακο ΘΒΕνδ αντιβοδψ, 1:, ον τηε Λειχα ΒονδΜαξ, ΕΡ1 ανδ Ρεφινε δετεχτιον κιτ. Φιγ 5. Οπτιµαλ δεµονστρατιον οφ Χ 31 ιν τηε ΥΚ ΝΕΘΑΣ αππενδιξ σεχτιον. Χ 31 νοτ ονλψ σταινσ ενδοτηελιαλ χελλσ ανδ ϖεσσελσ, βυτ αλσο µαντλε ζονε Β χελλσ ανδ Τ χελλσ ασ σεεν ιν τηισ ιµαγε. Σεχτιον σταινεδ ωιτη τηε ακο (ϑχ/7α) αντιβοδψ, 1:1, υσινγ τηε ακο ΠΤ Λινκ, αυτοσταινερ ανδ ΦΛΕΞ κιτ. Φιγ 6. Οπτιµαλ δεµονστρατιον οφ Χ 31 ιν τηε ΥΚ ΝΕΘΑΣ ΓΙΣΤ σεχτιον. Τηε τυµουρ ισ νεγατιϖε φορ Χ 31 βυτ τηε ενδοτηελιαλ χελλσ αρε στρονγλψ σταινεδ ασ εξπεχτεδ ανδ τηε βαχκγρουνδ ισ χλεαν. Σεχτιον σταινεδ ωιτη τηε ακο (ϑχ/7α) αντιβοδψ, 1:1, υσινγ τηε ακο ΠΤ Λινκ, αυτοσταινερ ανδ ΦΛΕΞ κιτ. Πριντεδ ατ 16:36 ον Τυεσδαψ, 31 ϑυλψ,

14 ΡΥΝ 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε Σελεχτεδ Ιµαγεσ σηοωινγ Οπτιµαλ ανδ Συβ οπτιµαλ Ιµµυνοσταινινγ Φιγ 7. Οπτιµαλ δεµονστρατιον οφ ΤΤΦ 1 ιν τηε ΥΚ ΝΕΘΑΣ νορµαλ λυνγ σεχτιον, σηοωινγ στρονγ διστινχτ νυχλεαρ σταινινγ ιν τηε επιτηελιαλ χελλσ. Σηοων το βεττερ αδϖανταγε ιν τηε ηιγη ποωερ ινσερτ ϖιεω. Σεχτιον σταινεδ ωιτη τηε Νοϖοχαστρα ΣΠΤ24 αντιβοδψ, 1:15, ον τηε Λειχα ΒονδΜαξ, ΕΡ2 ανδ Ρεφινε κιτ. Φιγ 8. Οπτιµαλ δεµονστρατιον οφ ΤΤΦ 1 ιν τηε ΥΚ ΝΕΘΑΣ λυνγ τυµουρ σεχτιον. ςιρτυαλλψ αλλ τηε νεοπλαστιχ χελλσ σηοω στρονγ διστινχτ νυχλεαρ σταινινγ, ωιτη α χλεαν βαχκγρουνδ. Σεχτιον σταινεδ ωιτη τηε Νοϖοχαστρα ΣΠΤ24 αντιβοδψ, 1:15, ον τηε Λειχα ΒονδΜαξ, ΕΡ2 ανδ Ρεφινε κιτ. Φιγ 9. Γοοδ ΤΤΦ 1 σταινινγ οφ τηε ΥΚ ΝΕΘΑΣ λυνγ τυµουρ σεχτιον. Τηε νεοπλαστιχ χελλσ ανδ βασαλ επιτηελιαλ χελλσ λινινγ τηε βρονχηιαλ δυχτ σηοω στρονγ, διστινχτ νυχλεαρ σταινινγ, ωηιλε τηε λυµιναλ επιτηελιαλ χελλσ σηοω ωεακ νυχλεαρ σταινινγ. Σεχτιον σταινεδ ωιτη τηε ακο 8Γ7Γ3/1 ανδτιβοδψ, 1:2, ον τηε ςεντανα Βενχηµαρκ, ΧΧ1 στανδαρδ. Φιγ 1. Συβ οπτιµαλ δεµονστρατιον οφ ΤΤΦ 1 ιν τηε ΥΚ ΝΕΘΑΣ νορµαλ λυνγ σεχτιον. Αλτηουγη τηε επιτηελιαλ χελλσ αρε σταινινγ ασ εξπεχτεδ, τηερε ισ αλσο ιναππροπριατε βαχκγρουνδ σταινινγ. Σεχτιονσ σταινεδ ωιτη τηε ακο 8Γ7Γ3/1 αντιβοδψ, ον τηε Λειχα ΒονδΜαξ, ΕΡ2 ανδ Ρεφινε κιτ. Φιγ 11. Ποορ δεµονστρατιον οφ ΤΤΦ 1 ιν τηε ΥΚ ΝΕΘΑΣ λυνγ τυµουρ σεχτιον. Τηε σταινινγ ισ ϖερψ ωεακ ανδ διφφυσε (χοµπαρε ωιτη Φιγσ 8 & 9). Σεχτιον σταινεδ ωιτη τηε ςεχτορ ΣΠΤ24 αντιβοδψ, 1:4, πρε τρεατεδ ιν τηε µιχροωαϖε ανδ σταινεδ ον τηε ΒιοΓενεξ ΓενοΜΞ αυτοσταινερ ωιτη τηε ΒιοΓενεξ ΣΣ Πολψµερ κιτ. Φιγ 12. Συβ οπτιµαλ δεµονστρατιον οφ ΤΤΦ 1 ιν τηε ΥΚ ΝΕΘΑΣ λυνγ τυµουρ σεχτιον, σηοωινγ ιναππροπριατε νον σπεχιφιχ χψτοπλασµιχ σταινινγ ανδ λιττλε σπεχιφιχ νυχλεαρ σταινινγ. Σεχτιονσ σταινεδ ωιτη τηε ακο 8Γ7Γ3/1 αντιβοδψ, 1:5, πρε τρεατεδ ιν τηε ακο Πασχαλ ανδ σταινεδ ον τηε αυτοσταινερ ωιτη τηε ΡΕΑΛ κιτ. Πριντεδ ατ 16:36 ον Τυεσδαψ, 31 ϑυλψ,

15 Ρυν 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε ΓΡΑΠΗΙΧΑΛ ΡΕΠΡΕΣΕΝΤΑΤΙΟΝ ΟΦ ΠΑΣΣ ΡΑΤΕΣ 8 ΡΥΝ 98Α Χ 31 / Χ 34 ον ΝΕΘΑΣ Σεχτιονσ Ινδιϖιδυαλ 4 = (%) 14 ΡΥΝ 98Β Χ 31 / Χ 34 ον ιν ηουσε Σεχτιονσ Ινδιϖιδυαλ 4 = (%) 7 5 = (%) 6 = (%) 12 5 = (%) 6 = (%) 6 7 = (%) 8 = 9 (3%) 1 7 = (%) 8 = (%) νο. οφ ρετυρνσ = 5 (1%) 1 = 7 (2%) 11 = 13 (4%) 12 = 22 (6%) 13 = 25 (7%) νο. οφ ρετυρνσ = 1 (%) 1 = (%) 11 = 2 (1%) 12 = 15 (4%) 13 = 15 (4%) 2 14 = 2 (6%) 15 = 41 (12%) 4 14 = 1 (3%) 15 = 22 (6%) 1 16 = 79 (23%) 17 = 55 (16%) 2 16 = 129 (38%) 17 = 53 (16%) = 33 (1%) 19 = 23 (7%) 2 = 9 (3%) = 41 (12%) 19 = 28 (8%) 2 = 23 (7%) Συµµαρψ Συµµαρψ 16 2 = 199 (58%) 16 2 = 274 (81%) = 86 (25%) = 47 (14%) 1 12 = 42 (12%) 1 12 = 17 (5%) 9 = 14 (4%) 9 = 1 (%) Μεδιαν = 14. Μεδιαν = 15. ΡΥΝ 98Χ Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) ον ΝΕΘΑΣ Σεχτιονσ 6 Ινδιϖιδυαλ 4 = (%) ΡΥΝ 98 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) ον ιν ηουσε Σεχτιονσ 14 Ινδιϖιδυαλ 4 = 1 (%) 5 = (%) 5 = (%) 5 6 = (%) 7 = 1 (%) 12 6 = (%) 7 = (%) νο. οφ ρετυρνσ = 7 (2%) 9 = 4 (1%) 1 = 6 (2%) 11 = 11 (3%) 12 = 34 (1%) 13 = 15 (5%) νο. οφ ρετυρνσ = 4 (1%) 9 = (%) 1 = 3 (1%) 11 = 3 (1%) 12 = 19 (6%) 13 = 8 (2%) 2 14 = 13 (4%) 15 = 36 (11%) 4 14 = 14 (4%) 15 = 35 (11%) 1 16 = 59 (18%) 17 = 52 (16%) 2 16 = 137 (42%) 17 = 49 (15%) 18 = 48 (15%) 18 = 32 (1%) = 26 (8%) 2 = 13 (4%) = 13 (4%) 2 = 5 (2%) Συµµαρψ Συµµαρψ 16 2 = 198 (61%) 16 2 = 236 (73%) = 64 (2%) = 57 (18%) 1 12 = 51 (16%) 1 12 = 25 (8%) 9 = 12 (4%) 9 = 5 (2%) Μεδιαν = 13.5 Μεδιαν = 14. Πριντεδ ατ 13:9 ον Τηυρσδαψ, 23 Αυγυστ,

16 Ρυν 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε ΑΝΤΙΒΟ ΙΕΣ ΑΝ ΟΤΗΕΡ ΤΕΧΗΝΙΧΑΛ ΠΑΡΑΜΕΤΕΡΣ ΕΜΠΛΟΨΕ ΒΨ ΠΑΡΤΙΧΙΠΑΝΤΣ ΙΝ ΤΗΕ ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ ΜΟ ΥΛΕ Τηε φολλοωινγ ταβλεσ ρεχορδ τηε νυµβερ οφ παρτιχιπαντσ (Ν) υσινγ εαχη πριµαρψ αντιβοδψ ανδ τηε περχενταγε (%) οφ τηεσε παρτιχιπαντσ αχηιεϖινγ αχχεπταβλε σταινινγ (α σχορε >12/2) ον ΥΚ ΝΕΘΑΣ σεχτιονσ. Γενεραλ Πατηολογψ Ρυν: 98 Γενεραλ Πατηολογψ Ρυν: 98 Πριµαρψ Αντιβοδψ : Χ 31 / Χ 34 Αντιβοδψ εταιλσ Ν % Αβχαµ Χ 31 αβ9498 (ϑχ/7α) 1 1 Βεχκµαν Χουλτερ Χ 34 ΙΜ786 (ΘΒενδ1) 1 1 ΒιοΓενεξ ΑΜ236 5Μ (ΘΒενδ1) Χ 34 1 Χελλ Μαρθυε Χ Μ/ (ϑχ7) 1 1 Χελλ Μαρθυε Χ Μ/76 262/ΧΜΧ33 (ΘΒενδ) 4 75 ακο Χ 31 ΡΤΥ ΦΛΕΞ ΙΡ61 (ϑχ7α) 4 75 ακο Χ 34 ΡΤΥ ΦΛΕΞ ΙΡ632 (ΘΒενδ1) 11 1 ακο Χ 34 ΡΤΥ Ν1632 (ΘΒενδ1) 6 83 ακο Μ823 (ϑχ/7α) Χ ακο Μ7165 (ΘΒενδ1) Χ Λαβςισιον Χ 34 ΜΣ363Π (ΘΒενδ1) 2 1 Λεχια/Νοϖοχαστρα Χ 31 ΝΧΛ Χ 31 1Α1 (1Α1) 7 71 Λειχα/Νοϖοχαστρα Χ 31 ΡΤΥ ΠΑ25 (1Α1) 2 5 Λειχα/Νοϖοχαστρα Χ 34 ΝΧΛ ΕΝ (ΘΒενδ) Λειχα/Νοϖοχαστρα Χ 34 ΡΤΥ ΠΑ212 (ΘΒενδ) 12 1 Οτηερ 6 67 Σεροτεχ Χ 34 ΜΧΑ 547 (ΘΒενδ1) 3 33 ΣΚΨΒΙΟ Χ 34 (ΘΒενδ1) 1 1 Τηερµο Σχι Χ 31 ΜΣ353 (ϑχ/7α) 1 1 ςεχτορ Χ 34 ςπ Χ345 (ΘΒενδ1) 11 1 ςεντανα Χ (ϑχ/7α) 2 5 ςεντανα Χ (ϑχ7) 2 1 ςεντανα Χ (ΘΒενδ1) Πριµαρψ Αντιβοδψ : Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Αντιβοδψ εταιλσ Ν % Βιογενεξ ΜΥ397 ΥΧ 2 5 ΒΟΝ ΡΤΥ ΤΤΦ 1 (ΣΠΤ24) ΠΑ Χελλ Μαρθυε 343Μ 95/96/97 (8Γ7Γ3/1) 9 56 Χελλ Πατη ΜΟΒ 285 (8Γ7Γ3/1) 3 1 ακο ΙΡ56 (8Γ7Γ3/1) 6 1 ακο ΙΣ56 (8Γ7Γ3/1) 3 67 ακο Μ3575 (8Γ7Γ3/1) Ινϖιτρογεν (8Γ7Γ3/1) 4 5 Λειχα Βονδ ΤΤΦ1 ΣΠΤ24 (ΡΤΥ) ΠΑ Λειχα ΝΧΛ Λ ΤΤΦ Λ Μεναρινι ΤΤΦ1 ΜΠ 87 χµ5 1 1 Νεοµαρκερ ΤΤΦ 1 ΜΣ 699 ΠΟ 2 5 Νεοµαρκερσ ΜΣ 69 ΞΞ (8Γ7Γ3/1) 5 8 Νοϖοχαστρα ΝΧΛ Λ ΤΤΦ 1 (ΣΠΤ24) Νοϖοχαστρα ΝΧΛ ΤΤΦ 1 (ΣΠΤ24) 22 1 Νοϖοχαστρα ΠΑ364 (ΣΠΤ24) 5 1 Οτηερ 3 67 ΤΗερµο Σχιεντιφιχ ΜΣ/699Π ςεχτορ ςπ Τ483 (ΣΠΤ24) 1 9 ςεντανα (8Γ7Γ3/1) 1 7 ςεντανα ΤΤΦ 1 (8Γ7Γ311) Ζψµεδ (8Γ7Γ3/1) 5 2 Ζψµεδ (8Γ7Γ3/1) 3 33 Γενεραλ Πατηολογψ Ρυν: 98 Γενεραλ Πατηολογψ Ρυν: 98 Χ 31 / Χ 34 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Ηεατ Μεδιατεδ Ρετριεϖαλ Ν % Ν % Βιοχαρε εχλοακινγ Χηαµβερ ακο Πασχαλ ακο ΠΤΛινκ Λαβ ϖισιον ΠΤ Μοδυλε Λειχα Βονδ ΙΙΙ ΕΡ Λειχα Βονδ ΙΙΙ ΕΡ Λειχα ΒονδΜαξ ΕΡ Λειχα ΒονδΜαξ ΕΡ Μιχροωαϖε Οϖεν ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Οτηερ Πρεσσυρε Χοοκερ Στεαµερ ςεντανα Βενκ ΧΧ1 (Εξτενδεδ) 4 1 ςεντανα Βενκ ΧΧ1 (Μιλδ) ςεντανα Βενκ ΧΧ1 (Στανδαρδ) ςεντανα Βενκ ΧΧ1# (8µινσ) 3 33 ςεντανα Βενκ ΥΛΤΡΑ ΧΧ1 (Εξτεν.) 2 5 ςεντανα Βενκ ΥΛΤΡΑ ΧΧ1 (Μιλδ) ςεντανα Βενκ ΥΛΤΡΑ ΧΧ1 (Σταν.) ςεντανα Βενκ ΥΛΤΡΑ ΧΧ1# (8µινσ) 1 ςεντανα Βενκ ΞΤ ΧΧ1 (Εξτενδεδ) ςεντανα Βενκ ΞΤ ΧΧ1 (Μιλδ) ςεντανα Βενκ ΞΤ ΧΧ1 (Στανδαρδ) ςεντανα Βενκ ΞΤ ΧΧ2 (Ε) 1 1 Ωατερ βατη 68 ΟΧ Ωατερ βατη ΟΧ Ενζψµε Μεδιατεδ Ρετριεϖαλ Χ 31 / Χ 34 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 Ν % Ν % ΑΣ ΠΕΡ ΚΙΤ ακο Προτεινασε Κ (Σ32) 1 1 ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Οτηερ 1 1 ςβσ Βονδ Ενζψµε ςβσ Βονδ Ενζψµε ςεντανα Προτεασε 1 1 ςεντανα Προτεασε 1 (76 218) 3 33 Πριντεδ ατ 13:9 ον Τηυρσδαψ, 23 Αυγυστ,

17 Ρυν 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε Γενεραλ Πατηολογψ Ρυν: 98 Γενεραλ Πατηολογψ Ρυν: 98 Χ 31 / Χ 34 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 ετεχτιον Ν % Ν % Α Μενερινι Πολψµερ (ΜΠ ΞΧΠ) ΑΣ ΠΕΡ ΚΙΤ Βιοχαρε πολψµερ (Μ4Υ534) ΒιοΓενεξ ΣΣ Πολψµερ (Θ 42 ΨΙΚΕ) ΒιοΓενεξ ΣΣ Πολψµερ (Θ 43 ΞΑΚΕ) ακο Ενςισιον ΦΛΕΞ ( Κ8/1) ακο Ενςισιον ΦΛΕΞ+ ( Κ82/12) ακο Ενϖισιον ΗΡΠ/ ΑΒ ( Κ57) ακο Ενϖισιον+ ΗΡΠ µουσε Κ44/5/6/7 2 1 ακο ΗερΧεπ Τεστ Αυτο ρ (Κ527) 1 1 ακο ρβ α µο Ιγ (Ε354) ακο ΡΕΑΛ ΗΡΠ/ ΑΒ (Κ51 ) Λαβςισιον Υλτραςισιον ΛΠ ΗΡΠ (ΤΛ 125 ΗΛϑ) 1 1 Λαβςισιον Υλτραςισιον ΛΠ ΗΡΠ (ΤΣ 125 Η ) Λαβςισιον Υλτραςισιον ΟΝΕ Πολψµερ ( ΤΛ 12/5 Η ϑ/τ Λειχα Βονδ Πολψµερ εφινε ( Σ9713) Λειχα Βονδ Πολψµερ Ρεφινε ( Σ98) ΜεναΠατη Ξ Χελλ Πλυσ (ΜΠ ΞΧΠ) Νονε ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Νοϖοχαστρα Νοϖολινκ Π Σ (ΡΕ7 14/15/28/29 Κ) Οτηερ Ποωερ ςισιον ΠςΒ999 ΗΡΠ ςεχτορ Ελιτε ΑΒΧ Κιτ (ΠΚ 72) ςεχτορ Ελιτε Υνιϖερσαλ ΑΒΧ (ΠΚ 62) ςεχτορ ΙµµΠΡΕΣΣ Υνιϖερσαλ (ΜΠ 75) ςεντανα ιςιεω σψστεµ (76 91) ςεντανα Οπτιςιεω Κιτ (76 7) ςεντανα Υλτραςιεω Κιτ (76 5) Χηροµογεν Χ 31 / Χ 34 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Ν % Ν % Α. Μεναρινι Λιθυιδ Σταβλε ΑΒ κιτ ΑΣ ΠΕΡ ΚΙΤ ΒιοΓενεξ ΑΕΧ 1 ΒιοΓενεξ ΑΒ (Θ 43) ΒιοΓενεξ Λιθυιδ ΑΒ (ΗΚ153 5Κ) ΒιοΓενεξ λιθυιδ ΒΑ (ΗΚ 124 7Κ) ακο ΑΒ Λιθυιδ (Κ3465) 1 ακο ΑΒ+ Λιθυιδ (Κ3468) ακο ΑΒ+ ΡΕΑΛ ετεχτιον (Κ51) ακο Ενςισιον Πλυσ κιτσ ακο ΦΛΕΞ ΑΒ ακο ΡΕΑΛ Ενςισιον Κ57 ΑΒ ακο ΡΕΑΛ Κ51 ΑΒ 2 1 Λαβςισιον (ΤΑ 125 Η ) Λαβςισιον ΑΒ µεναπατη ξχελλ κιτ ΑΒ (ΜΠ 86) Οτηερ Σιγµα ΑΒ ( 56 5) 1 1 Σιγµα ΑΒ ( 5637) Σιγµα ΑΒ ( 595) ςβσ Βονδ Πολψµερ Ρεφινε κιτ ( Σ98) ςεντανα ΑΒ ςεντανα ιϖιεω ςεντανα Υλτραϖιεω ΑΒ ςισιον ΒιοΣψστεµσ Βονδ Ξ ΑΒ Γενεραλ Πατηολογψ Ρυν: 98 Αυτοµατιον Χ 31 / Χ 34 Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Ν % Ν % ΒιοΓενεξ ΓενοΜΞ 6ι ΒιοΓενεξ Οπτιµαξ ακο Αυτοσταινερ ακο Αυτοσταινερ Λινκ ακο Αυτοσταινερ πλυσ ακο Αυτοσταινερ Πλυσ Λινκ ακο ΤεχηΜατε Λαβςισιον Αυτοσταινερ Λειχα Βονδ Μαξ Λειχα Βονδ ΙΙΙ Μεναρινι Ιντελλιπατη ΦΛΞ Νονε (Μανυαλ) Οτηερ Σηανδον Σεθυενζα ςεντανα Βενχηµαρκ ςεντανα Βενχηµαρκ ΥΛΤΡΑ ςεντανα Βενχηµαρκ ΞΤ ΒΕΣΤ ΜΕΤΗΟ Σ Γολδ Στανδαρδ Αντιβοδψ Α σελεχτιον φροµ ϕυστ α φεω οφ τηε βεστ µετηοδσ εµπλοψεδ βψ παρτιχιπαντσ Χ 31 / Χ 34 Μετηοδ 1 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 19/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: ακο Μ7165 (ΘΒενδ1) Χ 34, 3 Μινσ, 21 Χ ιλυτιον 1: 5 Αυτοµατιον: ακο Αυτοσταινερ Λινκ 48 Μετηοδ: ακο ΦΛΕΞ κιτ Μαιν Βυφφερ: ακο ΦΛΕΞ ωαση βυφφερ ΗΜΑΡ: ακο ΠΤΛινκ, Βυφφερ: ακο ηιγη πη ταργετ ρετριεϖαλ σολυτιον, ΠΗ: 9 ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Χηροµογεν: ακο ΦΛΕΞ ΑΒ, 21 Χ., Τιµε 1: 1 Μινσ ετεχτιον: ακο Ενςισιον ΦΛΕΞ ( Κ8/1), 3 Μινσ, 21 Χ Πρεδιλυτεδ Πριντεδ ατ 13:9 ον Τηυρσδαψ, 23 Αυγυστ,

18 Ρυν 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε Χ 31 / Χ 34 Μετηοδ 2 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 18/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: Λειχα/Νοϖοχαστρα Χ 34 ΝΧΛ ΕΝ (ΘΒενδ), 15 Μινσ ιλυτιον 1: 25 Αυτοµατιον: Λειχα Βονδ ΙΙΙ Μετηοδ: ςβσ ΒονδΜΑξ Ρεφινε ΚΙΤ Μαιν Βυφφερ: Βονδ Ωαση Βυφφερ (ΑΡ959) ΗΜΑΡ: Λειχα Βονδ ΙΙΙ ΕΡ2 Χηροµογεν: ςβσ Βονδ Πολψµερ Ρεφινε κιτ ( Σ98) ετεχτιον: Λειχα Βονδ Πολψµερ Ρεφινε ( Σ98), 1 Μινσ Χ 31 / Χ 34 Μετηοδ 3 Παρτιχιπαντ σχορεδ 19/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 2/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: Λειχα/Νοϖοχαστρα Χ 34 ΝΧΛ ΕΝ (ΘΒενδ), 6 Μινσ, 37 Χ ιλυτιον 1: 4 Αυτοµατιον: ςεντανα Βενχηµαρκ ΞΤ Μετηοδ: ςεντανα Υλτραςιεω ΑΒ Μαιν Βυφφερ: ςεντανα ρεαχτιον βυφφερ (95 3) ΗΜΑΡ: ςεντανα Βενκ ΥΛΤΡΑ ΧΧ1 (Σταν.) ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Χηροµογεν: ςεντανα Υλτραϖιεω ΑΒ, 27 Χ., Τιµε 1: 8 Μινσ ετεχτιον: ςεντανα Υλτραςιεω Κιτ (76 5), 8 Μινσ, 37 Χ Πρεδιλυτεδ Χ 31 / Χ 34 Μετηοδ 4 Παρτιχιπαντ σχορεδ 18/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 19/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: ακο Μ7165 (ΘΒενδ1) Χ 34, 3 Μινσ, 2 Χ ιλυτιον 1: 1 Αυτοµατιον: ΒιοΓενεξ ΓενοΜΞ 6ι Μετηοδ: ακο ΦΛΕΞ+ κιτ Μαιν Βυφφερ: ακο ΦΛΕΞ ωαση βυφφερ ΗΜΑΡ: ακο ΠΤΛινκ, Βυφφερ: Ασ Περ Κιτ, ΠΗ: 9 Χηροµογεν: ακο ΦΛΕΞ ΑΒ, 2 Χ., Τιµε 1: 1 Μινσ ετεχτιον: ακο Ενςισιον ΦΛΕΞ+ ( Κ82/12), 15 Μινσ, 2 Χ Πρεδιλυτεδ ΒΕΣΤ ΜΕΤΗΟ Σ Σεχονδαρψ Αντιβοδψ Α σελεχτιον φροµ ϕυστ α φεω οφ τηε βεστ µετηοδσ εµπλοψεδ βψ παρτιχιπαντσ Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Μετηοδ 1 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 18/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: Νοϖοχαστρα ΝΧΛ Λ ΤΤΦ 1 (ΣΠΤ24), 3 Μινσ, 37 Χ ιλυτιον 1: 5 Αυτοµατιον: ςεντανα Βενχηµαρκ ΥΛΤΡΑ Μετηοδ: ςεντανα Υλτραςιεω ΑΒ Μαιν Βυφφερ: ςεντανα ρεαχτιον βυφφερ (95 3) ΗΜΑΡ: Οτηερ Χηροµογεν: ςεντανα Υλτραϖιεω ΑΒ ετεχτιον: ςεντανα Υλτραςιεω Κιτ (76 5) Πριντεδ ατ 13:9 ον Τηυρσδαψ, 23 Αυγυστ,

19 Ρυν 98 ΓΕΝΕΡΑΛ ΠΑΤΗΟΛΟΓΨ Μοδυλε Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Μετηοδ 2 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 18/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: Ινϖιτρογεν (8Γ7Γ3/1), 3 Μινσ, 22 Χ ιλυτιον 1: 2 Αυτοµατιον: ακο Αυτοσταινερ Λινκ 48 Μετηοδ: ακο ΦΛΕΞ+ κιτ Μαιν Βυφφερ: ακο ΦΛΕΞ ωαση βυφφερ, ΠΗ: 7.4 ΗΜΑΡ: ακο ΠΤΛινκ, ΠΗ: 9 Χηροµογεν: ακο ΦΛΕΞ ΑΒ, 22 Χ., Τιµε 1: 7 Μινσ ετεχτιον: ακο Ενςισιον ΦΛΕΞ+ ( Κ82/12), 15 Μινσ, 22 Χ Πρεδιλυτεδ Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Μετηοδ 3 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 16/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: ςεχτορ ςπ Τ483 (ΣΠΤ24), 3 Μινσ, 2 Χ ιλυτιον 1: 2 Αυτοµατιον: Λαβςισιον Αυτοσταινερ Μετηοδ: Στ ΑΒΧ Μαιν Βυφφερ: ΤΒΣ + Τωεεν ΗΜΑΡ: ακο ΠΤΛινκ, Βυφφερ: ακο ηιγη πη ΝΟΤ ΑΠΠΛΙΧΑΒΛΕ Χηροµογεν: Λαβςισιον (ΤΑ 125 Η ) ετεχτιον: ςεχτορ Ελιτε ΑΒΧ Κιτ (ΠΚ 72), 3 Μινσ, 2 Χ Πρεδιλυτεδ Τηψροιδ Τρανσχριπτιον Φαχτορ 1 (ΤΤΦ1) Μετηοδ 4 Παρτιχιπαντ σχορεδ 2/2 (ΥΚ ΝΕΘΑΣ Σλιδε) ανδ 16/2 (Ιν Ηουσε σλιδε) υσινγ τηισ µετηοδ. Πριµαρψ Αντιβοδψ: Νοϖοχαστρα ΝΧΛ Λ ΤΤΦ 1 (ΣΠΤ24), 3 Μινσ, 25 Χ ιλυτιον 1: 1/5 Αυτοµατιον: Μεναρινι Ιντελλιπατη ΦΛΞ Μετηοδ: µεναπατη ξχελλ πλυσ Μαιν Βυφφερ: ΤΒΣ + Τωεεν ΗΜΑΡ: Βιοχαρε εχλοακινγ Χηαµβερ, Βυφφερ: χιτρατε, ΠΗ: 6 Χηροµογεν: µεναπατη ξχελλ κιτ ΑΒ (ΜΠ 86), 25 Χ., Τιµε 1: 5 Μινσ ετεχτιον: ΜεναΠατη Ξ Χελλ Πλυσ (ΜΠ ΞΧΠ), 1 Μινσ, 25 Χ Πρεδιλυτεδ Πριντεδ ατ 13:9 ον Τηυρσδαψ, 23 Αυγυστ,

20 The Breast Hormonal Receptor Module Run 98 Suzanne Parry and Merdol Ibrahim Antigens Assessed: Progesterone Receptors (PR) Tissue Sections circulated: Number of Registered Participants: 325 Sections from a composite block (see table below) Circulated Tissue The table below left shows the staining characteristics of the tissue sections circulated during Run98, which composed of three infiltrating ductal carcinomas (IDCs) with differing levels of receptor expression. The staining of the tumours were characterised using the PGR 16 (A) antibody clone. Tissue Sections % positivity Staining Intensity Allred / Quick Score A. IDC 85-95% High 7-8 B. IDC 11-33% Medium-High 4-6 C. IDC % Negative* General Guideline Used in The Assessment of Slides SCORE Slide not returned by Participant. STAINING PATTERN 1 or 2 No staining or staining of considerably fewer nuclei than expected in one or more of the distributed tissue sections, or inappropriate staining of nuclei in cells not expected to stain. 3 Staining of 1% or greater of tumour nuclei in each of the positive tumour sections, though substantially less than expected to stain, or staining is weaker than expected. 4 or 5 Demonstration of the expected proportion of nuclei stained in the invasive tumours, with roughly the expected staining intensity. Marks are also deducted when correct clinical interpretation of staining may be hindered due to factors such as: - False positive / false negative / non-specific or inappropriate staining - Excessive cytoplasmic or diffuse nuclear staining - Excessively strong or weak haematoxylin counterstain - Excessive Antigen retrieval resulting in morphological damage - Poor quality/inadequate choice of in-house control tissue ( poor/inadequate fixation, damaged cell morphology, over retrieval etc.) In-House Tissue Recommendations & Assessment Participants in-house control tissue MUST consist of composite breast tissue (cell line controls are an acceptable alternative), placed onto a single slide as outlined below: i. >8% tumour positivity with high intensity (Allred/Quick score 7-8) ii. 3-7% tumour positivity with low-moderate intensity (Allred/Quick score 4-6) iii. Negative tumour, with normal positively stained glands (Allred/Quick score <1) Participants NOT using a composite control are assessed a maximum score of a 'borderline' pass (1-12/2). UK NEQAS ICC & ISH. No part of this document can be copied or used without prior written consent 18