User`s Manual User`s Manual
Contents
1. Index INDEX Index A Alcohols 14 16 B Bath solution height 1 10 5 1 CellFyx Solution 5 1 Cleaning Stain Baths 1 10 Clearing Agents 1 7 Clearing agents 1 17 coverslip drying time 1 16 1 17 Coverslipping 1215 Customer Service 3 1 F Fixation 1 3 O Ordering Information 3 1 P Papanicolaou Stain 12 Precautions 1 1 Procedure Notes automated staining 1 10 Protocols for Staining 5 1 Q Quality 1 21 ThinPrep Stain User s Manual Index 1 INDEX 5 Stain Bath Washing 1 10 Staining 1 4 Staining dishes 1 10 Staining Protocols 5 1 Staining reagents 1 5 Staining required materials 1 4 T ThinPrep Bluing II Solution 1 6 ThinPrep Bluing Solution 1 5 ThinPrep Solution 51 ThinPrep EA Solution 1 6 ThinPrep Nuclear Stain 1 5 ThinPrep Orange G Solution 1 6 ThinPrep Rinse II Solution 1 5 ThinPrep Rinse Solution 1 5 Troubleshooting staining 1 12 Warnings 1 1 X Xylene 1 7 Index 2 ThinPrep Stain User s Manual User s Manual ThinPrep STAIN HOLOGIC Hologic Inc 250 Campus Drive Marlborough 01752 US 1 800 442 9892 1 508 263 2900 www hologic com Hologic UK Ltd Link 10 Napier Way Crawley West Sussex e RH10 9 RA UK 44 0 1293 522 0802. Jequinw uoissaooy 1527 JequnN uolsseooV 15114 aoueuaeluleN 1 MAINTENANCE SCHEDULES This page intentionally left blank 2 4 ThinPrepStain User s Manual sasodind doooloUd st sr Tenuen sasn wes oloejslleS 0 Sjueululo 2 4291 20 yoo qd Y29949 OUD urejs MAINTENANCE SCHEDULES This page intentionally left blank 2 6 ThinPrep Stain User s Manual sesodund doooloUd st a8ed Tenuen 5 199 uws SAep 09 Sr p Ieostq x JequnN 101 peuedo peuedgo 1 107 p u do p u do 1 JequnN 01 peuedo peuedgo JequnN 107 p u do p u do 9 pueosiq JequnN 301 peuedo 1 uonnjos asury uonnios dejquiu JealonN 1
3. The slides must remain in the alcohol for at least 10 minutes prior to staining Do not allow the slides to air dry at any time prior to staining e Stainer bath solutions should be kept covered when not in use to minimize evaporation ThinPrep Stain User s Manual 1 3 STAINING WARNING Poisonous Substances Flammable Liquids Refer to SDS section of this manual Follow local regulations or the manufacturer s recommendations for handling and cleanup of spills Refer to manufac turer s SDS for more information Staining For storage temperatures and opened bottle limits of stain solutions refer to the product label or its Instructions for Use Note Filter ThinPrep Nuclear Stain before initial use and when topping up per standard laboratory practice CAUTION For slides intended for use with the ThinPrep Imaging System be sure to follow the staining instructions and applicable protocols exactly as stated Do not substitute any solutions or stains required in the protocols as this may compromise the performance of the ThinPrep Imaging System REQUIRED MATERIALS FOR STAINING From Hologic ThinPrep Nuclear Stain ThinPrep Rinse Solution or ThinPrep Rinse II Solution ThinPrep Bluing Solution ThinPrep Bluing II Solution ThinPrep Orange G Solution ThinPrep EA Solution From Other Suppliers Alcohols 50 reagent or ethyl alcohol e 70 reagent or ethyl alcohol 95 reagent or ethyl al
4. 5 0111 06 65 MAINTENANCE SCHEDULES This page intentionally left blank 2 8 ThinPrep Stain User s Manual Ordering Information ORDERING INFORMATION Ordering Information Mailing Address Hologic Inc 250 Campus Drive Marlborough MA 01752 USA Remittance Address Hologic Inc PO Box 3009 Boston MA 02241 3009 USA Business Hours Hologic s business hours are 8 30 a m to 5 30 p m EST Monday through Friday excluding holidays Customer Service Product orders are placed through Customer Service by phone during business hours at 1 800 442 9892 Option 5 Orders can also be faxed to the attention of Customer Service at 1 800 409 7591 A copy of Hologic s limited warranty and other terms and conditions of sale may be obtained by contacting Customer Service at the numbers listed above ThinPrep Stain User s Manual 1 ORDERING INFORMATION Technical Support For questions about ThinPrep Stain issues and related application issues representatives from Technical Support are available by phone 7 00 a m to 7 00 p m EST Monday through Friday at 1 800 442 9892 Option 6 USA and Canada For Technical Support outside USA and Canada Asia 852 3526 0718 Netherlands 0800 022 6782 Australia 61 2 9888 8000 Norway 800 155 64 Austria 0800 291 919 Portugal 800 841 034 Belgium 0800 773 78 Spain 900 994 197 Denmark 8088 1378 South Africa 0800
5. high quality results for cytologic screening Substitution of reagents and or changes to the Hologic staining protocols may produce less than optimal staining results If protocols are not followed or reagents are substituted stain quality may be compromised Inadequately fixed specimens will not stain well 1 20 ThinPrep Stain User s Manual QUALITY CONTROL FOR THE PAPANICOLAOU STAIN Quality Control for the Papanicolaou Stain PURPOSE Regular monitoring of stain characteristics to Ensure consistent well stained slides for cytodiagnosis Detect stain quality issues prior to screening e Comply with regulatory and accrediting agencies Quality Control requirements PROCEDURE Microscopic review of stained slides based on your laboratory guidelines EXPECTED RESULTS General Expected Results Well stained cells that are transparent Distinct chromatin Good differential cytoplasmic staining e Even staining throughout the slide Reproducible and consistent results Standard for ThinPrep Nuclear Stain Nuclear stain blue to blue black Threads between nuclear segments are visible in flattened polymorphonuclear leukocytes PMNs Standard for ThinPrep Orange G Solution Pale orange to yellow tint in the cytoplasm of non keratinizing superficial squamous cells Bright orange in keratinized superficial cells ThinPrep Stain User s Manual 1 QUALITY CONTROL FOR THE PAPANICOLAOU STAIN Standard f
6. 1 hydrochloric acid in 70 reagent or ethyl alcohol This works particularly well for baths stained with Orange G and EA Solutions b Ifstep 4a does not clean the stain baths containing nuclear stain and those immediately following the nuclear stain use a 10 solution of bleach in water Do not exceed a 10 bleach solution 5 Rinseout each bath several times with water and follow with a final rinse using distilled water Extra care must be taken to thoroughly rinse containers washed using the procedure in 4b since any residual bleach can adversely affect the quality of subsequent staining 6 Allow the baths to air dry or wipe them dry with paper towels before filling them for the next use ThinPrep Stain User s Manual 1 11 STAINING TROUBLESHOOTING Staining Troubleshooting The following table is intended to guide the laboratory through standard troubleshooting practices if they are experiencing issues with the stain quality of slides stained with the ThinPrep Stain Solutions Table 1 Troubleshooting Category Facts Areas to Investigate What percentage of your work is being affected Is there a noticeable pattern in when the problems occur Does it happen when the stainer is changed General Frequency onset duration When was the issue initially seen Is there anyone new performing staining Imager information Are you seeing Imager slide events Are any of your stain s
7. 6419 Xylene Sakura Finetek Xylene 1hour Richard Allan Shandon Xylene Mounting Medium Substitute Histosolve 00 USE with Tissue Tek glass 4111 Thermo Fisher Toluene ee Xylene coverslippers 1 hour also available as Scientific Check ClearVue Mountant Pro Par Clearant mfg recommendations 4211 Anatech Ltd Shandon Xylene CoverSafe American Master Tech Substitute Histosolve Check coverslipper mfg Mounting Medi D Limonene Scientifi dati 1 hour ounting Medium cientific Pro Par Clearant recommendations Anatech Ltd Xylene Pertex Medite Leica Histo Clear HS 200 Biosystems and various National Diagnostics Check coverslipper mfg also available as Xylene ae 1 hour Leica CV Mount distributors recommendations international Histolene JJ031 Fronine Lomb Sci Australia New Zealand DO NOT USE with Tissue Tek glass Thermo Fisher coverslippers Permount Toluene Scientific Xylene 3 hours Check coverslipper mfg recommendations Neutral Balsam Sinopharm Chemical Check coverslipper mfg Xylene Reagent Co Aye recommendations SUUS Electron Microscopy Sciences Leica DPX Mountant Biosystems USA Check coverslipper mfg Caution Xylene Xylene recommendations 1 hour auton Merck and various NOT DEPEX distributors international Leica Biosystems USA ald Xylene Leica Biosystems and Xylene icromoun other distributors recommendations i
8. s Manual MAN 01390 001 Stain Protocol Sakura 2000 One protocol 04329 001 Continuous using 95 alcohol fixed slides Stain Protocol Sakura 2000 One protocol 04330 001 Continuous using CellFyx d slides Stain Protocol Sakura Tissue Tek Prisma One protocol 04904 001 using 95 alcohol fixed slides Stain Protocol Sakura Tissue Tek Prisma One protocol 04905 001 using CellFyx d slides Expansion Stain Protocol Sakura Tissue Tek One protocol 06233 001 Prisma using 95 alcohol fixed slides Expansion Stain Protocol Sakura Tissue Tek One protocol 06234 001 Prisma using CellFyx d slides Stain Protocol Leica Autostainer XL using One protocol 04291 001 95 alcohol fixed slides Stain Protocol Leica Autostainer XL using One protocol 04265 001 CellFyx d slides Stain Protocol Leica 515020 Multistainer One protocol 04906 001 using 95 alcohol fixed slides Stain Protocol Leica 515020 Multistainer One protocol 05586 001 using CellFyx d slides Stain Protocol Shandon Varistain Gemini One protocol 04814 001 using 95 alcohol fixed slides Stain Protocol Medite TST 44 using 95 One protocol 05971 001 alcohol fixed slides ThinPrep Stain User s Manual 3 3 3 4 ORDERING INFORMATION Solutions on the Hologic Compass Stainer using 95 alcohol fixed slides Item Quantity Part Number Stain Protocol Medite TST 44 using CellFyx d One protocol 05972 001 slides Stain Protocol using Rinse
9. slide preparations ThinPrep Orange G Solution The ThinPrep Orange G Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Nuclear Stain ThinPrep Rinse Solution or ThinPrep Rinse II Solution ThinPrep Bluing Solution or ThinPrep Bluing II Solution and ThinPrep EA Solution The ThinPrep Orange G Solution is an alcohol based solution that serves to stain the cytoplasm of cells on cytologic slide preparations It will stain keratin in the cytoplasm of cells while maintaining nuclear staining and nuclear morphology ThinPrep EA Solution The ThinPrep EA Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Nuclear Stain ThinPrep Rinse Solution or ThinPrep Rinse II Solution ThinPrep Bluing Solution or ThinPrep Bluing II Solution and ThinPrep Orange G Solution The ThinPrep EA Solution is an alcohol based solution that serves to stain the cytoplasm of cells on cytologic slide preparations Reagent alcohol or ethyl alcohol Reagent alcohol a blend of 90 ethyl 4 6 methyl and 4 6 isopropyl alcohols or ethyl alcohol may be used in the staining procedure The 100 alcohol must be virgin but 95 may be from approved recycling systems Note For ThinPrep Imaging System applications see the next section RECY CLED SOLVENTS for approved systems and guidelines ThinPrep Stain User s Manual WARNING Poisonous Substance Flammable Liquid Re
10. 980 731 Finland 0800 114 829 Sweden 020 797 943 France 0800 913 659 Switzerland 0800 298 921 Germany 0800 183 0227 UK 0800 032 3318 Ireland Rep 1 800 554 144 Rest of the world 0041 21 633 39 26 Italy 800 786 308 Intl Fax number 0041 21 633 39 10 Protocol for Returned Goods For returns on warranty covered ThinPrep Stain items contact Technical Support Reordering ThinPrep Stain Supply Items Item Quantity Part Number ThinPrep Nuclear Stain One 4 liter bottle 70780 001 ThinPrep Rinse Solution One 4 liter bottle 70779 001 ThinPrep Bluing Solution One 4 liter bottle 70793 001 ThinPrep Orange G Solution One 4 liter bottle 70781 001 ThinPrep EA Solution One 4 liter bottle 70782 001 Stain Set 1 gallon bottle of each of the five 70897 001 stain solutions 32 ThinPrep Stain User s Manual ORDERING INFORMATION Item Quantity Part Number ThinPrep Rinse II Solution One 4 liter bottle ASY 04875 ThinPrep Bluing Solution One 4 liter bottle ASY 04876 ThinPrep Stain Set One 4 liter bottle each of Nuclear PRD 01617 Stain Solution Rinse Solution Bluing Solution Orange Solution and EA Solution Note ThinPrep Rinse and Bluing Solutions are designed to work together and cannot be mixed with or used in conjunction with the original ThinPrep Rinse and Bluing Solutions ThinPrep Stain User s Manual Additional User
11. LABORATORY SOLUTIONS 5 7 ThinPrep STAIN User s Manual CD D o lt D 5 C D HOLOGIC MAN 01390 001 Rev 005 ThinPrep Stain Users Manual HOLOGIC ul Hologic Inc Hologic UK Ltd Australian Sponsor 250 Campus Drive Unit 2 Link 10 Napier Way Hologic Australia Pty Ltd Marlborough MA 01752 USA Crawley West Sussex Suite 402 Level 4 Tel 1 800 442 9892 RH10 9RA 2 Lyon Park Road 1 508 263 2900 United Kingdom Macquarie Park NSW 2113 Fax 1 508 229 2795 Tel 44 0 1293 522 080 Australia Web www hologic com Fax 444 0 1293 528 010 Tel 02 9888 8000 01390 001 Hologic Inc 2014 rights reserved part of this publication may be reproduced transmitted transcribed stored in a retrieval system or translated into any language or computer language in any form or by any means elec tronic mechanical magnetic optical chemical manual or otherwise without the prior written permission of Hologic Inc 250 Campus Drive Marlborough Massachusetts 01752 United States of America Although this guide has been prepared with every precaution to ensure accu racy Hologic assumes no liability for any errors or omissions nor for any dam ages resulting from the application or use of this information Hologic CellFyx and ThinPrep are trademarks or registered trademarks of Hologic Inc in the United States and other countries All other trademarks
12. Staining Protocols The following staining protocols are available from Hologic Stainer Protocol Description Protocol Number Sakura Tissue Tek DRS 2000 Stain Protocol Sakura 2000 04329 001 Slide Stainer Continuous using 95 alcohol fixed slides Sakura Tissue Tek DRS 2000 Stain Protocol Sakura 2000 04330 001 Slide Stainer Continuous using CellFyx d slides Sakura Tissue Tek Prisma Stain Protocol Sakura Tissue Tek Prisma 04904 001 Automated Slide Stainer using 95 alcohol fixed slides Sakura Tissue Tek Prisma Stain Protocol Sakura Tissue Tek Prisma 04905 001 Automated Slide Stainer using CellFyx d slides Sakura Tissue Tek Prisma Expansion Stain Protocol Sakura Tissue 06233 001 Automated Slide Stainer Tek Prisma using 95 alcohol fixed slides Sakura Tissue Tek Prisma Expansion Stain Protocol Sakura Tissue 06234 001 Automated Slide Stainer Tek Prisma using CellFyx d slides Leica Autostainer XL Stain Protocol Leica Autostainer XL using 04291 001 95 alcohol fixed slides Leica Autostainer XL Stain Protocol Leica Autostainer XL using 04265 001 CellFyx d slides Leica 515020 Multistainer Stain Protocol Leica 515020 Multistainer 04906 001 using 95 alcohol fixed slides Leica 515020 Multistainer Stain Protocol Leica 515020 Multistainer 05586 001 using CellFyx d slides Thermo Shandon Varistain Stain Protocol Shandon Varistain Gemini 04814 001 Gemini Slide Stainer using 95 alcohol fixed sl
13. and Bluing One protocol AW 08954 001 Solutions on the Leica Autostainer XL Stain Protocol using Rinse and Bluing One protocol AW 08953 001 Solutions on the Leica 575020 Multistainer Stain Protocol using the Rinse Il and Bluing One protocol AW 08955 001 Solutions on the Sakura Tissue Tek DRS 2000 Stainer Stain Protocol using the Rinse and Bluing II One protocol AW 11301 001 Solutions on the Sakura Tissue Tek Prisma Autostainer Stain Protocol using the Rinse and Bluing One protocol AW 11302 001 Solutions on the Sakura Tissue Tek Prisma Stainer in Expansion Configuration Stain Protocol using the Rinse and Bluing ll One protocol AW 11303 001 Solutions on the Medite TST 44 Stainer Stain Protocol using the Rinse and Bluing II One protocol AW 11304 001 ThinPrep Stain User s Manual 5199116 ejed Safety Data Sheets SAFETY DATA SHEETS Safety Data Sheets ThinPrep Nuclear Stain Solution ThinPrep Rinse Solution ThinPrep Bluing Solution ThinPrep Orange G Solution ThinPrep EA Solution CellFyx Solution ThinPrep Rinse II Solution ThinPrep Bluing 1 Solution The Safety Data Sheet SDS for each solution may be requested from Hologic Technical Support at 1 800 442 9892 Option 6 USA and Canada ThinPrep Stain User s Manual 1 SAFETY DATA SHEETS This page intentionally left blank 42 ThinPrep Stain User s Manual 5 0203014 SUTUTE S Staining Protocols STAINING PROTOCOLS
14. are the property of their respective companies Document Number AW 03790 001 Rev 004 Contents OF 5 Table Contents INTRODUCTION gcn ion CORO a Qe 11 THE PAPANICOLAOU STAIN PAP STAIN 1 2 Principle 1 2 Specimen Requirement 12 FIXATION 1 3 Principle 1 3 Fixation Procedure 1 3 STAINING 14 Required Materials for Staining 14 Reagents 1 5 PROCEDURES FOR STAINING 1 10 Procedure Notes for Automated Staining 1 10 STAINING 5 1 12 COVERSLIPPINU 4242235 ER VASE pa coco P a 1 15 Purpose L21 Coverslipping Requirements 1 15 LIMITATIONS OF FIXATION STAINING AND COVERSLIPPING 445342 p ER 1 20 QUALITY CONTROL FOR THE PAPANICOLAOU STAIN 121 1 21 Procedure 1 21 Expected Results 121 Notes 1 22 BIBLIOGRAPHY 1 23 ThinPrep Stain User s Manual iii iv TABLE OF CONTENTS N MAINTENANCE SCHEDULES 21 ORDERING 31 SAFETY DATA EUM CE P 41 STAINING ORAS S 5 1 INDEX ThinPrep Stain User s Manual uorpnpodju Introduction INTRODUCTION Introducti
15. ble 4 Sakura Tissue Tek FILM Coverslipper Settings for Use with the ThinPrep Imaging System Setting Tissue Tek FILM Automated Coverslipper Coverslip Length 45 mm Xylene Dispensing Range 1 5 Setting 1 Xylene Holding Bath Full Bath Spring Tension Setting Adjust to maximum tension Remove staining racks from instrument immediately after cover slipping is complete Do not let them sit in the output carousel Remove excess xylene dab base of stain rack on Kimwipe or Slide Drying paper towel and stand the rack up so the slides are vertical Immediately remove slides from rack and wipe excess xylene from back if slide with Kimwipe Slides are now ready to label if needed Coverslip Review If any of the following are noted prior to imaging of the slide the slide should be re coverslipped per standard laboratory practice e bubbles CAUTION Large numerous air bubbles can obscure cellular material Retraction of mounting medium e Coverslip is not straight or overhangs the edge of the slide Reagents Follow manufacturer s recommendations for handling and cleaning up spills Refer to manufacturer s 505 for further information ThinPrep Stain User s Manual 1 19 LIMITATIONS OF FIXATION STAINING AND COVERSLIPPING Limitations of Fixation Staining and Coverslipping The ThinPrep reagents and protocols have been developed by Hologic to produce consistent and standardized
16. cohol 100 reagent or ethyl alcohol Clearing Agents e Xylene Shandon Xylene Substitute Thermo Fisher Scientific Clearant Anatech Ltd e Histo Clear HS 200 National Diagnostics e Histolene JJ031 Fronine Lomb Scientific Australia New Zealand 14 ThinPrepStain User s Manual STAINING Water Quality Distilled or deionized water that meets these Hologic specifications for Instrument Feed Water CLSI C3 A4 standard 2006 gt 1 0 microSiemens cm conductivity or gt 1 0 megOhm cm resistivity REAGENTS ThinPrep Nuclear Stain The ThinPrep Nuclear Stain is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Rinse Solution or ThinPrep Rinse Solution ThinPrep Bluing Solution or ThinPrep Bluing II Solution ThinPrep Orange G Solution and ThinPrep EA Solution The ThinPrep Nuclear Stain is an aqueous solution that serves to stain the nuclei of cells on cytologic slide preparations while maintaining cytoplasmic morphology ThinPrep Rinse Solution The ThinPrep Rinse Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Nuclear Stain ThinPrep Bluing Solution ThinPrep Orange G Solution and ThinPrep EA Solution The ThinPrep Rinse Solution is an aqueous solution that removes excess ThinPrep Nuclear Stain from cellular components prior to bluing in cytologic slide preparations enhancing nuclear staining and clar
17. fer to manufac turer s SDS for more information STAINING Clearing Agents Xylene For ThinPrep applications or for slides intended for use with the ThinPrep Imaging System the following xylene alternatives are approved Shandon Xylene Substitute Thermo Fisher Scientific Pro Par Clearant Anatech Ltd Histo Clear HS 200 National Diagnostics Histolene 1031 Fronine Lomb Scientific Australia New Zealand CAUTION ThinPrep Imaging System applications require the clearing agent to be matched to the coverslipping media See Table 3 on page 1 18 RECYCLED SOLVENTS Systems and Guidelines Alcohols recycled via the following systems are acceptable for use with the ThinPrep Imaging System Filtration Systems Alcohol Creative Waste Solutions Inc Bench Top Alcohol Recycling System www cwsincorp com or tel 888 795 8300 SunCycle Technologies Inc Gravity Recycling Alcohol Cartridge GRAC www suncycletech com or tel 866 786 2925 Distillation Systems Alcohol and Xylene CBG Biotech www cbgbiotech com tel 800 941 9484 B R Instrument www brinstrument com or tel 410 820 8800 Recycled Alcohol Guidelines General Guidelines applies to all alcohol recycling Laboratories must follow manufacturer s recommendations for alcohol recycling e Only Hologic recommended alcohols should used reagent or ethyl alcohol ThinPrep Stain User s Manual 1 7 18 STAINING Laboratories are respo
18. gic screening SPECIMEN REQUIREMENT ThinPrep Pap Test slides processed on all approved ThinPrep Processors ThinPrep Stain User s Manual FIXATION Fixation PRINCIPLE Fixatives are applied to specimens to preserve cytologic morphology Immediate and proper fixation is critical to accurate cytologic diagnosis FIXATION PROCEDURE Slides prepared on the ThinPrep 3000 Processor are automatically treated with fixative CellFyx Solution in the instrument No further fixation is required All other slides are deposited into a bath containing standard laboratory alcohol fixative defined as 95 ethyl or reagent alcohol For slides not intended for use with the ThinPrep Imaging System certain laboratory procedures may require alternate fixatives in the bath or for it to remain empty For slides which must be shipped prior to staining CellFyx Solution fixative must be applied Refer to the CellFyx Solution Instructions for Use for details of applying CellFyx Solution to slides prior to shipping Contact Hologic Customer Service for ordering CAUTION No other fixative has been validated for transport For slides fixed in alcohol and intended for use with the ThinPrep Imaging System Note that the alcohol in the fixative bath should be changed after 100 slides or daily whichever comes first Slides should be placed in a multi slide holder staining rack immersed in a bath of 95 reagent alcohol or 95 ethyl alcohol
19. he coverslips are centrally placed and do not overhang any edges of the slide Glass Coverslip Drying Time After coverslipping slides must be allowed to adequately dry before being placed in the Thin Prep Imaging System A slide can be considered dry when the medium has cured sufficiently so there is no movement of the coverslip in response to normal laboratory handling Table 3 illustrates drying times for slides placed in an oven set at 37 C 3 C after automatic coverslipping Note Drying times will differ depending upon variables such as the type of mounting medium quantity of medium used mounting method tem perature and humidity Manually coverslipped slides may have drying times longer than those indicated in Table 3 It is the responsibility of the laboratory to monitor and ensure that adequate but not excessive medium is applied to the slides and that they dry sufficiently prior to being placed on the ThinPrep Imaging System CAUTION Failure to allow slides to dry sufficiently may compromise the performance of the Imager 116 ThinPrep Stain User s Manual COVERSLIPPING Table 2 ThinPrep Imaging System Mounting Media Clearing Agent Validation Summaries Mounting Media Distributor or Automatic Coverslip Minimum Dry Time Mounting Media Media Solvent Manufacturer Clearing Agent System 37 C gt 0 Tissue Tek Glas Check coverslipper mfg
20. ides Medite TST 44 Slide Stainer Stain Protocol Medite TST 44 using 95 05971 001 alcohol fixed slides Medite TST 44 Slide Stainer Stain Protocol Medite TST 44 using 05972 001 CellFyx d slides ThinPrep Stain User s Manual 5 1 5 2 STAINING PROTOCOLS Stainer Protocol Description Protocol Number Leica Autostainer XL Stain Protocol using Rinse and Bluing AW 08954 001 Solutions on the Leica Autostainer XL Leica ST5020 Multistainer Stain Protocol using Rinse Il and Bluing AW 08953 001 Solutions on the Leica 515020 Multistainer Sakura Tissue Tek DRS 2000 Stain Protocol using the Rinse and AW 08955 001 Slide Stainer Bluing Solutions on the Sakura Tissue Tek DRS 2000 Stainer Sakura Tissue Tek Prisma Automated Slide Stainer Stain Protocol using the Rinse Il and Bluing Solutions on the Sakura Tissue Tek Prisma Autostainer AW 11301 001 Sakura Tissue Tek Prisma Automated Slide Stainer Stain Protocol using the Rinse Il and Bluing Solutions on the Sakura Tissue Tek Prisma Stainer in Expansion Configuration AW 11302 001 Medite TST 44 Slide Stainer Stain Protocol using the Rinse Il and Bluing Solutions on the Medite TST 44 Stainer AW 11303 001 Hologic Compass Stainer Stain Protocol using the Rinse and Bluing Solutions on the Hologic Compass Stainer using 95 alcohol fixed slides AW 11304 001 ThinPrep Stain User s Manual
21. ifying cytoplasms ThinPrep Rinse II Solution The ThinPrep Rinse II Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Nuclear Stain ThinPrep Bluing II Solution ThinPrep Orange G Solution and ThinPrep EA Solution The ThinPrep Rinse Solution is alcohol based solution that removes excess ThinPrep Nuclear Stain from cellular components on slide preparations for cytologic evaluation enhancing nuclear staining and clarifying cytoplasms ThinPrep Bluing Solution The ThinPrep Bluing Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Rinse Solution ThinPrep Nuclear Stain ThinPrep Orange G Solution and ThinPrep EA Solution The ThinPrep ThinPrep Stain User s Manual 1 5 16 STAINING Bluing Solution is an aqueous alkaline solution causes cellular components stained with ThinPrep Nuclear Stain to darken to blue violet enhancing nuclear morphology in cytologic slide preparations ThinPrep Bluing II Solution The ThinPrep Bluing II Solution is intended to be used in a Papanicolaou staining procedure in conjunction with the ThinPrep Rinse II Solution ThinPrep Nuclear Stain ThinPrep Orange G Solution and ThinPrep EA Solution The ThinPrep Bluing II Solution is an aqueous alkaline solution that darkens the cellular components stained with the ThinPrep Nuclear Stain to blue violet enhancing nuclear morphology in cytologic
22. ional Research Council Recommendations Concerning Chemical Hygiene in Laboratories Non Mandatory 1910 1450 App A Gill Gary W 1999 The Papanicolaou Stain Quality Control and Quality Assurance SCAN 10 2 18 9 21 Gill Gary W 2001 WHAT IS THE PAPANICOLAOU STAIN ANYWAY SCAN 12 2 33 60 Triol JH and Goodell RM editors ASCT Cytopathology Quality Assurance Guide Second Edition Volume 1 1992 1 24 ThinPrep Stain User s Manual Maintenance Schedules MAINTENANCE 9CHEDULES Maintenance Schedules maintenance schedules the following pages may be used for photocopy purposes e ThinPrep Stain Maintenance ThinPrep Stain Quality Control Check e ThinPrep Stain Solutions Inventory ThinPrep Stain User s Manual 1 MAINTENANCE SCHEDULES This intentionally left blank 2 2 ThinPrepStain User s Manual sasodind doooloUd st ased 5141 TenueJN sasn ureys 5 05 IP SE INOA jr 04 jo Jur 1 41949 epis 1 spenbo sepi s jo y UeUA suonngos asueyD ald
23. ng drying guidelines Sample Biology Certain Imaging System slide events for example 6615 or 6630 may be associated with patient biology when slides have excessive amounts of inflammation bacteria certain lubricants or blood Contact Hologic Technical Support for guidance if there is an increase in such slide events Commonly Asked Questions and Answers 1 Does Hologic have a recommended stainer bath washing procedure Yes please see page 1 10 of this manual for details Can a lab destain re stain and then image slides if an applications issue arises which would normally call for this process Hologic does not support destaining and re staining slides intended for imaging with the ThinPrep Imaging System Does Hologic support staining slides by hand that are intended for imaging Hologic does not support hand staining slides that are intended for imaging Does Hologic have a Non Gyn protocol which uses the ThinPrep stain solutions The ThinPrep Stain may be used for Non Gyn slides however Hologic does not provide a protocol for this Can I substitute non Hologic stain solutions in stainers for slides that are intended for imaging ThinPrep Stain User s Manual 3 STAINING TROUBLESHOOTING Hologic does not recommend using other stain solutions for slides intended for imaging with the ThinPrep Imaging System 6 Cana lab self validate stainers protocols and mounting medium clearing agent combinations for
24. nsible for ensuring that recycled alcohol concentration used in the ThinPrep staining protocol meets Hologic s recommendation Filtration Systems Alcohol only Periodically have the output product independently tested i e annually at a minimum to confirm proper function of the recycling system Contact the manufacturer regarding this service Only the 95 and 100 alcohols from the ThinPrep Stain setup should be put through the filtration system 95 and 100 alcohols combined on the filtration system will result in an output alcohol concentration that will need to be adjusted to 95 prior to use in the ThinPrep Stain setup Distillation Systems Alcohol Periodically have the output product independently tested i e annually at a minimum to confirm proper function of the recycling system Contact the manufacturer regarding this service Only alcohols from the ThinPrep Stain setups can be distilled together e Alcohols from non ThinPrep stain setups must not be co mingled for recycling with ThinPrep alcohols e ONLY 70 95 amp 100 Reagent Alcohol or Ethyl Alcohol may be combined for recycling Note 50 alcohol must be discarded according to the laboratory s hazardous chemical waste disposal guidelines If included for recycling its high water content will reduce the recovery volume and signifi cantly lower the output alcohol concentration Testing Test the collected waste alcohol
25. nternational ThinPrep Stain User s Manual 1 17 COVERSLIPPING Use of Coverslipping Tape Film 1 Follow manufacturer s recommendations for proper storage and operating conditions of the coverslipping product Place coverslipped slides flat in cardboard slide folders for a five minute dry time at room temperature Slides are then ready to be labeled if needed and imaged Slides may be imaged up to twenty eight 28 days after coverslipping If using the Sakura Tissue Tek FILM Coverslipper Refer to Table 3 for setup guidelines Remove staining racks from the instrument immediately after coverslipping in complete Do not let them remain in the output carousel Remove excess xylene by blotting the base of the stain rack on a lint free wipe or paper towel Then stand the rack up so slides are vertical Promptly remove slides from the rack one at a time wiping excess xylene from the back of each slide with a lint free wipe Place slides flat in slide tray for drying Table 3 Coverslipping Film Clearing Agent Validation Summary NET Minimum Dry Time Coverslip Distributor Coverslip Solvent Manufacturer Clearing Agent at Room Temperature Tissue Tek Coverslipping Film Xylene activated Sakura Finetek Xylene 5 min 4470 KP Tape Klinipath 3020 activated The Netherlands Xylene 5 min 1 18 ThinPrep Stain User s Manual COVERSLIPPING Ta
26. olutions expired Stains Check Solution Lot your Nuclear Stain solution open for 60 days or less numbers Did you filter your Nuclear Stain solution before use Verify staining protocol Refer to the protocol Verify stain bath layout Refer to the protocol Verify solutions in bath at the appropriate fill level Staining Verify slides fully submerged in the post stain solution baths Are you following required 0 throughout the process Stainer stainer staining protocols Are you within the maximum allowable throughput limits for the stain table Are you changing your post Nuclear Stain water per protocol Are you cleaning stainer baths per recommended procedure Refer to the Recommended Stain Bath Washing Procedure on page 10 112 ThinPrep Stain User s Manual STAINING TROUBLESHOOTING Table 1 Troubleshooting Category Facts Areas to Investigate Staining Other Solutions Water quality Source How often or last time the system was Water verified Do you make alcohols or buy alcohols If you make alcohols did you check the concentration of alcohols Alcohol before use Are you using recycled alcohol and following manufacturer s guidelines Are you using an approved mounting medium clearing agent Clearing Agents and combination Refer to tables 2 and 3 of this section Mounting Medium Are you following the slide handli
27. on This User s Manual is intended for customers using Hologic products to stain ThinPrep Pap Test slides for cytologic screening The ThinPrep Stain User s Manual provides our customers with standardized fixation and staining protocols using Hologic products in order to achieve consistent high quality results Technical information on reagents and protocols basic troubleshooting techniques and Maintenance Schedules are included The ThinPrep Stain User s Manual is written in a format to make it convenient for our customers to incorporate this information into their individual Laboratory Procedure Manuals WARNINGS PRECAUTIONS For proper handling storage and disposal of each chemical refer to the recommendations on the manufacturer s Safety Data Sheet SDS As with all laboratory procedures universal precautions should be followed ThinPrep Stain User s Manual 1 12 THE PAPANICOLAOU STAIN STAIN The Papanicolaou Stain Pap Stain PRINCIPLE Papanicolaou staining is the universal method for staining gynecologic slides for cytologic diagnosis When performed properly it results in Crisp nuclear detail Differential cytoplasmic staining Transparent cells The ThinPrep Stain is used in conjunction with the ThinPrep Stain protocol and recommended reagents The ThinPrep reagents and protocols have been developed by Hologic to produce consistent and standardized high quality results for cytolo
28. or ThinPrep EA Solution Non keratinizing superficial and intermediate cells vibrant pinks and greens Nuclei and edges of overlapping cells are visible through the cytoplasm PROCEDURE NOTES e Record results of microscopic review Record problems discrepancies and the remedial action taken 1 22 ThinPrep Stain User s Manual BIBLIOGRAPHY Bibliography Allen K editor ASCT A Guide to Cytopreparation 1998 Atkinson B Atlas of Diagnostic Cytopathology WB Saunders Company 1992 Bibbo M and Day L editors Comprehensive Cytopathology WB Saunders Company 2 edition 1997 CLSI Preparation and Testing of Reagent Water in the Clinical Laboratory Approved Guideline Fourth edition C3 A4 2006 Current Regulations page CDC web site Available at www cdc gov clia regs toc aspx Accessed November 7 2011 Commission on Laboratory Accreditation Laboratory Accreditation Program Cytopathology Checklist 2001 Hologic Inc ThinPrep 2000 Processor Operator s Manual Part Number MAN 01408 001 Hologic Inc ThinPrep 3000 Processor Operator s Manual Part Number MAN 01399 001 Hologic Inc ThinPrep 5000 Processor Operator s Manual Part Number 71723 001 Hologic Inc Image Processor Operator s Manual Part Number MAN 01445 001 Keebler CM Somrak TM editors The Manual of Cytotechnology 7th Edition 1997 ThinPrep Stain User s Manual 1 23 BIBLIOGRAPHY Regulations Standards 29 CFR Nat
29. same note for length required for ThinPrep plus Imaging slides Required Materials Table 2 lists the mounting media clearing agents coverslips and combinations of these items that have been evaluated and approved for use with the ThinPrep Imaging System Table 3 lists coverslipping films approved for use with the ThinPrep Imaging System Automated Glass Coverslipping Refer to manufacturer s instructions for use of automated coverslipping instruments Note Refer to Table 4 for recommendations for setting up a Sakura Tissue Tek FILM coverslipper for use with a ThinPrep Imaging System Note Refer to detailed instructions for use of coverslipping tape film on page 1 18 ThinPrep Stain User s Manual 5 COVERSLIPPING Manual Glass Coverslipping 1 Quickly drain excess clearing agent from the slide 2 Immediately apply sufficient mounting medium to the slide to minimize cornflaking air bubbles or retraction Avoid excessive medium which may ooze out from under the coverslip and run down the edges of the slide 3 Apply the coverslip using standard laboratory practice Using a lint free wipe moistened in clearing agent wipe the edges of the coverslip to clear any residual mounting medium 5 Allow slides to dry completely a minimum of 16 hours at room temperature or in a slide oven set at 37 C 3 C for 3 to 12 hours before placing in Imager slide cassettes Refer to the table on page 1 18 CAUTION Make sure t
30. slides intended for imaging with the ThinPrep Imaging System Hologic does not recommend that self validating does not provide the level of quality management that good lab practices recommend Hologic validates stainers stain protocols and mounting medium clearing agent combinations for slides intended for imaging with the ThinPrep Imaging System 114 ThinPrep Stain User s Manual COVERSLIPPING Coverslipping PURPOSE The mounting medium creates a permanent bond between the slide and the coverslip It protects the cellular material from air drying and shrinkage and acts as a seal to deter fading of the stain over time COVERSLIPPING REQUIREMENTS It is recommended that the bath in which the slides are held prior to coverslipping has the clearing agent filled to a level above the cell spot but below the frosted area of the slide This will reduce the amount of mounting media overspreading the label area after the coverslip is applied Various mounting media have been evaluated by Hologic for use with ThinPrep slides These are found in the table on page 1 17 Glass coverslips 1 thickness 24 mm x 50 mm are recommended Note that ThinPrep slides for use on the ThinPrep Imaging System require coverslips long enough to cover the area including the fiducial marks that are printed on the microscope slide Hologic approved film or tape coverslip material for use with automated coverslipping instrumentation is also acceptable with the
31. solution to ensure it is free of xylene prior to recycling Otherwise the quality of the output alcohol will be compromised Testing for Xylene Contaminant in Alcohol 1 Mixthe contents of the alcohol collection container thoroughly 2 pipette 20 mL into a clean dry vessel e g graduated cylinder 3 Add 20 mL of tap water cover and invert several times to mix a If any cloudiness appears within the solution occurs almost immediately xylene is present Then DISCARD do not recycle all the container s contents according to ThinPrep Stain User s Manual STAINING the laboratory s guidelines for hazardous chemical waste disposal Clean the container using fresh absolute alcohol and dry Begin new alcohol collection b If no cloudiness is present no xylene is present Proceed to recycle the collected alcohol Test the output alcohol 1 For xylene contaminant See method above 2 Usea hydrometer to determine concentration 3 Adjust with water or absolute alcohol to 95 70 or 50 as needed Special note for CBG Biotech units which are used to recycle both alcohol and xylene e When alcohol recycling is to follow xylene recycling assure the System flush is properly performed to prevent contaminating alcohol with residual xylene Refer to the CBG Biotech Installation and Operating Instructions manual for details After a System flush test output from the first recycled alcohol run to en
32. sure xylene is not present Caution Failure to follow the recommendations for fractional distillation in this section may result in recycled product which is not acceptable for use with the ThinPrep Imaging System Distillation Systems Xylene Observe each manufacturer s setup maintenance and operation recommendations Ensure the recycled xylene quality is suitable for use in ThinPrep staining protocols Contact the respective manufacturer for their recommended laboratory test procedure Periodically have the output product independently tested i e annually at minimum to confirm proper function of the recycling system Contact the manufacturer regarding this service e Only xylenes from ThinPrep Stain protocols can be distilled together Xylenes from non ThinPrep stain setups must not be co mingled for recycling with ThinPrep xylenes ThinPrep Stain User s Manual 1 9 STAINING Caution Failure to follow all recommendations for fractional distillation in this section may result in recycled product which is not acceptable for use with the ThinPrep Imaging System 110 ThinPrep Stain User s Manual WARNING Poisonous Substances Flammable Liquids Refer to the SDS for more information PROCEDURES FOR STAINING Procedures For Staining PROCEDURE NOTES FOR AUTOMATED STAINING The following information is to be used in conjunction with the applicable staining protocol Bath solution heights should comple
33. tely cover the slides at full immersion Add fresh reagent as needed to maintain bath heights It is recommended that the final clearing agent bath be filled to a level above the cell spot but below the frosted area of the slide This will reduce the amount of mounting media overspreading the label area after the coverslip is applied Staining baths should be covered when not in use to minimize evaporation of solutions and oxidative effects on the stains Change all solutions when the number of slides stained equals 1 slide for every 1 mL of bath volume or once a week whichever comes first For example if your bath volume is 450 mL then change the bath after 450 slides or once a week whichever comes first Note A list of the staining protocols including stain station maps for automated stainers is located at the Protocols tab of this manual The stainers listed are the ones that have been validated for use with the ThinPrep Stain Solutions and ThinPrep Pap slides processed on ThinPrep Processor Recommended Stain Bath Washing Procedure 1 After emptying all stain baths separate the baths for the clearing agent Xylene ProPar etc from the others 2 Clean clearing agent baths by wiping out any debris with a paper towel and letting them air dry in a hood Do not use water in these baths 3 Wash out all of the other baths with a brush and water 4 For baths that do not come clean with a brush and water Usea solution of
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