InviMag Blood DNA Mini Kit/ IG User manual
Contents
1. Unload button B 3 The plate can be reloaded by pressing on the offered plate in C For a successful run the InviGenius needs one free lane in the incubator position seven free lanes in the working position and one free lane in the eluate position Please make sure that the depicted lanes on the monitor are consistent with the real lanes in the corresponding positions To avoid contaminations we strongly recommend to not wash reuse disposed plates 19 InviMag Blood DNA Mini Kit IG 0515 Waste management Please make sure that the waste tray is capacity is sufficient for your planned assay If the capacity is not sufficient empty the solid waste Note The waste is potentially infectious Waste management Waste capacity 336 Dropshaft status In place Fill level 0 disposables 0 Number of wasted 20 disposable sheaths 0 Number of wasted disposable fips 0 Microplates Back Batch definition loading Status Ready m User developer Figure 11 Waste management screen If the waste tray was emptied exchanged please use the Empty solid waste button A Batch definition Please select the appropriate assay and check the samples you want to process in this run You can select the assays by using the two arrow buttons A By default all loaded samples are selected Batch definition Assay descriptions suitable to loaded reagents in Samples SA2. Remaining time 00 00 00 Duration UV decontamination status There was no UV decontamination completed N Status Ready mem mi User service Figure 17 UV decontamination screen When the decontamination is finished go back to the main menu by using the Back button The device is now decontaminated and can be either switched off or used for sample processing We recommend decontaminating the instrument daily 23 InviMag Blood DNA Mini Kit IG 0515 Appendix Example data The below shown gDNA was derived with the InviMag Blood DNA Mini Kit IG 200 ul whole blood stored at 20 C The isolated DNA was analyzed by agarose gel electrophoresis and the DNA was suitable for real time PCR amplification which is demonstrated by the successful amplification of the GAPDH fragment in the samples Figure A Figure B Amplification Plot 10000 ARn 1000 1 24 20 28 E 2 34 E 3 a t 18 xa 2 Cycle The average purity OD 2so OD so of all isolated samples was about 1 7 0 1 and represents pure DNA The overall yield of all samples was about 6 0 4 ug The elution volume was 200 ul The photometric measurement was performed on a Nanodrop 1000 instrument using 2ul sample whereas 10 ul of the respective eluates were used for gel electrophoresis The performed PCR proved that no inhibition occurred All samples were fully functional and showed a consistent amplification picture The PCR was performed on a
3. stratecee molecular es User manual InviMag Blood DNA Mini Kit IG for useonthe InviGenius STRATEC Molecular GmbH for automated purification of DNA from 200 ul whole blood samples with magnetic beads 2431120100 past STRATEC Molecular GmbH D 13125 Berlin v Instruction for InviMag Blood DNA Mini Kit IG The InviMag Blood DNA Mini Kit IG combines the advantages of the innovative Invisorb technology with easy handling of magnetic particles in combination with the InviGenius robotic platform for a very efficient and reliable isolation of nucleic acids with a high purity The InviMag Blood DNA Mini Kit IG is the ideal tool for a walk away automated isolation and purification of highly pure genomic and mitochondrial DNA from 200 ul whole blood samples EDTA or citrate stabilized but not heparin The kit is designed for the InviGenius workstation The interplay of the DNA extraction and purification chemistry provided by the InviMag Blood DNA Mini Kit IG was intensely tested and validated The DNA binding magnetic particles are characterized by a high surface area uniform size distribution good suspension stability and are highly suitable for high throughput processing Due to the high purity the isolated DNA is ready to use in a broad panel of downstream applications or can be stored at 20 C for subsequent use The InviGenius is a compact walk away DNA RNA extraction platform with full in process
4. For long term storage we recommend freezing the samples at 20 C or 80 C Avoid multiple thawing and freezing cycles of the sample s before isolating the DNA Various different blood collection tubes e g Sarstedt Greiner and anticoagulants except heparin can be used to collect blood samples for the InviMag Blood DNA Mini Kit IG procedure STRATEC Molecular will not take responsibility if other sample types than described above are used or if the sample preparation advices are modified Principle and procedure The InviMag Blood DNA Mini Kit IG procedure comprises following steps lysis of blood cells and protein digestion binding the genomic DNA to the magnetic beads washing of the bead bound DNA and elimination of ethanol elution of genomic DNA O O OO After lysis the DNA binds to the magnetic beads whereas contaminations and enzyme inhibitors are efficiently removed during the following three washing steps Finally highly purified DNA is eluted in Elution Buffer Lysis Samples are lysed at chaotropic conditions in a 2 0 ml Deep Well Plate at elevated temperatures in the presence of Lysis Buffer HLT and Proteinase K Binding of the genomic DNA After addition of Binding Solution and MAP Solution B magnetic beads the DNA is bound to the surface of the beads Removing residual contaminants Contaminants are efficiently removed using Wash Buffer HLT prepared by the system and Wash Buffer II while the DNA remains bo
5. amount whereas a detailed inventory checks helps to eliminate human errors data storage backup and archiving Due to the high purity the eluted total genomic and mitochondrial DNA is ready to use for a broad panel of downstream applications PCR qPCR oequencing Restriction Enzyme Digestion HLA Typing o Southern Blot O O O O For the isolation of DNA from a single blood sample STRATEC Molecular offers the Invisorb Spin Blood Mini Kit or for 8 96 samples the Invisorb DNA Blood Mini HTS 96 Kits for use on a centrifuge or other robotic workstations For further information please contact phone 49 0 30 9489 2901 2910 in Germany and from foreign countries phone 49 0 30 9489 2907 2903 or ask your local distributor The PCR process is covered by US Patents 4 683 195 and 4 683 202 and foreign equivalents owned by Hoffmann La Roche AG T InviMag Blood DNA Mini Kit IG 0515 Sampling and storage of starting material For reproducible and high yields appropriate sample storage is essential Yields may be varying from sample to sample depending on factors such as health of the donor sample age kind of sample transport and storage conditions Blood Best results are obtained using fresh whole blood samples Mammalian blood samples stabilized with EDTA or citrate but not heparin can be stored at room temperature 18 25 C for 2 3 hours For short time storage up to 24 h samples should be stored at 4 8 C
6. 4 Loading screen of the InviGenius software Select Samples to proceed with the sample loading screen Samples loading SAMPLE 1 edited SAMPLE 2 edited SAMPLE 3 edited SAMPLE 4 edited SAMPLE 5 edited SAMPLE 6 edited SAMPLE 7 edited SAMPLE 8 edited SAMPLE 9 edited SAMPLE 10 edited SAMPLE 11 edited SAMPLE 12 edited DELeOOOO Q Q2OO000 Reagent loading Status Ready m User service Figure 5 Sample loading screen of the InviGenius software Insert the samples into the sample rack and decap the tubes If available the primary tubes should be used directly as sample tubes If the samples are not provided in primary tubes please prepare the sample rack with primary tubes that are prefilled with samples from which the DNA shall be extracted Sample tubes are not provided with the kit and can be ordered at e g Sarstedt order no 55 476 5 ml tubes 75x12 mm PS or see recommendation at page 10 chapter Reagents and equipment to be supplied by user For each reaction a sample volume of 200 ul is processed However it is recommended that the provided sample volume should be at least 400 ul to ensure stable processing Ensure that only the first 12 positions of the sample rack can be processed due to the limited number of wells per row of the plastic For correct identification of the sample tubes the unique bar codes must face to the bar code scanner located at the right side of the loading bay After inse
7. Elution Plate E Sheath Box Microtube Cap Sealing Foils Incubator Stripe Foils Initial steps 8 x 12 preps reagent sufficient for 2431120100 50 ml 8 runs for 2 7 ml working solution 1 tube per 8 runs 2 x 2 6 ml 1 tube per 4 runs empty bottle final volume 80 ml 8 runs 2 x 36 ml 4 runs per bottle final volume 2 x 120 ml P 80 ml 8 runs per bottle 1 8 runs per plate 4 2 runs per plate 1 8 runs per plate 1 2 racks 48 sheaths 4 runs per rack 8 4 Add 80 ml of 99 7 Isopropanol to the bottle Binding Solution Resuspend lyophilized Proteinase K by addition of 2 7 ml of distilled water 1x PBS to the tube mix thoroughly and store unused Proteinase K at 20 C Add 84 ml of 96 100 ethanol to each bottle Wash Buffer Il Mix thoroughly and always keep the bottle firmly closed 3 InviMag Blood DNA Mini Kit IG 0515 Symbols poet Manufacturer Lor Lot number Catalogue number Expiry date Consult operating instructions Temperature limitation Do not reuse o HKA E Storage All buffers and kit contents of the InviMag Blood Mini Kit IG except dissolved Proteinase K and MAP Solution B magnetic beads should be stored at room temperature and are stable for at least 12 months at this condition Proteinase K Dissolved and unused Proteinase K must be stored at 20 C MAP Solution B The magnetic bead solution should be stored at 4 8 C Wash Buffer II Wash Buffer II charged with et
8. If a new kit is used add ethanol to Wash Buffer II and isopropanol to the Binding Solution Before starting a new run dissolve the Proteinase K tube by adding 2 7 ml distilled water Preparing the system Switch on the InviGenius system using the power switch located on the right side of the back part of the instrument The InviGenius software will be automatically loaded after the system has booted up Please keep the door of the InviGenius system closed during system initialization After successful initialization of the InviGenius system a login screen appears Figure 2 Login use the provided user name and password Login User identifier ucc Password i Status Ready sss EMEN Ser Figure 2 Login screen of the InviGenius software After logging in the main screen of the InviGenius software appears Figure 3 Select Loading to start loading the system and prepare for starting a run Select Processing to define and run an assay if the system has been already loaded Main Menu Loading Processing Shutdown InviGenius Status Ready m ser developer Figure 3 Main menu of the InviGenius software 14 InviMag Blood DNA Mini Kit IG 0515 Sample Loading After selecting Loading the main loading screen appears Loading Samples Disposable tips Disposable Assay Waste management Status Ready Ott 307 mamum User developer Back Figure
9. PCR process is covered by US Patents 4 683 195 and 4 683 202 and foreign equivalents owned by Hoffmann La Roche AG 2015 STRATEC Molecular all rights reserved 1 InviMag Blood DNA Mini Kit IG 0515 Contents Kit contents of InviMag Blood DNA Mini Kit IG 3 Symbols 4 Storage 4 Quality control 4 Intended use 9 Product use limitation 5 Safety information 6 Product characteristics of the InviMag Blood DNA Mini Kit IG 7 Sampling and storage of starting material 8 Principle and procedure 8 Yield and quality of genomic DNA 9 Important notes 9 Preparing reagents and buffers 9 Reagents and equipment to be supplied by user 9 Important indications 10 Scheme of the InviMag Blood DNA Mini Kit IG 11 Preparing the samples for processing on the InviGenius 12 General overview of the InviGenius system 13 Preparing and loading of the InviGenius amp system 14 Daily Maintenance UV sterilization 20 Appendix 24 Example data 24 General notes on handling DNA 24 Troubleshooting 25 Ordering information 26 2 InviMag Blood DNA Mini Kit IG 0515 Kit contents of InviMag Blood DNA Mini Kit IG Store the MAP Solution B at 2 8 C store dissolved Proteinase K at 20 C Store all other kit components at room temperature RT Component Catalogue No Lysis Buffer HLT Proteinase K MAP Solution B Binding Solution fill with 99 7 Isopropanol Wash Buffer II Elution Buffer M Incubation Plate A Working Plate A
10. Status Ready User developer Figure 8 Disposable tip loading screen There are three tip rack positions on the InviGenius system Fig 8 A1 A3 Remaining tip numbers are shown in field B Tip numbers can be changed by pressing the number field directly Empty tip racks can be unloaded and reloaded by 1 Pressing the Loading Position directly The software will focus this loading position on the main screen 2 Pressing the Unload Button C 3 The loading position can be refilled with a new tip rack by pressing on the corresponding tip rack on D Each position can be filled either with 50 ul or 1100 ul filter tips However for the Blood Mini assays only 1100 ul filtered tips can be used and will be selectable Attention It is very important to allocate the type of tips correctly in the software that have been loaded into the instrument In case of false tip allocation overfilling of the tip will irreparably destroy the pipettor head All protocols should be used in combination with filter tips to ensure efficient prevention of sample or reagent cross contaminations STRATEC Molecular will give no guarantee or responsibility if contaminations occur due to the use of non filtered tips Note Disposable tips are not supplied within the kit We recommend the use of validated conductive tips which can be ordered at STRATEC Molecular STRATEC Molecular offers 50 ul conductive tips 10x 96 pieces order no 5011120100 an
11. StepOnePlus cycler from Applied Biosystems in which 2ul sample were analyzed The C values derived by the PCR illustrated in Fig B demonstrate that all Cz values are in the same range with a very low standard deviation General notes on handling DNA Nature of DNA The length and delicate physical nature of DNA requires careful handling to avoid damage due to shearing and enzymatic degradation Other conditions that affect the integrity and stability of DNA include acidic and alkaline environments high temperature and UV irradiation Careful isolation and handling of high molecular weight DNA is necessary to ensure its functionality in various downstream applications Damaged DNA could perform poorly in applications such as genomic Southern blotting long template PCR and construction of cosmid libraries Handling fresh and stored material before the extraction of DNA For the isolation of genomic DNA use either fresh samples or samples that have been quickly frozen in liquid nitrogen and stored at 70 C This procedure minimizes degradation of crude DNA by limiting the activity of endogenous nucleases Storage of DNA Store genomic DNA at 20 C However be careful because if the DNA is exposed to repeated freezing and thawing cycles this may cause shearing of the material 24 InviMag Blood DNA Mini Kit IG 0515 Troubleshooting Problem pipetting distribution errors low concentration of extracted DNA degraded DNA no
12. Buffer HLT and 20 ul Proteinase K Incubation at 56 C for 10 min is performed 260 ul Binding Solution follow preparing instructions and 40 ul MAP Solution B are added to the lysate DNA binds to magnetic particles magnetic separation washing of the particle fixed genomic DNA 1 x 900 ul Wash Buffer HLT 1 x 950 ul Wash Buffer Il 1 x 700 ul Wash Buffer Il magnetic separation elution of genomic DNA in 100 ul 200 ul Elution Buffer M magnetic separation removal of beads transfer of eluate to the eluate plate rc ex cd 11 InviMag Blood DNA Mini Kit IG 0515 Preparing the samples for processing on the InviGenius Please read the instructions carefully and conduct the prepared procedure Important Note The protocol has been optimized for the isolation of genomic DNA from up to 200 ul of whole Blood EDTA Citrate stabilized To prevent possible distribution errors it is highly recommend to provide at least 400 ul of sample in total to ensure stable processing Attention Before you start dissolve one vial of Proteinase K with 2 7 ml of distilled water PBS Reconstitute the Wash Buffer ll Be aware that the Binding Solution has to be prepared see instruction at page 9 1 Extraction of genomic DNA from whole blood This type of sample can be processed directly without any preparations Ensure to supply at least 400 ul sample volume DBLD E100 S200 DNA from Blood E Elution volume 100 ul S Sample vol
13. H Robert Rossle Str 10 13125 Berlin Germany Phone 49 30 94 89 29 01 Fax 49 30 94 89 29 09 E mail info berlin stratec com www stratec com 1G3a01 IG 05 2015
14. InviGenius software Insert all provided reagents into the provided reagent rack of the InviGenius system Verify that the bar code labels face to the right side of the loading bay and decap the bottles and tubes The order of the inserted reagents is not crucial because the type and position of a reagent is identified by the unique bar code However the possible loading positions are limited by the size of the used bottles After rack insertion the loading status of the reagents will be shown In case of unsuccessful reagent allocation remove the rack check the bar code orientation and try again slowly Assay Selection Assay selection Assay description suitable to loaded reagents DBLD E100 200 DBLD E200 200 No Assay Selected Version 1 Reagent loading Disp tip loading Status Ready m ser service Figure 7 Assay Selection screen of the InviGenius software 16 InviMag Blood DNA Mini Kit IG 0515 Select the appropriate protocol and proceed with disposable tip loading If no assay file is visible one or more reagents were not recognized correctly during the reagent loading procedure go back to Reagent Loading and repeat the loading process Disposable Tip Loading Disposable tips loading Load Pos 1 Tip Load Position 1 99999999990 SSCCCCCCCCCSD I oo 999999999990 B 22222222221 0 Tip 1100 333553255225 ul filter Assay selection Disp sheaths loading
15. MPLE SAMPLE2 SAMPLE3 SAMPLE4 SAMPLE5 DBLD E200 200 DBLD E100 S200 No Assay Selected SAMPLES SAMPLE7 emove SAMPLE8 from batch SAMPLES SAMPLE10 SAMPLE11 SAMPLE12 Version 30 01 00312 Aliquots Volume B si 200 v management Status Ready mmm ee User developer Figure 12 Batch definition screen Please select the desired assay and recheck the allocated samples that should be processed in this run It is possible to switch between the offered assays by using the two arrow buttons A By default all loaded samples are selected to be processed in this run If samples have to be excluded from the batch exclude them by selecting the corresponding sample and clicking on the Hemove from batch button B 20 InviMag Blood DNA Mini Kit IG 0515 Batch checking This screen shows a summary of all checked disposables samples and reagents in one informational screen Please make sure that all required components are loaded correctly In case of any error the problem will be highlighted in by a red font If no errors during the loading steps occurred proceed by pressing the button Batch processing To solve any error click on the red highlighted field and follow the instructions printed on the instrument screen Batch checking Selected assay DBLD_E100_S200 E ti 1 Od0000000000 n GBHBHHHBHBHHE HOC a Batch processing eooooooo
16. STS TRAINED IN MOLECULAR BIOLOGICAL TECHNIQUES It is designed to be used with any downstream application employing enzymatic amplification or other enzymatic modifications followed by signal detection or amplification Any diagnostic results generated by using the sample preparation procedure in conjunction with any downstream diagnostic assay should be interpreted with regard to other clinical or laboratory findings To minimize irregularities in diagnostic results adequate controls for downstream applications should be used The kit is in compliance with EU Directive 98 79 EC on in vitro medical devices But it is not for in vitro diagnostic use in countries where the EU Directive 98 79 EC on in vitro medical devices is not recognized Product use limitation The kit is validated for the use of whole blood samples only The isolation of DNA from other sources like stool tissue bacteria fungi or virus was neither tested nor validated The included chemicals are only useable once Differing of starting material may lead to inoperability Therefore neither a warranty nor guarantee in this case will be given implied or expressed The user is responsible to validate the performance of the STRATEC Molecular product for any particular use STRATEC Molecular does not provide validation of performance characteristics of the product with respect to specific applications STRATEC Molecular products may be used in clinical diagnostic laboratory sy
17. assay selectable eluted nucleic acids are brownish colored Probable cause pipetting of Proteinase K failed samples transfer failed incomplete reagent buffer transfer failed incomplete blood components seitled No too much ethanol added to Wash Buffer Il incorrect storage of starting material old material combination of reagents from different kits missing required reagent Residual magnetic particles are left in eluate 25 Comments and suggestions ensure that the lyophilized Proteinase K is dissolved with the appropriate volume of distilled water before usage the sample tube must contain at least 400 ul sample ensure that the supplied Wash Buffer Il Binding Solution are filled up properly with either ethanol or isopropanol do not reuse bottles more often than described in Tab 1 because they will be rejected by the system in case of large sample volumes gt gt 1 ml carefully premix the sample tube before inserting it into the sample rack ensure that the Wash Buffer II have been filled up properly with ethanol as indicated in Tab 1 ensure that the storage of starting material is correct avoid multiple freezing and thawing cycles of the material ensure that the starting material is fresh or stored at appropriate conditions for long time storage at 20 C avoid multiple thawing and freezing cycles of the material old material may contain degraded DNA ensure th
18. at only and all reagents belonging to one kit type are used a combination of reagents belonging to different kit types is not supported centrifuge the eluate plate at full speed for 1 min and transfer supernatant to a new plate tube InviMag Blood DNA Mini Kit IG 0515 Ordering information Product InviMag Blood DNA Mini Kit IG Related products InviMag Blood DNA Giga Kit IG Invisorb Spin Blood Mini Kit Invisorb Spin Blood Mini Kit Invisorb Spin Blood Midi Kit Invisorb Spin Blood Midi Kit InviGenius and consumables InviGenius Starting Box I IG Sheath Box Conductive filter tips 1 ml 2 x 2 rack pack 384 pieces 5 Waste Trays 120 sample tubes Sheath Bundle Sheaths Conductive filter tips 1 ml Package size 8 x 12 preps 8 x 12 preps 50 preparations 250 preparations 50 preparations 250 preparations 1 unit 1 box 10 x 48 pieces 1000 pieces 10 x 96 pieces Waste tray IG 25 pieces Possible suppliers for Isopropanol Carl Roth Applichem 2 Propanol 2 Propanol f r die Molekularbiologie Rotipuran gt 99 7 p a ACS ISO Order no A3928 Order no 6752 26 InviMag Blood DNA Mini Kit IG Catalogue No 2431120100 2431320400 1031100200 1031100300 1031110300 1031110500 9011100000 2400110100 5011100300 5011100200 5011100400 5011100100 Sigma 2 Propanol Order no 59304 1L F 0515 Stratecee molecular STRATEC Molecular Gmb
19. aterials Contamination of the waste with residual infectious materials is unlikely but cannot be excluded completely Therefore the waste has to be considered infectious and should be handled and discarded accordingly to local safety regulations Subsequently European Community risk and safety phrases for the components of the InviMag Universal Kit IG to which they apply are listed Lysis Buffer HLT PKC Tube Proteinase K Carrier RNA contains guanidine hydrochloride H302 315 319 P280 505 351 338 H315 319 334 335 P280 305 351 338 310 405 H315 Causes skin irritation H319 Causes serious eye irritation H334 May cause allergy or asthma symptoms or breathing difficulties if inhaled H335 May cause respiratory irritation P280 Wear protective gloves protective clothing eye protection face protection P305 P351 P338 IF IN EYES Rinse cautiously with water for several minutes If present remove contact lenses and continue rinsing P310 Immediately call a POISON CENTER or doctor physician P405 Store locked up P210 Keep away from heat sparks open flames hot surfaces No smoking Emergency medical information can be obtained 24 hours a day from infotrac outside of USA 1 352 323 3500 inside of USA 1 800 535 5053 6 InviMag Blood DNA Mini Kit IG 0515 Product characteristics of the InviMag Blood DNA Mini Kit IG The InviMag Blood DNA Mini Kit IG is the ideal tool for an efficie
20. control including the following modules such as pipettor heatable incubator barcode reader magnetic separation head integrated PC and touch screen barcode labelled sample rack for primary tubes and a barcode labelled reagent rack which helps to deliver premium quality nucleic acid for routine laboratories The workstation eliminates human errors standardizes the extraction process and offers an integrated solution for data storage backup and archiving The unique bar codes for samples and reagents avoid unwanted transpositions The kit is not validated for the isolation of genomic DNA from cultured or isolated cells from tissue blood cards dried blood stains or urine The application of the kit for isolation and purification of viral DNA has not been evaluated C Compliance with EU Directive 98 79 EC on in vitro medical devices Not for in vitro diagnostic use in countries where the EU Directive 98 79 EC on in vitro medical devices is not recognized Trademarks InviMag Invisorb and InviGenius Registered marks trademarks etc used in this document even when not specifically marked as such are not to be considered unprotected by law The Invisorb technology is covered by patents and patent applications US 6 110363 US 6 043 354 US 6 037 465 EP 0880535 WO 9728171 WO 9534569 EP 0765335 DE 19506887 DE 10041825 2 WO 0034463 InviMag Invisorb and InviGenius are registered trademarks of STRATEC Biomedical AG The
21. d 1100 ul conductive tips 10x 96 pieces order no 5011120200 Be sure that conductive tips are used otherwise the tip detection unit installed in the pipetting unit will reject the tips and no run will be possible 17 InviMag Blood DNA Mini Kit IG 0515 Disposable Sheaths Loading The sheaths are used as protection devices for the magnetic rods The sheaths are picked up automatically during the run and provided in the kits Disposable sheaths loading Sheath Load Position amp eee 4 amp amp amp e 9 amp e 9 9 e 9 anes amp amp amp 99e LOKORKOKORORKORORKORKORKORORKO Sheath Type Microplates Status Ready OI User developer Figure 9 Disposable sheaths loading screen The loading procedure of the disposable sheaths works analogous to the disposable tip loading screen For a run always 12 disposable sheaths one row in the sheaths rack are used regardless of the processed sample numbers This is done to assure that the rods are always protected against contaminations In general the number of sheaths supplied within the kit is sufficient for the amount of runs printed on the kit package If you are lacking sheaths they can be ordered separately at STRATEC Molecular 100 pieces bulk order no 5011120300 or 10 x 48 pieces order no 5011120400 Comparable to the disposable tips loading it is possible to define the number of rows left in the tip rack by pressing on the display
22. ect quantities If there are any unconformities notify STRATEC Molecular in writing with immediate effect upon inspection thereof If buffer bottles are damaged contact the STRATEC Molecular Technical Services or your local distributor In case of liquid spilage refer to Safety Information s see page 6 Do not use damaged kit components because their use may lead to poor kit performance o When working with chemicals always wear a suitable lab coat disposable gloves and protective goggles Discard contaminated gloves immediately Never combine components of different kits Avoid microbial contamination of the kit reagents To minimize the risk of infections from potentially infectious material we recommend working under laminar air flow until the samples are prepared o This kit should only be used by trained personnel O O O Preparing reagents and buffers Before starting a run equilibrate all reagents at room temperature Where necessary gently mix and redissolve any precipitates by incubating at 30 C Swirl gently to avoid foaming Lysis Buffer HLT and Elution Buffer M are ready to use 8 x 12 DNA extractions Add 80 ml of 99 7 Isopropanol to the bottle Binding Solution Resuspend the lyophilized Proteinase K tube by addition of 2 7 ml of distilled water or 1x PBS Mix thoroughly and store diluted and used Proteinase K at 20 C Add 84 ml of 96 100 ethanol to the bottle Wash Buffer II Mix thoroughly and always kee
23. ed number area Make sure that the disposable sheaths are loaded and displayed consistent to the manually loaded sheaths in the rack to ensure correct sheaths pick up Don t remove single disposable sheaths within a row of the sheaths rack if less than 12 samples are processed within one run because there is a sheaths detection sensor installed in the device If less than 12 sheaths picked up by the instrument a warning will be displayed and all picked up sheath will be discarded into the waste before a next row of sheaths will be picked up for testing To avoid contaminations we strongly recommend to not wash reuse any disposed sheaths 18 InviMag Blood DNA Mini Kit IG 0515 Plate Loading Analogous to the previous loading screens the incubation working and elution plate are loaded within the plate loading screen Figure 10 Eluate Position Free lanes 8 6729 woz naaa aun goaagaogaagaadgadrurc coooooooooooo nooooooo Elution Plate E Disp sheaths loading Waste management User service Status Ready Figure 10 Plate loading screen In general the Incubation Plate A and Working Plate A identical are used at the incubator and working position whereas at the eluate position the Elution Plate E is used Used plates can be unloaded and reloaded by 1 Pressing the plate position directly A The software will focus at the plate position on the main screen 2 Pressing the
24. hanol should be stored at room temperature and must be appropriately sealed if not used If there are any precipitates visible within the provided solution solve them by carefully warming up to 30 C Room temperature RT is defined as range from 15 30 C Quality control and product warranty STRATEC Molecular warrants the correct function of the InviMag Blood Mini Kit IG for applications as described in this manual Purchasers must determine the suitability of the product for its particular use Should any product fail to perform the applications as described in the manual STRATEC Molecular will check the lot and if lot connected problem is investigated STRATEC Molecular will replace the product free of charge STRATEC Molecular reserves the right to change alter or modify any product to enhance its performance and design at any time In accordance with STRATEC Molecular s ISO 9001 2000 and ISO EN 13485 certified Quality Management System the performance of all components of the InviMag Blood Mini Kit IG have been tested separately against predetermined specifications routinely on lot to lot to ensure consistent product quality In case of any question or problem regarding any aspect of InviMag Blood Mini Kit IG or other STRATEC Molecular products please do not hesitate to contact us A copy of STRATEC Molecular s terms and conditions can be obtained upon request or are presented at the STRATEC Molecular webpage For technical
25. he InviGenius system is equipped with an internal UV lamp 254 nm wavelength that should be used daily either at the end of the working day or in the morning before a run is started The suggested decontamination time is about 20 min To start the UV decontamination go to the main menu of the InviGenius software and select Maintenance Main Menu Loading Data management Maintenance Administration InviGenius Status Ready mm User developer Processing Shutdown Figure 15 Main screen of the InviGenius software When the sub item Maintenance is opened select UV decontamination UV Periodical decontamination maintenance Cleanu P Diagnostics procedures K Maintenance Sisari Recove drop catcher Y Status Ready ma User service Figure 16 Maintenance screen of the InviGenius software 22 InviMag Blood DNA Mini Kit IG 0515 In the UV decontamination menu adjust the exposure time A and finally press the Start button B During the decontamination process the instrument door will be locked to prevent any UV radiation release in the lab Warning UV radiation is harmful It causes serious burns of the skin and leads to irreparable damage of the eyes and skin Ensure that no lab personnel is submitted to direct UV light Do not try to force open the instrument door during the decontamination process UV decontamination A 00 15 00 HH MM SS
26. nt and fully automated DNA extraction and purification from fresh or frozen whole blood samples using magnetic beads in combination with the InviGenius robotic platform preparation 1 200 ul fresh or frozen whole blood 2 10 ug depends on the about 90 min per blood sample storage run 12 samples and source The DNA isolation process is based on the interaction of nucleic acids with silica coated magnetic particles at adapted buffer conditions The InviGenius instrument will automatically perform all steps of sample and reagent distribution The DNA purification procedure is performed without any user intervention except the initial loading of the system thus allowing safe handling of potentially infectious samples Sample cross contaminations and reagent cross overs are effectively eliminated by the automated purification process The use of unique bar codes for samples and reagents avoids unwanted transpositions The InviGenius instrument uses magnetic rods to transport the DNA binding magnetic particles through the various purification phases such as lysis binding washing and elution The volume of buffers and other liquids necessary for DNA isolation is reduced to a minimum Eliminating the direct liquid handling and increasing the automation level results in a fast reliable and robust technique The process on InviGenius is total in process control including e g completely reagent tracking on the bottle shelf live
27. oooooQ User service ooo Batch definition Status Ready Figure 13 Batch definition screen Batch processing After closing the system door the assay can be started by pressing the Start Button A The door will be locked during the run and the system will start with sample processing The door will only be unlocked after a run has been successfully finished or if an error occurs that requires user interaction Do not try to force open the door during a run This will cause an abort of the run Batch processing Batch identifier Assay description Actual step s Remaining time 00 00 00 Process state Idle Events Status Ready i User service Figure 14 Batch processing screen At the end of the process the nucleic acid containing elates are located in the appropriate eluate position and can be used for any further downstream application Note The complete process will take approximately 90 minutes 21 InviMag Blood DNA Mini Kit IG 0515 After a run After a run is completed and no additional run shall be started unload all plates and reagents and store them according to GLP guidelines Please keep in mind that the plates could contain infectious material As with all medical clinical and diagnostically equipment all waste liquids tips sheaths and plates should be treated as potentially dangerous bio hazard waste Daily maintenance UV decontamination T
28. p the unused bottle firmly closed Reagents and equipment to be supplied by user ddH O Vortexer 96 100 ethanol Primary tubes e g see below Isopropanol o Measuring cylinder 250 ml o Conductive pipette tips see ordering information page 26 o Disposable gloves o PBS buffer O O O O 9 InviMag Blood DNA Mini Kit IG 0515 The InviMag Blood DNA Mini Kit IG is validated with 2 Propanol Rotipuran gt 99 7 p a ACS ISO Order no 6752 from Carl Roth Possible suppliers for Isopropanol Carl Roth Applichem Sigma 2 Propanol 2 Propanol fur die Molekularbiologie 2 Propanol Rotipuran gt 99 7 p a ACS ISO Order no A3928 Order no 59304 1L F Order no 6752 Possible primary tubes manufacturer cat no Venosafe 5 5 ml Ref VF O76SDK Terumo Vacuette 2 ml Ref A110500I Greiner bio one Vacuette 9 ml Ref 455036 Greiner bio one BD Vacutainer 2 7 ml Ref 363048 BD Vacutainer 6 ml Ref 367864 BD Vacutainer 10 ml Ref 367525 BD Vacutainer 5 0 ml Re Sarstedt Monovette 8 5 ml PS Tube Sarstedt 5 ml Ref 55 476 Sarstedt Monovette 4 5 ml Sarstedt Monovette 7 5 ml Sarstedt Monovette 9 0 ml Important indications 1 Minimum volume of samples in primary tubes The procedure of the InviMag Blood DNA Mini Kit IG has been optimized for the isolation of genomic DNA from up to 200 ul whole blood We advise to provide at least 400 ul blood per sample tube to prevent pipetting distrib
29. rting the sample rack in the very left lane of the loading bay an updated screen will show the identifiers read from the sample bar codes Figure 5 In case of unsuccessful sample identification remove the rack check the bar code orientation and reinsert the rack slowly It is 15 InviMag Blood DNA Mini Kit IG 0515 also possible to rename the samples by selecting the corresponding sample by using the arrow fields followed by the Edit button After a certain time about 5 min the bar code scanner is inactivated In that case the user has to restart the scanner with the START SCANNER button if the loading procedure is not finished After successful loading of the samples proceed with reagent loading by selecting Reagent loading on the bottom right hand side of this screen Reagent Loading The reagent loading process is analogous to the sample loading procedure Reagents loading Barcode Exp date Unknown Unknown PA124045130701771506 Lysis B HLT PA 2015 06 30 PA706026340108801506 Proteinase K PA 2015 06 30 PA218068100001491506 Binding Sol PA 2015 06 30 PA618025300202521506 MAP Sol B PA 2015 06 30 PA306108140102791506 Wash B II PA 2015 06 30 Unknown Unknown PA410068130602971506 Elution B M PA 2015 06 30 iA ACACA ACACACA A ACACACI N Unknown Unknown Sample loading Status Ready memm User Service _ e L Figure 6 Reagent loading screen of the
30. stems conditioned upon the complete diagnostic system of the laboratory after the laboratory has been validated pursuant to CLIA 88 regulations in the U S or equivalents in other countries All products sold by STRATEC Molecular are subject to extensive quality control procedures according to ISO 9001 2000 and ISO EN 13485 and are warranted to perform as described herein Any problems incidents or defects shall be reported to STRATEC Molecular immediately upon detection thereof The chemicals and the plastics are for laboratory use only They should be stored in the laboratory and must not be used for other purposes than intended The product with its contents is not suitable for consumption 9 InviMag Blood DNA Mini Kit IG 0515 Safety information When and while working with chemicals always wear a suitable lab coat disposable gloves and protective goggles Avoid skin contact Adhere to the legal requirements for working with biological material For more information please consult the appropriate material safety data sheets MSDS These are available online in convenient and compact PDF format at www stratec com for each STRATEC Molecular Product and its components If buffer bottles are damaged or leaking WEAR GLOVES AND PROTECTIVE GOGGLES when discarding the bottles in order to avoid any injuries STRATEC Molecular has not tested the waste generated by the InviMag Universal Kit IG procedures for residual infectious m
31. support or further information please contact from Germany 49 0 30 9489 2901 2910 from abroad 49 0 30 9489 2907 or contact your local distributor 4 InviMag Blood DNA Mini Kit IG 0515 Intended use The InviMag Blood DNA Mini Kit IG is designed for fully automated extraction and purification of genomic DNA from up to 200 ul whole blood ranging from 1 12 samples using magnetic beads and the InviGenius instrument The nucleic acid isolation protocol is suitable for routinely walk away automated preparation of DNA from fresh or frozen blood samples For reproducible and high yields the appropriate sample storage Is critical see Sampling and storage of the starting material page 8 Common blood collection tubes not provided and anticoagulants EDTA and citrate but not heparin can be used to gather a set of blood samples All utilities reagents and plastics beside filtered tips necessary for preparation of DNA from blood are provided by the InviMag Blood DNA Mini Kit IG The procedure of the InviMag Blood DNA Mini Kit IG has been optimized for the isolation of genomic DNA from up to 200 ul starting material However we advise to provide a sample volume of at least 400 ul per a 12 mm tube diameter more if bigger tubes are used to prevent pipetting distribution errors The final processed sample volume is 200 ul THE PRODUCT IS INTENDED FOR USE BY PROFESSIONALS SUCH AS TECHNICIANS PHYSICIANS AND BIOLOGI
32. ume 200 ul DBLD E200 S200 DNA from Blood E Elution volume 200 ul S Sample volume 200 ul 2 Extraction of genomic DNA from swab samples Rinse each swab with 400 ul distilled water or PBS provided in a primary tube Remove the swab after sqeezing it out inside the tube wall and load it into the sample rack 3 Extraction of genomic DNA from buffy coat Maximum 250 yl buffy coat can be used Ensure that this sample is diluted with PBS and supply at least 400 ul 12 InviMag Blood DNA Mini Kit IG 0515 General overview of the InviGenius system IN J Figure 1 Frontal view of the InviGenius system Figure 1 shows the plate positions A elution position E working position and I incubation position Disposable tips are placed on position B1 B3 and disposable sheaths on F The waste tray C is located on the lower side of the InviGenius system behind the red flap The waste shaft D is completely stainless steel and easy to remove for autoclaving The loading bay is divided into sample loading bay J1 and reagent loading bay J2 The magnetic separator head MSH H is located on top of the incubator I and can reach all necessary positions The single head pipettor G starting positions are in the right front of the machine All movable parts only work when the InviGenius machine is closed and locked 13 InviMag Blood DNA Mini Kit IG 0515 Preparing and loading of the InviGenius system Preparing the reagents
33. und to the magnetic beads Elution The DNA is finally eluted in Elution Buffer M The eluted DNA is ready to use in different subsequent downstream applications like PCR amplification restriction enzymes analysis Southern hybridizations HLA typing etc The blood assays stored in the instrument memory contains two different elution volumes protocols the user can choose DBLD E100 S200 DNA from Blood E Elution 100ul S Sample 200ul DBLD E200 S200 DNA from Blood E Elution 200ul S Sample 200ul 8 InviMag Blood DNA Mini Kit IG 0515 Yield and quality of genomic DNA The amount of purified DNA derived by the InviMag Blood DNA Mini Kit IG procedure from whole blood strongly depends on the leucocytes content sample source transport storage and age Typically a volume of 200 ul of a whole blood sample from a healthy individual with an elevated white blood cell content ranging from 3x10 to 1x10 cells ml will yield at least 3 ug of genomic DNA The typical yield usually expected from the InviMag Blood DNA Mini Kit IG is in the range of 2 10 ug DNA If a whole blood sample is mixed with anticoagulant containing buffer solutions the overall leukocyte concentration decreases and the yield of the DNA extraction procedure is slightly reduced Important notes Important points before starting a protocol Immediately upon receipt inspect the product and its components as well as the package for any apparent damages and corr
34. ution errors during processing 2 Sample volume smaller than 200 pl For samples of a smaller volume than 200 ul please adjust the volume to 200 ul using either distilled water or 1x PBS 3 Elution volume The final processed sample volume is 200 ul The procedure also offers a variation of the elution volume by offering two assays with respectively 100 ul or 200 ul elution volume Typically 200 ul eluate yields in app 2 10 ug DNA The blood assays stored in the instrument memory contains two different elution volumes protocols The user can choose between DBLD E100 S200 DNA from Blood E Elution volume 100 ul S Sample volume 200 ul DBLD E200 S200 DNA from Blood E Elution volume 200 ul S Sample volume 200 ul Important DBLD E100 S200 assay should only be used if whole blood with a very low content of leukocytes will be used Using normal whole blood will very likely result in an incomplete elution due to the high concentration of leukocytes present in normal blood and will most probably lead in a slightly decreased yield compared to 200 ul elution volume Prevention of Cross contamination To comply with the demanding guidelines of in vitro Diagnostics we programmed the InviGenius to route the pipettor in such a way that possible contamination risks are minimized 10 InviMag Blood DNA Mini Kit IG 0515 Scheme of the InviMag Blood DNA Mini Kit IG 200 ul sample will be mixed with 200 ul Lysis
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