NucleoMag® Blood 3 mL
Contents
1. Genomic DNA from blood User manual NucleoMag Blood 3 mL March 2012 Rev 02 MACHEREY NAGEL MN Genomic DNA from blood Table of contents 1 Components 1 1 Kit contents 1 2 Material to be supplied by the user 4 2 Product description 5 2 1 The basic principle 5 2 2 Kit specifications 5 2 3 Elution procedures 6 3 Storage conditions and preparation of working solutions 7 4 Safety instructions 4 1 Risk and safety phrases 4 2 GHS classification 5 Protocol for the isolation of genomic DNA from 3 mL blood samples using KingFisher Flex 24 12 6 Appendix 14 6 1 Troubleshooting 14 6 3 Product use restriction warranty 16 MACHEREY NAGEL 03 2012 Rev 02 3 Genomic DNA from blood 1 Components 1 1 Kit contents NucleoMag Blood 3 mL 1x 96 preps REF 744502 1 NucleoMag B Beads 18 mL Lysis Buffer MBL1 125 mL Binding Buffer MBL2 550 mL Wash Buffer MBL3 1000 mL Wash Buffer MBL4 500 mL Elution Buffer MBL5 125 mL Proteinase K lyophilized 12x75 mg Proteinase Buffer PB 2x35 mL User manual 1 1 2 Material to be supplied by the user Magnetic separator KingFisher Flex 24 instrument Separation plates elution plates KingFisher 24 deep well plates KingFisher 24 well tip comb 80 ethanol for the washing step Elution Buffer MBL5 5 mM Tris pH 8 5 For preparation of working solutions and storage conditions see section 3 4 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from b2. 03 2012 Rev 02 NucleoMag Blood 3 mL After adding the samples add 575 uL Buffer MBL1 to each well of the two lysis plates Start isolation on King Fisher Flex 24 instrument Start method NucleoMag _Blood_3mL method is available from MN on request Insert plates as indicated on the KingFisher instrument display Method starts with a mixing step sample lysis after setting up the last plate to the instrument After mixing steps for Iysis approx 10 min the instrument will ask for addition of Buffer MBL2 and NucleoMag B Beads Addition of Binding Buffer MBL2 and NucleoMag B Beads to lysis plate 1 Add 2 3 mL Buffer MBL2 and 150 uL NucleoMag B Beads to each well of the lysis plate 1 Mix up NucleoMag B Beads before use Return lysis plate 1 to the instrument and continue Addition of Binding Buffer MBL2 to lysis plate 2 Add 2 3 mL Buffer MBL2 to each well of the lysis plate 2 Return lysis plate 2 to the instrument and continue All further steps are now processed without further user interaction Remove eluted DNA The instrument stops after the final elution step Follow the instructions on instrument display and unload the plates from the instrument Purified DNA should be centrifuged before UV measurement MACHEREY NAGEL 03 2012 Rev 02 13 Genomic DNA from blood 6 Appendix 6 1 Troubleshooting Problem Possible cause and suggestions Elution bu
3. EL products shall in any event only be used wearing adequate PROTECTIVE CLOTHING For detailed information please refer to the respective Material Safety Data Sheet of the product MACHEREY NAGEL products shall exclusively be used in an ADEQUATE TEST ENVIRONMENT MACHEREY NAGEL does not assume any responsibility for damages due to improper application of our products in other fields of application Application on the human body is STRICTLY FORBIDDEN The respective user is liable for any and all damages resulting from such application DNA RNA PROTEIN purification products of MACHEREY NAGEL are suitable for IN VITRO USES ONLY ONLY MACHEREY NAGEL products specially labeled as IVD are also suitable for IN VITRO diagnostic use Please pay attention to the package of the product IN VITRO diagnostic products are expressly marked as IVD on the packaging IF THERE IS NO IVD SIGN THE PRODUCT SHALL NOT BE SUITABLE FOR N VITRO DIAGNOSTIC USE ALL OTHER PRODUCTS NOT LABELED AS IVD ARE NOT SUITED FOR ANY CLINICAL USE INCLUDING BUT NOT LIMITED TO DIAGNOSTIC THERAPEUTIC AND OR PROGNOSTIC USE No claim or representations is intended for its use to identify any specific organism or for clinical use included but not limited to diagnostic prognostic therapeutic or blood banking It is rather in the responsibility of the user or in any case of resale of the products in the responsibility of the reseller to inspect and assure the use of the DNA
4. ESS OR IMPLIED INCLUDING WITHOUT LIMITATION AS TO THE SUITABILITY REPRODUCTIVITY DURABILITY FITNESS FOR A PARTICULAR PURPOSE OR USE MERCHANTABILITY CONDITION OR ANY OTHER MATTER WITH RESPECT TO MACHEREY NAGEL PRODUCTS In no event shall MACHEREY NAGEL be liable for claims for any other damages whether direct indirect incidental compensatory foreseeable consequential or special including but not limited to loss of use revenue or profit whether based upon warranty contract tort including negligence or strict liability arising in connection with the sale or the failure of MACHEREY NAGEL products to perform in accordance with the stated specifications This warranty is exclusive and MACHEREY NAGEL makes no other warranty expressed or implied The warranty provided herein and the data specifications and descriptions of this MACHEREY NAGEL product appearing in MACHEREY NAGEL published catalogues and product literature are MACHEREY NAGEL s sole representations concerning the product and warranty No other statements or representations written or oral by MACHEREY NAGEL s employees agent or representatives except written statements signed by a duly authorized officer of MACHEREY NAGEL are authorized they should not be relied upon by the customer and are not a part of the contract of sale or of this warranty Product claims are subject to change Therefore please contact our Technical Service Team for the most up to date info
5. RNA protein purification products of MACHEREY NAGEL for a well defined and specific application MACHEREY NAGEL shall only be responsible for the product specifications and the performance range of MN products according to the specifications of in house quality control product documentation and marketing material This MACHEREY NAGEL product is shipped with documentation stating specifications and other technical information MACHEREY NAGEL warrants to meet the stated specifications MACHEREY NAGEL s sole obligation and the customer s sole remedy is limited to replacement of products free of charge in the event products fail to perform as warranted Supplementary reference is made to the general business terms and conditions of MACHEREY NAGEL which are printed on the price list Please contact us if you wish to get an extra copy There is no warranty for and MACHEREY NAGEL is not liable for damages or defects arising in shipping and handling transport insurance for customers excluded or out of accident or improper or abnormal use of this product defects in products or 16 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from blood components not manufactured by MACHEREY NAGEL or damages resulting from such non MACHEREY NAGEL components or products MACHEREY NAGEL makes no other warranty of any kind whatsoever and SPECIFICALLY DISCLAIMS AND EXCLUDES ALL OTHER WARRANTIES OF ANY KIND OR NATURE WHATSOEVER DIRECTLY OR INDIRECTLY EXPR
6. UGEN Einige Minuten lang behutsam mit Wasser sp len Vorhandene Kontaktlinsen nach M glichkeit entfernen Weiter sp len Call a POISON CENTER or doctor physician if you feel unwell Bei Unwohlsein GIFTINFORMATIONSZENTRUM oder Arzt anrufen Rinse mouth Mund aussp len IF skin irritation occurs Get medical advice attention Bei Hautreizung rztlichen Rat einholen rztliche Hilfe hinzuziehen Get medical advice attention Bei anhaltender Augenreizung rztliche Rat einholen rztliche Hilfe hinzuziehen If experiencing respiratory symptoms Call a POISON CENTER or doc tor physician Bei Symptomen der Atemwege Giftinformationszentrum oder Arzt anrufen Store in a well ventilated place Keep cool Beh lter dicht verschlossen an einem gut bel fteten Ort aufbewahren 10 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from blood Precaution phrases P 403 235 Store in a well ventilated place Keep cool K hl an einem gut bel fteten Ort aufbewahren For further information please see Material Safety Data Sheets www mn net com Weiterf hrende Informationen finden Sie in den Sicherheitsdatenbl ttern www mn net com MACHEREY NAGEL 03 2012 Rev 02 11 NucleoMag Blood 3 mL 5 Protocol for the isolation of genomic DNA from 3 mL blood samples using KingFisher Flex 24 The script necessary to run the NucleoMag Blood 3 mL kit on the KingFisher Flex 24 is available through MN technical s
7. cht Hautreizugen Causes serious eye irritation Verursacht schwere Augenreizung May cause allergy or asthma symptoms or breathing difficulties if inhaled Kann bei Einatmen Allergie asthmaartige Symptome oder Atembeschwerden verursa chen May cause respiratory irritation Kann die Atemwege reizen Precaution phrases P 210 P 233 P 261 P 280 P 301 312 P 302 352 P 304 340 P 305 351 338 P 312 P 330 P 3324313 P 337 313 P 3424311 P 403 233 Keep away from heat sparks open flames hot surfaces No smoking Von Hitze Funken offener Flamme heiBen Oberfl chen fernhalten Nicht rauchen Keep container tightly closed Beh lter dicht verschlossen halten Avoid breathing dust Einatmen von Staub vermeiden Wear protective gloves eye protection Schutzhandschuhe Augenschutz tragen IF SWALLOWED Call a POISON CENTER or doctor physician if you feel unwell BEI VERSCHLUCKEN Bei Unwohlsein GIFTINFORMATIONSZENTRUM oder Arzt anrufen IF ON SKIN Wash with plenty of soap and water BEI KONTAKT MIT DER HAUT Mit viel Wasser und Seife waschen IF INHALED Remove victim to fresh air and keep at rest in a position com fortable for breathing BEI EINATMEN An die frische Luft bringen und in einer Position rihigstellen die das Atmen erleichtert IF IN EYES Rinse continuously with water for several minutes Remove contact lenses if present and easy to do continue rinsing BEI KONTAKT MIT DEN A
8. e beads in Buffer MBL4 is of DNA in sufficient to remove ethanol However if necessary include a downstream 10 min airdrying step following the Buffer MBL4 wash step applications Low purity See above Time for magnetic separation too short Increase separation time to allow the beads to be completely attracted to the magnets Carry over of beads Incomplete magnetic bead separation High amounts of eluted DNA increase the viscosity and prevent the beads from being attracted completely to the magnets Increase elution buffer volume u lag Chase san Overfilling of wells from the 24 well separation plate mination oes contamination by splashing Do not overfill the wells of the separation plates to avoid cross 6 2 Ordering information Product REF NucleoMag Blood 3 mL 744502 1 1 x 96 preps NucleoMag Blood 200 uL 744501 1 1 x 96 preps 744501 4 4 x 96 preps Visit www mn net com for more detailed product information MACHEREY NAGEL 03 2012 Rev 02 15 Genomic DNA from blood 6 3 Product use restriction warranty NucleoMag Blood 3 mL kit components are intended developed designed and sold FOR RESEARCH PURPOSES ONLY except however any other function of the product being expressly described in original MACHEREY NAGEL product leaflets MACHEREY NAGEL products are intended for GENERAL LABORATORY USE ONLY MACHEREY NAGEL products are suited for QUALIFIED PERSONNEL ONLY MACHEREY NAG
9. ed completely at room temperature Elution at 55 C will increase the yield by about 15 20 NucleoMag B Beads are highly reactive superparamagnetic beads with a high binding capacity NucleoMag Blood 3 mL kit has been developed for use with ThermoFisher s KingFisher Flex 24 instrument A script is available on request from MACHEREY NAGEL The maximum sample volume of 3 mL is splitted into two aliquots of 1 5 mL each For processing smaller blood sample volumes use of liquid handling robots other than the KingFisher Flex 24 or manual extraction please inquire with MN technical support for details For smaller blood sample volumes MN offers the NucleoMag Blood 200 uL kit see ordering information MACHEREY NAGEL 03 2012 Rev 02 5 Genomic DNA from blood 2 3 Elution procedures Purified genomic DNA can be eluted directly with the supplied Elution Buffer MBL5 Elution can be carried out in a volume of gt 1 mL Smaller elution buffer volumes may result in incomplete bead separation For efficient elution the magnetic bead pellet should be resuspended completely in the elution buffer 6 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from blood 3 Storage conditions and preparation of working solutions Attention Buffers MBL1 MBL2 and MBL3 contain chaotropic salt Wear gloves and goggles CAUTION Buffer MBL1 contains guanidine hydrochloride which can form highly reactive compounds when combi
10. ffer volume insufficient Beads pellet must be covered completely with elution buffer Insufficient performance of elution buffer during elution step Remove residual buffers during the separation steps completely Remaining buffers decrease efficiency of subsequent washing and elution steps Beads dried out Poor DNA Do not let the beads dry as this might result in lower elution yield efficiency Partial elution in Wash Buffer MBL4 already Do not resuspend beads in Buffer MBL4 and do not incubate beads in this buffer for more than 2 min as this buffer is water based and might elute the DNA already Incubation after dispensing beads to lysate Mix immediately after dispensing NucleoMag B Beads and Binding Buffer MBL2 to the lysate Poor blood quality Be sure that no blood clots are transferred to the lysis plates Blood can be stored at 2 8 C for two weeks Freeze samples if stored for longer periods Low purity Incomplete magnetic bead separation High amounts of eluted DNA increase the viscosity and prevent the beads from being attracted completely to the magnets Increase elution buffer volume 14 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from blood Problem Possible cause and suggestions Carry over of ethanol from ethanol wash step e Be sure to remove all of the ethanol from the ethanol wash Suboptimal step Carry over of ethanol may interfere with downstream performance applications Typically washing th
11. hutz tragen S 37 39 Wear suitable gloves and eye face protection Bei der Arbeit geeignete Schutzhandschuhe und Schutzkleidung tragen 4 2 GHS classification Only harmful features do not need to be labeled with H and P phrases until 125 mL or 125 g Mindergef hrliche Eigenschaften m ssen bis 125 mL oder 125 g nicht mit H und P S tzen gekennzeichnet werden Component Hazard contents GHS symbol Hazard Precaution phrases phrases Inhalt Gefahrstoff GHS Symbol H S tze P S tze MBL1 Guanidine hydrochloride Danger 302 315 280 301 312 50 66 319 302 352 Guanidinhydrochlorid 50 66 Gefahr 305 351 338 330 332 313 337 313 MBL2 Sodium perchlorate 20 Warning 226 302 210 233 40 ethanol 35 55 301 312 330 Natriumperchlorat 20 40 Achtung 403 235 Ethanol 35 55 MBL3 Sodium perchlorate A Warning 226 210 233 5 20 ethanol 20 35 403 235 Natriumperchlorat 5 20 Achtung Ethanol 20 35 Proteinase K Proteinase K lyophilized Danger 315 319 261 280 Proteinase K Iyophilisiert O Gefahr 334 335 3024352 304 340 305 351 338 312 332 313 337 313 342 311 403 233 MACHEREY NAGEL 03 2012 Rev 02 9 Genomic DNA from blood Hazard phrases H 226 H 302 H 315 H 319 H 334 H 335 Flammable liquid and vapour Fl ssigkeit und Dampf entz ndbar Harmful if swallowed Gesundheitssch dlich bei Verschlucken Causes skin irritation Verursa
12. ilized x Xn R 36 37 38 S 22 24 Proteinase K lyophilisiert 42 26 36 37 Risk phrases R10 Flammable Entz ndlich R22 Harmful if swallowed Gesundheitssch dlich bei Verschlucken R 36 37 38 Irritating to eyes respiratory system and skin Reizt die Augen Atmungsorgane und die Haut R 36 38 Irritating to eyes and skin Reizt die Augen und die Haut R 42 May cause sensitization by inhalation Sensibilisierung durch Einatmen und Hautkontaktm glich Hazard labeling not necessary if quantity per bottle below 125g or mL certificate of exemption according to 67 548 EEC Art 25 1999 45 EC Art 12 and German GefStoffV 20 3 and TRGS 200 7 1 For further information see Material Safety Data Sheet 8 MACHEREY NAGEL 03 2012 Rev 02 Genomic DNA from blood Safety phrases 13 Keep away from food drink and animal foodstuffs Von Nahrungsmitteln Getr nken und Futtermitteln fernhalten S 16 Keep away from sources if ignition No smoking Von Z ndquellen fernhalten Nicht rauchen S 22 Do not breathe dust Staub nicht einatmen S 24 Avoid contact with the skin Ber hrung mit der Haut vermeiden S26 In case of contact with eyes rinse immediately with plenty of water and seek medical advice Bei Ber hrung mit den Augen sofort gr ndlich mit Wasser absp len und Arzt konsultieren S 36 37 Wear suitable protective clothing and gloves Bei der Arbeit geeignete Schutzhandschuhe und Schutzbrille Gesichtssc
13. lood 2 Product description 2 1 The basic principle The NucleoMag Blood 3 mL procedure is based on reversible adsorption of nucleic acids to paramagnetic beads under appropriate buffer conditions Whole blood is lysed with Lysis Buffer MBL1 and Proteinase K Following lysis incubation magnetic beads are added and binding conditions under which the DNA binds to the magnetic beads are adjusted by addition of Binding Buffer MBL2 After magnetic separation and removal of supernatant the paramagnetic beads are washed three times to remove contaminants and salt There is no need for a drying step as ethanol from previous wash steps is removed by Wash Buffer MBL4 Finally highly purified DNA is eluted with low salt Elution Buffer MBL5 and can directly be used for downstream applications NucleoMag Blood 3 mL is recommended for use on KingFisher Flex 24 instrument 2 2 Kit specifications The NucleoMag Blood 3 mL kit is made for isolation of genomic DNA from blood samples This kit provides reagents and magnetic beads for isolation of genomic DNA from 96 samples of up to 3 mL The purified DNA can be used directly as template for PCR blotting or any kind of enzymatic reactions The kit provides reagents for the purification of up to 100 130 ug of pure genomic DNA from 3 mL whole blood with an A A ratio 1 6 1 9 Fresh frozen or blood treated either with EDTA or citrate can be used NucleoMag Blood 3 mL kit can be process
14. ned with bleach sodium hypochlorite DO NOT add bleach or acidic solutions directly to the sample preparation waste All components of the NucleoMag Blood 3 mL kit should be stored at room temperature 18 25 C and are stable for up to one year All buffers are delivered ready to use Before starting NucleoMag Blood 3 mL protocol prepare the following Add the indicated volume of Proteinase Buffer PB to dissolve lyophilized Proteinase K see table below Proteinase K solution is stable at 20 C for up to 6 months NucleoMag Blood 3 mL 1 x 96 preps REF 744502 1 Proteinase K Add 3 75 mL Proteinase Buffer PB to each vial MACHEREY NAGEL 03 2012 Rev 02 T Genomic DNA from blood 4 Safety instructions The following components of the NucleoMag Blood 3 mL kits contain hazardous contents Wear gloves and goggles and follow the safety instructions given in this section 4 1 Risk and safety phrases Component Hazard contents Hazard Risk Safety symbol phrases phrases Inhalt Gefahrstoff Gefahrstoff R S tze S S tze symbol MBL1 Guanidine hydrochloride 50 66 x Xi R 22 36 38 S 26 Guanidinhydrochlorid 50 66 37 39 MBL2 Sodium perchlorate 20 40 R 10 22 S 13 16 ethanol 35 55 Natriumperchlorat 20 40 Ethanol 35 55 MBL3 Sodium perchlorate 5 20 R 10 S 16 ethanol 20 35 Natriumperchlorat 5 20 Ethanol 20 35 Proteinase K Proteinase K Iyoph
15. rmation on MACHEREY NAGEL products You may also contact your local distributor for general scientific information Applications mentioned in MACHEREY NAGEL literature are provided for informational purposes only MACHEREY NAGEL does not warrant that all applications have been tested in MACHEREY NAGEL laboratories using MACHEREY NAGEL products MACHEREY NAGEL does not warrant the correctness of any of those applications Last updated 07 2010 Rev 03 Please contact MACHEREY NAGEL GmbH amp Co KG Tel 49 24 21 969 270 tech bio mn net com Trademarks KingFisher is a registered trademark of Thermo Fisher Scientific NucleoMag is a registered trademark of MACHEREY NAGEL GmbH amp Co KG All used names and denotations can be brands trademarks or registered labels of their respective owner also if they are not special denotation To mention products and brands is only a kind of information i e it does not offend against trademarks and brands and can not be seen as a kind of recommendation or assessment Regarding these products or services we can not grant any guarantees regarding selection efficiency or operation MACHEREY NAGEL 03 2012 Rev 02 17
16. upport Before starting the preparation Check if Proteinase K was prepared according to section 3 Lyse sample Prepare KingFisher 24 deep well plate with buffers label deep well blocks before use Wash and elution buffers Fill 1 mL Elution Buffer MBL5 to each well of an empty Thermo KingFisher 24 deep well plate Fill 4 8 mL Wash Buffer MBL4 to each well of an empty Thermo KingFisher 24 deep well plate Fill 4 8 mL 80 ethanol to each well of an empty Thermo KingFisher 24 deep well plate Fill 4 8 mL Wash Buffer MBL3 to each well of an empty Thermo KingFisher 24 deep well plate Fill 4 8 mL Wash Buffer MBL3 to each well of a second empty Thermo KingFisher 24 deep well plate Fill 150 uL of Proteinase K working solution to each well of the two lysis plates Thermo KingFisher 24 deep well plates Please note that 3 mL blood samples have to be split and distributed into two plates 1 5 mL for each plate Fill 1 5 mL blood sample to a well of the lysis plate Thermo KingFisher 24 deep well plate with 150 uL Proteinase K per well Fill 1 5 mL blood of the same sample to the well at the same position of the second lysis plate Make sure that one sample is distributed into the same position of each deep well plate e g sample 1 to position A1 of lysis plate 1 and position A1 of lysis plate 2 sample 2 to position A2 of lysis plate 1 and position A2 of lysis plate 2 etc 12 MACHEREY NAGEL
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