Mag-Bind®Blood DNA HDQ 96 Kit
Contents
1. Vortex at maximum speed for 10 minutes Add 320 uL HDQ Binding Buffer and 20 uL Mag Bind Particles HDQ to each sample Vortex at maximum speed for 10 minutes 10 11 12 13 Mag Bind Blood DNA HDQ 96 Protocols Place the plate on a magnetic separation device to magnetize the Mag Bind Particles HDQ Let sit at room temperature until the Mag Bind Particles HDQ are completely cleared from solution Note If MSD 01B is used the Mag Bind Particles HDQ should collect at the corner of each well adjacent to the magnet Aspirate and discard the cleared supernatant Do not disturb the Mag Bind Particles HDQ Remove the plate containing the Mag Bind Particles HDQ from the magnetic separation device Add 600 uL VHB Buffer to each sample Note VHB Buffer must be diluted with ethanol prior to use Please see Page 4 for instructions Resuspend the Mag Bind Particles HDQ by pipetting up and down 20 times or vortexing for 1 minute Note Complete resuspension of the Mag Bind Particles HDQ is critical for obtaining good purity Place the plate on the magnetic separation device to magnetize the Mag Bind Particles HDQ Let sit at room temperature until the Mag Bind Particles HDQ are completely cleared from solution Aspirate and discard the cleared supernatant Do not disturb the Mag Bind Particles HDQ Remove the plate containing the Mag Bind Particles HDQ from the magnetic separation device2. must be at room temperature Dry the Mag Bind Particles HDQ before elution Ordering Information The following components are available for purchase separately Call Toll Free at 1 800 832 8896 Elution Buffer EB Buffer 100 mL Elution Buffer EB Buffer 500 mL RNase A 400 uL RNase A 5 mL Omega Homogenizer Columns 50 Omega Homogenizer Columns 200 1 5 mL DNase RNase free Microcentrifuge Tubes 2 mL DNase RNase free Microcentrifuge Tubes Magnetic Separation Device for Microplates 96 well Round well Plates 1 2 mL 96 well Microplates 500 uL Sealing Film HiBind E Z N A and MicroElute are registered trademarks of Omega Bio tek Inc Tissue Tearor and Tissuemizer are trademarks of their respective companies PCR is a patented process of Hoffman La Roche Use of the PCR process requires a license Notes
3. Ca OMEGA Innovations in nucleic acid isolation bio tek Product Manual Mag Bind Blood DNA HDQ 96 Kit M6399 00 1 x 96 preps M6399 01 4 x 96 preps M6399 02 20 x 96 preps August 2013 For research use only Not intended for diagnostic testing Mag Bind Blood DNA HDQ 96 Kit Table of Contents Introduction and OVEFVIEW ssssssecseccssecseccssecssessecssccssecssccssesseecseeessess Kit Contents Storage and Stability ssssssecsecssecneecseesseeseeneese Preparing Reagents ssesseseseesesserssseeessseessseesssesssseesssssessseessseerssseeessseess Mag Bind Blood DNA HDQ 96 Protocol for 100 200 uL Troubleshooting GUIDE xicissscisesssieericisnnncceaniomnicimeonnnimeninsie OrderiNg ereer E E RA A EAS Manual Revision August 2013 N OMEGA bio tek Innovations in nucleic acid isolation Introduction and Overview Introduction The Mag Bind Blood DNA HDQ 96 Kit is designed for rapid and reliable isolation of high quality genomic DNA from 100 200 uL blood samples All heating steps that limit robotic applications have been removed to allow for faster processing Mag Bind Particles HDQ provide quick magnetic response time reducing overall processing time This system combines the reversible nucleic acid binding properties of Mag Bind paramagnetic particles with the time proven efficiency of Omega Bio tek s blood DNA isolation system to provide a fast and convenient method to isolated DNA from fre
4. Repeat Steps 8 12 for a second VHB Buffer wash step 14 15 16 17 18 19 20 21 22 23 24 Mag Bind Blood DNA HDQ 96 Protocols Add 600 uL SPM Wash Buffer to each sample Note SPM Wash Buffer must be diluted with ethanol prior to use Please see Page 4 for instructions Resuspend the Mag Bind Particles HDQ by pipetting up and down 20 times or vortexing for 1 minute Let sit at room temperature for 1 minute Place the plate on the magnetic separation device to magnetize the Mag Bind Particles HDQ Let sit at room temperature until the Mag Bind Particles HDQ are completely cleared from solution Aspirate and discard the cleared supernatant Do not disturb the Mag Bind Particles HDQ Leave the plate on the magnetic separation device for 10 minutes to air dry the magnetic particles Remove any residue liquid with a pipettor Remove the plate containing the Mag Bind Particles HDQ from the magnetic separation device Add 100 200 uL Elution Buffer or nuclease free water to elute DNA from the Mag Bind Particles HDQ Resuspend the Mag Bind Particles HDQ by pipetting up and down 50 times Let sit at room temperature for 10 minutes Place the plate on the magnetic separation device to magnetize the Mag Bind Particles HDQ Let sit at room temperature until the Mag Bind Particles HDQ are completely cleared from solution Transfer the cleared supernatant containing purified DNA t
5. e at room temperature ee O ooe OE 3 Prepare HDQ Binding Buffer as follows and store at room temperature M6399 01 160 mL M6399 02 800 mL 4 Shake or vortex the Mag Bind Particles HDQ to fully resuspend the particles before use The particles must be fully suspended during use to assure proper binding Mag Bind Blood DNA HDQ 96 Protocols Mag Bind Blood DNA HDQ 96 Protocol 100 200 uL Blood The procedure below has been optimized for use with 100 200 uL FRESH or FROZEN blood samples Buffy coat can also be used Materials and Reagents to be Supplied by User 100 ethanol 100 isopropanol Magnetic separation device for microplates Cat MSD 01B Vortexer 96 well Microplate 500 uL Cat EZ9604 or desired elution plate 96 well Round well Plate 1 2 mL Cat SS11780 Sealing film Cat AC1200 Optional PBS or nuclease free water Optional RNase A 10 mg mL Before Starting Prepare SPM Wash Buffer HDQ Binding Buffer and VHB Buffer according to the Preparing Reagents section on Page 4 Add blood samples to a 96 well Round well Plate 1 2 mL Bring the volume up to 200 uL with PBS not provided or Elution Buffer provided with this kit if volume of blood is less than 200 uL Add 10 uL Proteinase K Solution to each sample Vortex or pipet up and down 20 times to mix Optional Add 5 uL RNase A to each sample Vortex or pipet up and down 20 times to mix Add 230 uL AL Buffer to each sample
6. o a clean microplate not supplied Store the DNA at 20 C Troubleshooting Guide Please use this guide to troubleshoot any problems that may arise For further assistance please contact the technical support staff toll free at 800 832 8896 Problem Low DNA yield Mag Bind Particles HDQ do not completely clear from solution Gel like material in the eluted DNA Problems in downstream applications Incomplete resuspension of Mag Bind Particles HDQ Frozen blood samples not mixed properly after thawing Loss of Mag Bind Particles HDQ during operation DNA remains bound to Mag Bind Particles HDQ DNA washed off Ethanol is not added into VHB buffer Too short of magnetizing time Blood is too old Salt carry over Ethanol carry over Resuspend the Mag Bind Particles HDQ by vortexing vigorously before use Thaw the frozen blood at room temperature and gently mix the blood by inverting Avoid disturbing the Mag Bind Particles HDQ during aspiration Increase elution volume and incubate at for 15 minutes pipet up and down 50 to 100 times Dilute SPM Wash Buffer by adding appropriate volume of ethanol prior to use see Page 4 for instructions Make sure to add ethanol to the VHB Buffer see Page 4 for instructions Increase collection time on the magnet Remove the gel like material by centrifugation recommend using fresh blood Use 8 mM NaOH as elution buffer SPM Wash Buffer
7. sh or frozen blood Utilizing paramagnetic particles provides high quality DNA that is suitable for direct use in most downstream applications such as amplification and enzymatic reactions Overview If using the Mag Bind Blood DNA HDQ 96 Kit for the first time please read this booklet in its entirety to become familiar with the procedures Blood cells are lysed in a specially formulated buffer DNA is isolated from the lysates in one step by binding to Mag Bind Particles surfaces The paramagnetic particles are separated from the lysates by using a magnetic separation device After a few rapid wash steps to remove trace contaminants DNA is eluted in Elution Buffer Kit Contents ae Sw Mag Bind Particles HDQ 40 mL AL Buffer 600 mL HDQ Binding Buffer 200 mL VHB Buffer 2x 440 mL SPM Wash Buffer 2x 300 mL Proteinase K Solution 10 mg mL 22 mL Elution Buffer 400 mL Storage and Stability All of the Mag Bind Blood DNA Kit components are guaranteed for at least 12 months from the date of purchase when stored as follows Mag Bind Particles HDQ should be stored at 2 8 C for long term use Proteinase K Solution can be stored at room temperature for up to 12 months For long term storage store Proteinase K Solution at 2 8 C Preparing Reagents 1 Dilute SPM Wash Buffer with 100 ethanol as follows and store at room temperature M6399 01 280 mL M6399 02 700 mL per bottle 2 Prepare VHB Buffer as follows and stor
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