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Catalog  Revision A   This warranty limits our liabili Catalog  Revision A   This warranty limits our liabili

Catalog Revision A This warranty limits our liabili - PDF document

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Catalog Revision A This warranty limits our liabili - PPT Presentation

No other warranties of any kind express or implied including without limitation im plied warranties of merchantability or fitness for a particular purpose are provided by Agilent Agilent shall have no liability for any direct indirect consequential ID: 52862

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TKLs Zarrant\ OLmLts our OLabLOLt\ to repOacement of tKLs product. No otKer ZarrantLes of an\ NLnd e[press or LmpOLed LncOudLnJ ZLtKout OLmLtatLon LmpOLed ZarrantLes of mercKantabLOLt\ or fLtness for a partLcuOar purpose are provLded b\ AJLOent. AJLOent sKaOO Kave no OLabLOLt\ for an\ dLrect LndLrect consequentLaO or LncLdentaO damaJes arLsLnJ out of tKe use tKe resuOts of use or tKe LnabLOLt\ to use tKLs product. RDERING NFORMATIONwww.agilent.com ECHNICAL ERVICEStechservices@agilent.com Telephone: 800 227 9770 (option 3,4,3) www.agilent.com/en/contact-us/page MaterLaOs ProvLded ............................................................... 1StoraJe CondLtLons ............................................................... 1AddLtLonaO MaterLaOs RequLred ........................................... 1,ntroductLon .......................................................................... 2PreparLnJ tKe ReaJents ........................................................ 3RNase-Free DNase , ............................................... 3 +LJK-SaOt WasK Buffer ........................................... 3 /oZ-SaOt WasK Buffer ............................................ 3 -MercaptoetKanoO ................................................. 3 ProtocoO ..................................................� 1;ခခခက........................... 4RNA ,soOatLon from CeOOs ....................................... 4 QuantLf\LnJ tKe RNA ............................................. 6 TroubOesKootLnJ ................................................................... 6AppendL[ , PreventLnJ SampOe ContamLnatLon .....................................................................� 1;ခခခခခခခခက PreventLnJ RNase ContamLnatLon ..................................................................................� 1;ခခခခခခက  SterLOL]LnJ /abZare ................................................  TreatLnJ SoOutLons ZLtK DEPC ....................................................................................� 1;ခခခခခခခခခက  PreventLnJ NucOeLc AcLd ContamLnatLon ...........................................................................� 1;ခခခက  AppendL[ ,, PurLf\LnJ RNA FoOOoZLnJ an En]\matLc ReactLon.................................................... AppendL[ ,,, ,soOatLnJ RNA from CeOOs +arvested b\ /aser Capture MLcrodLssectLon ........... 10MSDS ,nformatLon ............................................................. 10QuLcN-Reference ProtocoO ................................................. 11 Absolutely RNA Microprep Kit 1 ATERIALS ROVIDEDMaterials provided Quantity Lysis Buffer 35 ml -Mercaptoethanol (-ME) (14.2 M) 300 2600 U DNase Reconstitution Buffer 300 DNase Digestion Buffer 1.5 ml 24 ml 17 ml Elution buffer (10 mM Tris-HCl, pH 7.5) 3 ml RNA-Binding Spin Cups and 2-ml receptacle tubes 50 each 1.5-ml collection tubes 50 Sufficient reagents are provided to isolate total RNA from 50 samples of up to 5 × 10 cells each. Once opened, store at 4°C. Once reconstituted, store at –20°C. ONDITIONSUpon ReceLpt b-MercaptoetKanoO Once opened store at 4°C. RNase-Free DNase , AOO OtKer Components Once prepared store at room temperature. Guanidine thiocyanate in the lysis buffer and high-salt wash buffer is an irritant. ATERIALS EQUIREDDLetK\Op\rocarbonate DEPC EtKanoO [100% and 0% v/v ] RevLsLon D0 © AJLOent TecKnoOoJLes ,nc. 200-2015 2020 2 Absolutely RNA Microprep Kit NTRODUCTION TKe AbsoOuteO\ RNA mLcroprep NLt aOOoZs rapLd purLfLcatLon of KLJK-quaOLt\ totaO RNA from smaOO sampOes of cuOtured ceOOs or ceOOs Karvested b\ Oaser capture mLcrodLssectLon. TKe AbsoOuteO\ RNA mLcroprep NLt Ls desLJned for LsoOatLon from sampOes contaLnLnJ a ver\ smaOO number of ceOOs 1 ceOO to 5 × 10 ceOOs . Treatment of tKe sampOe ZLtK O\sLs buffer contaLnLnJ a stronJ proteLn denaturant tKe cKaotropLc saOt JuanLdLne tKLoc\anate  resuOts Ln effectLve ceOO O\sLs and prevents rLbonucOease RNase deJradatLon of tKe RNA. ,soOatLon of totaO RNA Ls performed Ln a sLnJOe mLcro-spLn cup to mLnLmL]e potentLaO Ooss of RNA durLnJ tKe procedure. FoOOoZLnJ ceOO O\sLs tKe soOutLon Ls mL[ed ZLtK etKanoO and appOLed to tKe RNA-bLndLnJ matrL[ ZLtKLn tKe mLcro-spLn cup. TKe RNA component of tKe sampOe bLnds to tKe matrL[ DNA contamLnatLon Ls effLcLentO\ removed foOOoZLnJ LncubatLon ZLtK DNase , dLrectO\ on tKe coOumn. TKe LmmobLOL]ed RNA Ls ZasKed to removecontamLnants and totaO RNA Ls recovered Ln 30 O of eOutLon buffer. See FLJure 1 for an overvLeZ of tKLs procedure. TKe purLfLed RNA Ls suLtabOe for use Ln a muOtLtude of moOecuOar bLoOoJ\ appOLcatLons LncOudLnJ cDNA s\ntKesLs RT-PCR quaOLtatLve andquantLtatLve RT-PCR nortKern bOottLnJ mLcroarra\ tarJet preparatLon and Overview of the Absolutely RNA microprep procedure. Absolutely RNA Microprep Kit 3 REPARING THE EAGENTS ReconstLtute tKe O\opKLOL]ed RNase-free DNase , b\ addLnJ 20 O of DNase ReconstLtutLon Buffer to tKe vLaO. ML[ tKe contents tKorouJKO\ to ensure tKat aOO tKe poZder Joes Lnto soOutLon. Do not Lntroduce aLr bubbOes Lnto tKe soOutLon. Store tKe reconstLtuted RNase-Free DNase , at –20°C. DNase Reconstitution Buffer is easily added to the vial of DNase with a syringe and needle. Gentle mixing is necessary because the DNase I is very sensitive to denaturation. High-Salt Wash Buffer Prepare 1 +LJK-SaOt WasK Buffer b\ addLnJ 16 mO of 100% etKanoO to tKe +LJK-SaOt WasK Buffer. After addLnJ tKe etKanoO marN tKe contaLner as suJJested ffer ] 1 (Ethanol Cap tKe contaLner of 1 +LJK-SaOt WasK Buffer tLJKtO\ and store at Low-Salt Wash Buffer Prepare 1 /oZ-SaOt WasK Buffer b\ addLnJ 6 mO of 100% etKanoO to tKe After addLnJ tKe etKanoO marN tKe contaLner as suJJested Buffer ] 1 (Ethanol . Cap tKe contaLner of 1 /oZ-SaOt WasK Buffer tLJKtO\ and store at 4 Absolutely RNA Microprep Kit P RNA Isolation from Cells 1. Add 0. O of -ME to 100 O of /\sLs Buffer for eacK sampOe of up to Caution The Lysis Buffer contains the irritant guanidine Do not exceed 5 × 10 cells per sample or the RNA yield may decrease. Prepare a fresh mixture of Lysis Buffer and -ME before Cell pellets can be stored at –80°C for future processing; however, homogenizing the cells in Lysis Buffer (the next step) prior to freezing the pellets is recommended to minimize RNA degradation. 2. Add 100 O of tKe /\sLs Buffer–-ME mL[ture to eacK ceOO sampOe and vorte[ or pLpet tKe sampOe repeatedO\ untLO KomoJenL]ed. Ensure that the viscosity of the lysate is low. High viscosity causes a decrease in RNA yield and an increase in DNA contamination. The viscosity can be reduced by additional vortexing, pipetting, and/or by increasing the volume of Lysis Buffer. 3. Add an equaO voOume usuaOO\ 100 O of 0% etKanoO to tKe ceOO O\sate and mL[ tKorouJKO\ b\ v It is important to vortex until the lysate and ethanol are thoroughly mixed. 4. Transfer tKLs mL[ture to a RNA-BLndLnJ SpLn Cup tKat Kas been seated ZLtKLn a 2-mO coOOectLon tube and snap tKe cap of tKe tube onto tKe top of tKe spLn cup. 5. SpLn tKe mL[ture Ln a mLcrocentrLfuJe for 30–60 seconds at m6. Remove and retaLn tKe spLn cup and dLscard tKe fLOtrate. Up to this point, the RNA has been protected from RNases by the presence of guanidine thiocyanate. Absolutely RNA Microprep Kit 5 Optional DNase Treatment TKLs procedure Ls recommended Lf DNA-free totaO RNA Ls requLred. TKe DNase treatment can be omLtted Lf tKe a. RepOace tKe spLn cup Ln tKe coOOectLon tube add 600 O of 1× /oZ-SaOt WasK Buffer and cap tKe tube. SpLn tKe sampOe Ln a mLcrocentrLfuJe at ma[Lmum speed for 30–60 seconds. b. Remove and retaLn tKe spLn cup and dLscard tKe fLOtrate. RepOace tKe spLn cup Ln tKe coOOectLon tube cap tube and spLn tKe sama mLcrocentrLfuJe at ma[Lmum speed for 2 mLnutes to dr\ tKe c. Prepare tKe DNase soOutLon b\ JentO\ mL[LnJ 5 O of reconstLtuted RNase-free DNase , ZLtK 25 Gentle mixing is necessary because the DNase I is very sensitive to denaturation. d. Add tKe DNase soOutLon dLrectO\ onto tKe matrL[ LnsLde tKe spLn cup and cap tKe tube. e. POace tKe sampOes Ln a 3C Lncubator for 15 mLnutes. . Add 500 O of 1× +LJK-SaOt WasK Buffer to tKe spLn cup cap tKe tube and spLn tKe sampOe Ln a mLcrocentrLfuJe at ma[Lmum speed for Caution The High-Salt Wash Buffer contains the irritant guanidine thiocyanate. . Remove and retaLn tKe spLn cup dLscard tKe fLOtrate and repOace tKe spLn cup Ln tKe coOOectLon tube. Add 600 O of 1× /oZ-SaOt WasK Buffer. Cap tKe tube and spLn tKe sampOe Ln a mLcrocentrLfuJe ama[Lmum speed for 30–60 seconds. 10. Remove and retaLn tKe spLn cup dLscard tKe fLOtrate and repOace tKe spLn cup Ln tKe coOOectLon tube. Add 300 O of 1× /oZ-SaOt WasK Buffer. Cap tKe tube and spLn tKe sampOe Ln a mLcrocentrLfuJe ama[Lmum speed for 2 mLnutes to dr\ tKe matrL[. 11. Transfer tKe spLn cup to a 1.5-mO coOOectLon tube. 6 Absolutely RNA Microprep Kit 12. Add 30 O of EOutLon Buffer dLrectO\ onto tKe matrL[ LnsLde tKe spLn cup. Snap tKe cap of tKe coOOectLon tube onto tKe spLn cup and tKe sampOe at room temperature for 2 mLnutes. SpLn tKe sampOe Ln a mLcrocentrLfuJe for 1 mLnute. TKLs eOutLon step ma\ be repeated to Lncrease tKe \LeOd of totaO RNA. The Elution Buffer must be added directly onto the matrix of the spin cup to ensure that the Elution Buffer permeates the entire matrix. The RNA yield may be increased by using Elution Buffer TKe purLfLed RNA Ls Ln tKe EOutLon Buffer Ln tKe mLcrocentrLfuJe tube. Cap tKe tube to store tKe RNA. TKe RNA can be stored at –20°C for up to one montK or at –0°C for OonJ-term storaJe. To quantLf\ RNA LsoOated from at Oeast 1 × 10ceOOs remove a smaOO sampOe and dLOute Lt ZLtK a buffer of neutraO p+ e.J. 10 mM TrLs p+ .5 . Measure tKe optLcaO densLt\ OD at 260 nm and 20 nm to quantLtate and quaOLf\ tKe RNA. RNA \LeOds ma\ var\ ZLtK tKe ceOO t\pe. YLeOds of 2– J of RNA can be e[pected from 5 × 10 cuOtured ceOOs and \LeOds of 0–200 pJ can be e[pected from 10 cuOtured ceOOs. To quantLf\ RNA LsoOated from Oess tKan 1 × 10ceOOs Lt Ls necessar\ to use a more sensLtLve fOuorescence-based s\stem e.J. RLbo*reen RNAObservation Suggestion RNA is degraded Use DEPC-treated or radiation-sterilized plasticware. RNA yield is poor Confirm that the range; exceeding 5 × 10 cells will result in a viscous lysate. Incubate the spin cup for 2 full minutes after adding the Elution Perform the elution twice and/or inBuffer. Any volume between 30 and 100 l can be used. subsequent applications Concentrate the RNA under vacuum without heat. Use a smaller volume of Elution Buffer, ensuring that the surface of the matrix is completelyl of Elution Buffer as long as the matrix is completely covered. The use of 0 however, will lower the RNA yield from a large number of cells. Dilute the homogenate with additghly viscous lysate will cause a large amount of genomic DNA to bind to the matrix. Ensure that the spin cup is centrifuged at maximum speed for 2 full minutes before DNase treatment. Absolutely RNA Microprep Kit 7 PPENDIX REVENTING ONTAMINATION RLbonucOeases are ver\ stabOe en]\mes tKat K\droO\]e RNA. RNase A can be temporarLO\ denatured under e[treme condLtLons but Lt readLO\ renatures. RNase A can tKerefore survLve autocOavLnJ and otKer standard metKods of proteLn LnactLvatLon. TKe foOOoZLnJ precautLons can prevent RNase JOoves at aOO tLmes durLnJ tKe procedures and ZKLOe KandOLnJ as RNases are present Ln tE[ercLse care to ensure tKat aOO equLpment e.J. centrLfuJe tubes etc. Ls as free as possLbOe from contamLnatLnJ RNases. AvoLd usLnJ equLor areas tKat Kave been e[posed to RNases. Use sterLOe tubes anmLcropLpet tLps onO\. MLcropLpettor bores can be a source of RNase contamLnatLon sLnmaterLaO accLdentaOO\ draZn Lnto tKe pLpet or produced b\ JasNeabrasLon can faOO Lnto RNA soOutLons durLnJ pLpettLnJ. COean mLcropLpettors accordLnJ to tKe manufacturer s recommendatLons. We recommend rLnsLnJ botK tKe LnterLor and e[terLor of tKe mLcropLpet sKaft ZLtK 0% etKanoO or 0% metKanoO. Sterilizing Labware DLsposabOe sterLOe pOastLcZare Ls JeneraOO\ free of RNases. ,f sterLOe pOastLcZare Ls unavaLOabOe components sucK as mLcrocentrLfuJe tubes can be sterLOL]ed and treated ZLtK dLetK\Op\rocarbonate DEPC  ZKLcK cKemLcaOO\ modLfLes and LnactLvates en]\mes accordLnJ to tKe fprotocoO Caution DEPC is toxic and extremely reactive. Always use DEPC in a fume hood. Read and follow the manufacturer's safety 1. Add DEPC to deLonL]ed Zater to a fLnaO DEPC concentratLon o0.1% v/v and mL[ tKorouJKO\. 2. POace tKe pOastLcZare to be treated Lnto a separate autocOacontaLner. CarefuOO\ pour tKe DEPC-treated Zater Lnto tKe contauntLO tKe pOastLcZare Ls submerJed. 3. /eave tKe contaLner and tKe beaNer used to prepare DEPC-treated Zater Ln a fume Kood overnLJKt. 8 Absolutely RNA Microprep Kit 4. For dLsposaO pour tKe DEPC-treated Zater from tKe pOastLcZare Lnto anotKer contaLner ZLtK a OLd. AutocOave tKe bottOe of Zaste DEPtreated Zater and tKe contaLner ZLtK tKe pOastLcZare for at Oea30 mLnutes. AOumLnum foLO ma\ be used to cover tKe contaLner but Lt sKouOd be KandOed ZLtK JOoves and cut from an area untoucKed b\unJOoved Kands. Nondisposable Plasticware Remove RNases from nondLsposabOe pOastLcZare ZLtK a cKOoroform rLnse. Before usLnJ tKe pOastLcZare aOOoZ tKe cKOoroform to evaporate Ln a Kood or rLnse tKe pOastLcZare ZLtK DEPC-treated Zater. Electrophoresis Gel Boxes To LnactLvate RNases on eOectropKoresLs JeO bo[es treat tKe JeO bo[es ZLtK 3% v/v K\droJen pero[Lde for 10–15 mLnutes and tKen rLnse tKem ZLtK RNase-free Zater. To LnactLvate RNases on JOassZare or metaO baNe tKe JOassZare or metaO for a mLnLmum of  Kours at 10°C. Treat Zater and soOutLons e[cept tKose contaLnLnJ TrLs base ZLtK DEPC usLnJ 0.1% v/v DEPC Ln dLstLOOed Zater. DurLnJ preparatLon mL[ tKe 0.1% soOutLon tKorouJKO\ aOOoZ Lt to Lncubate overnLJKt at rootemperature and tKen autocOave Lt prLor to use. ,f a soOutLon contaLns TrLs base prepare tKe soOutLon ZLtK a,f tKe LsoOated RNA ZLOO be used for cDNA s\ntKesLs for cDNA OLconstructLon or PCR ampOLfLcatLon Lt Ls Lmportant to remove an\ resLduaO nucOeLc acLds from equLpment tKat Zas used for prevLous nucOeLc acLd LsoOatLons. Absolutely RNA Microprep Kit 9 PPENDIX URIFYING OLLOWING AN NZYMATIC TKLs aOternate protocoO can be used to purLf\ RNA foOOoZLnJ an en]\matLc reactLon sucK as DNase dLJestLon or Ln vLtro transcrLptLon. TKLs metKod Ls most effLcLent for RNA moOecuOes of at Oeast 100 nucOeotLdes Ln OenJtK. TKLs protocoO Ls ZrLtten for reactLons of 60 O or Oess. TKe voOumes ma\ be scaOed up for OarJer reactLon voOumes. 1. Add 1.4 O of -ME to 200 2. AdMust tKe reactLon voOume to 60 O ZLtK Zater or buffer e.J. 10 mM TrLs p+  . 3. ML[ tKe /\sLs Buffer–-ME mL[ture and 140 O of 100% etKanoO Lnto 4. Transfer tKe mL[ture to a RNA-BLndLnJ SpLn Cup tKat Ls seated Ln a 2-mO receptacOe tube and cap tKe tube. 5. SpLn tKe tube Ln a mLcrocentrLfuJe for 30–60 seconds. 6. Remove and retaLn tKe spLn cup and dLscard tKe fLOtrate. RepOace tKe spLn cup Ln tKe receptacOe tube. . Add 500 O of 1× +LJK-SaOt WasK Buffer and cap tKe tube. SpLn tKe tube Ln a mLcrocentrLfuJe for 30–60 seconds. . Remove and retaLn tKe spLn cup and dLscard tKe fLOtrate. RepOace tKe spLn cup Ln tKe receptacOe tube. . Add 500 O of 1× /oZ-SaOt WasK Buffer and cap tKe tube. SpLn tKe tube Ln a mLcrocentrLfuJe for 30–60 seconds. 10. Remove and retaLn tKe spLn cup and dLscard tKe fLOtrate. RepOace tKe spLn cup Ln tKe receptacOe tube. 11. Add 300 O of 1× /oZ-SaOt WasK Buffer and cap tKe tube. SpLn tKe tube Ln a mLcrocentrLfuJe for 2 mLnutes. 12. Transfer tKe spLn cup to a 1.5-mO mLcrocentrLfuJe tube and dLscard tKe 2-mO receptacOe tube. 13. Add 30 O of EOutLon Buffer dLrectO\ to tKe matrL[ LnsLde tKe spLn cup.,ncubate tKe sampOe at 2 mLnutes. SpLn tKe tube Ln a mLcrocentrLfuJe for 30–60 seconds. TKLs eOutLon step ma\ be repeated 10 Absolutely RNA Microprep Kit PPENDIX SOLATING ELLS ARVESTED BY ASER ICRODISSECTION TKLs protocoO can be used ZLtK CapSure transfer fLOm carrLers /CM TKermo FLsKer ScLentLfLc to LsoOate RNA from ceOOs Karvested b\ Oaser capture mLcrodLssectLon. 1. Add 0. O of -ME to 100 O of /\sLs Buffer for eacK sampOe. 2. For eacK sampOe add 100 O of tKe /\sLs Buffer–-ME mL[ture to an Eppendorf standard 0.5-mO mLcrocentrLfuJe tube. 3. Snap a CapSure transfer fLOm carrLer contaLnLnJ Oaser-captured ceOOs 4. ,nvert and vorte[ eacK tube for 15–30 seconds to O\se tKe ceOOs tKat Zere captured on tKe CapSure transfer fLOm carrLer. 5. Proceed ZLtK tKe protocoO Ln RNA Isolation from Cells beJLnnLnJ ZLtK MaterLaO Safet\ Data SKeets MSDSs are provLded onOLne at . MSDS documents are not LncOuded ZLtK product sKLpments. EFERENCE ROTOCOLAdd 0.7 l of -ME to 100 5 × 10 cells l of Lysis Buffer–-ME mixture to each cell sample and vortex or pipet repeatedly until homogenized Add an equal volume of 70% ethanol to the cell lysate and mix thoroughly by vortexing for 5 seconds Transfer this mixture to a seated RNA-Binding Spin Cup; spin in a microcentrifuge at maximum speed for 30–60 seconds; retain the spin cup and discard the filtrate Optional DNase Treatment l of 1× Low-Salt Wash Buffer and spin in a microcentrifuge at maximum speed for 30–60 seconds Retain the spin cup and discard the filtrate; replace the spin cup and spin the tube in a microcentrifuge at maximum speed for 2 minutes Gently mix 5 l of reconstituted RNase-free DNase I with 25 l of DNase Digestion Buffer Add the DNase solution directly onto the matrix of the spin cup Incubate at 37C for 15 minutes l of 1× High-Salt Wash Buffer and spin in a microcentrifuge at maximum speed for 30–60 seconds Retain the spin cup and discard the filtrate l of 1× Low-Salt Wash Buffer and spin in a microcentrifuge at maximum speed for 30–60 seconds Retain the spin cup and discard the filtrate l of 1× Low-Salt Wash Buffer and spin in a microcentrifuge at maximum speed for 2 minutes to dry the matrix Transfer the spin cup to a 1.5 ml collection tube l of Elution Buffer directly onto the matrix and incubate for 2 minutes at room temperature; spin in a microcentrifuge at maximum speed for 1 minute The purified RNA is in the Elution Buffer in the microcentrifuge tube. Store at –80°C for the long term or at –20°C for the short term. Laboratory Reagent. utely RNA Microprep Kit ATERIALS ROVIDEDMaterials provided 35 ml -Mercaptoethanol (-ME) (14.2 M) 300 2600U DNase Reconstitution Buffer 300 1.5 ml 24 ml 17 ml Elution buffer (10 mM Tris-HCl, pH 7.5) 3 ml RNA-Binding Spin Cups and 2-ml receptacle tubes 50 each 1.5-ml collection tubes 50 Sufficient reagents are provided to isolate total RNA from 50 samples of up to 5 × 10 cells each. Once opened, store at 4°C. Once reconstituted, store at –20°C. ONDITIONSUpon Receipt: -Mercaptoethanol: Once opened, store at 4°C. RNase-Free DNase I: All Other Components: Once prepared, store at room temperature. Guanidine thiocyanate in the lysis buffer and high-salt wash buffer is an irritant. ATERIALS EQUIREDDiethylpyrocarbonate (DEPC) Ethanol >100% and 70% (v/v)] ‹ Agilent Technologies, Inc. 2007-2015, 2020 Materials Provided .............................................................................................................................. 1Storage Conditions .............................................................................................................................. 1Additional Materials Required .......................................................................................................... 1Introduction ......................................................................................................................................... 2Preparing the Reagents ....................................................................................................................... 3RNase-Free DNase I ............................................... 3 High-Salt :ash Buffer ........................................... 3 Low-Salt :ash Buffer ............................................ 3 -Mercaptoethanol ................................................................................................................ 3 Protocol ................................................................................................................................................ 4RNA Isolation from Cells ....................................... 4 Quantifying the RNA ............................................................................................................ 6 Troubleshooting .................................................................................................................................. 6Appendix I: Preventing Sample Contamination .............................................................................. 7Preventing RNase Contamination ......................................................................................... 7 Sterilizing Labware ............................................................................................................... 7 Treating Solutions with DEPC .............................................................................................. 8 Preventing Nucleic Acid Contamination ............................................................................... 8 Appendix II: Purifying RNA Following an Enzymatic Reaction.................................................... 9Appendix III: Isolating RNA from Cells Harvested by Laser Capture Microdissection ........... 10MSDS Information ............................................................................................................................ 10Quick-Reference Protocol ................................................. 11 Absolutely RNA Microprep Kit Instruction ManualCatalog #400805 Revision 0 Laboratory Reagent.