Eppendorf Eporator®
Contents
1. Fig 3 Display 1 Actual voltage value in V 2 Actual discharge time in ms 3 Voltage symbol 4 Cuvette symbol Voltage symbol kd appears after Cuvette symbol U appears when a cuvette electroporation and disappears after the is inserted cuvette carrier has been removed 5 Set voltage in V 5 2 Recommendations for sample preparation Independent of the device the success of an electroporation is influenced by a variety of factors Quality and concentration of the inserted DNA Quality and concentration of the cells Resuspension medium of the DNA and cells 5 2 1 DNA preparation DNA quality In order to obtain a high transformation efficiency the DNA solution should be pure and free of salts Buffer DNA dissolved in TE buffer is acceptable if the DNA was dissolved in approx ten times the quantity of electrocompetent cells Salt concentration DNA from enzyme reactions e g ligation can be immediately used for electroporation if the salt concentration is under 5 M If the ionic strength of the reaction combination is too high it can be reduced via dilution or ethanol precipitation After ethanol precipitation the DNA can be resuspended in sterile demineralized water or TE buffer Incubation Before electroporation do not incubate the DNA with the cell suspension too long Generally the DNA should be added to the cells one minute before electroporation and the solution should be incubated at 0 C Long incubati2. The DNA The quantity and quality of the DNA used should be checked before electroporation Incorrectly concentrated or degraded DNA leads to low transformation efficiency Salts and other components that can have a toxic effect on cells must be removed from the DNA preparation before the preparation process The DNA preparation should be added to the cells no longer than one minute before the electroporation DNase present in cell preparation can degrade the DNA and thereby cause a low transformation efficiency see DNA preparation on p 14 The temperature Electroporation cuvettes should be cooled to 0 C 4 C before electroporation see Temperature on p 15 This produces better results than with electroporation cuvettes at room temperature If frozen cells are used electroporation should be performed immediately after thawing Frozen cells can be stored a maximum of 6 12 months in 10 15 glycerine at 80 C Deviating voltages during transformation The voltage applied to the electroporation cuvette act greatly differs from the set voltage se A too low resistance has several causes The cells were washed and resuspended in a buffer with a too high ionic strength The cells were not sufficiently cleaned during the preparation After insufficient washing growth medium residue which has been carried along leaves behind unwanted salts Lysed cells are in the preparation They contribute to the reduction
3. aa 9 7 1 9 10 3 1 Intended sess vita caw ses 7 7 7 a A 10 32 User profiles 6 10 3 3 Information on product liability ah 10 3 4 Safety instructions on the device aa 10 3 5 Warnings for intended use 11 4 Installation ug san ne 12 4 1 Preparing installation sa na ane nn 12 4 2 Selecting the locaton dc 222 shina EIERE ane hd Pas d 12 4 3 Installing the instrument ea e ne 12 5 Operation oii 13 5 1 Control element overview es 13 5 2 Recommendations for sample preparation eeeeeee 14 5 2 1 DNA preparation NN NEIER Has an a a ewe ees 14 5 2 2 Electroporation medium 15 5 2 3 Growth and preparation of ces 15 5 24 Temperature 2 15 5 3 Performing electroporation a s ssaussssnussaranrrunrananrnrrsrrsrunas 16 5 3 1 Switching on the device aa aaa 16 5 3 2 Inserting cuvette aaa se ke nanen 16 5 3 3 Electroporation vi
4. Eppendorf Eporator Operating manual eppendorf Copyright 2012 Eppendorf AG Hamburg No part of this publication may be reproduced without the prior permission of the copyright owner Eppendorf and the Eppendorf logo are registered trademarks of Eppendorf AG Hamburg Germany Eppendorf Eporator is a registered trademark of Eppendorf AG Hamburg Germany Excel and Microsoft are registered trademarks of Microsoft Corporation in the United States and other countries Registered trademarks are not marked in all cases with TM or in this manual 4309 900 010 03 122012 Eppendorf Eporator Operating manual Table of contents 1 Operating instructions ca da ENEE TEE 5 1 1 Using thismanual eb Li a he 5 1 2 Danger symbols and danger levels aaa 5 1 2 1 Hazard SYMDOIS ng 244032 ea area 5058 5 1 2 2 Degrees of dange 5 1 3 Symbols US Q c 3 3 3 33 5 TA 0 GJOSSAKVs 7 7 4 45 0 0 6 2 Product description 7 2 1 Main lUSHALONI case ce 8 7 2 2 Delivery package AN AREND 8 2 3 Features u a aa 8 2 3 1 The principle of electroporation
5. in the business to business sphere to which this product is assigned may no longer be disposed of in municipal or domestic waste To document this they have been marked with the following identification Because disposal regulations may differ from one country to another within the EU please contact your supplier if necessary In Germany this is mandatory from March 23 2006 From this date the manufacturer has to offer a suitable method of return for all devices supplied after August 13 2005 For all devices supplied before August 13 2005 the last user is responsible for the correct disposal 29 ce c ce Ze C te DK ci CE TE C ve C e CS e t e ce Se ce C CE TE C ve C e C e CE e Ci ce Te CE Te C ve C t ec EG Konformit tserkl rung wa Ko EC Conformity Declaration dE Ci H Das bezeichnete Produkt entspricht den einschl gigen grundlegenden Anforderungen der Cc aufgef hrten EG Richtlinien und Normen Bei einer nicht mit uns abgestimmten nderung des 6 C PA ce Produktes oder einer nicht bestimmungsgem en Anwendung verliert diese Erkl rung ihre G ltigkeit D ce The product named below fulfills the relevant fundamental requirements of I A c te the EC directives and standards listed In the case of unauthorized modifications to the product ce t ce ce or an unintended use this declaration becomes invalid eS Produktbezeichnung Product name ce C Vorstand Board of
6. electroporation of various bacteria and yeast strains can be found on the Eppendorf home page www eppendorf com Eppendorf Eporator Operating manual 2 3 1 The principle of electroporation qmo CO KI S e DNA and bacteria Electrical charge DNA in bacteria 200 V 2 500 V approx 5 ms With the electroporation method macromolecules such as DNA can be placed in electrocompetent bacteria or yeast strains In the process small volume samples with high resistance are exposed to pulses with very high electrical field strengths The short high voltage pulses create temporary holes or pores in the cell membrane through which macromolecules e g plasmid DNA can diffuse into the cell The holes close after removal of the electrical field and a period of regeneration The inserted plasmid DNA can then be transcribed and replicated in the cell Unlike chemical transformation electroporation is characterized by high transformation efficiency and simple execution 10 Eppendorf Eporator Operating manual 3 Safety 3 1 Intended use The Eporator is intended for indoor use only and enables the simple and safe electroporation of bacteria and yeast strains using standard protocols 3 2 User profile This device must only be used by skilled personnel with the appropriate training Before using the device read the operating manual carefully and familiarize yourself with the device s mo
7. of the resistance of the medium The salt concentration in the DNA preparation is too high Eppendorf Eporator Operating manual 6 2 Error messages If the proposed measures repeatedly fail to remedy the fault contact your local Eppendorf partner The addresses of our distributors can be found on our website www eppendorf com The addresses of our sales offices can be found on the second to last page of this operating manual gt Quit all error messages with the Exit key 6 2 1 Operational error Symptom message Cause Remedy Display remains dark The device was not connected to the mains supply or the mains power switch was not switched on gt Check the mains connection and the power cable gt Switch on the device function not The display shows Akey that is not available in the current device state was gt Message disappears after about 2 seconds located in the device available pressed e g the Exit key in the main display The display shows The Start key was pressed 1 Insert an electroporation no cuvette before an electroporation cuvette cuvette was inserted 2 Start electroporation see Inserting cuvette on p 16 The display shows The export command was 1 Insert a USB stick in the USB no USB stick activated before a USB stick port of the device was inserted in the USB port of 2 Repeat the export command
8. the device see Exporting data on p 20 The display shows A USB stick with no storage 1 Insert a USB stick with USB stick full capacity was inserted in the sufficient storage capacity in USB port of the device the USB port of the device 2 Repeat the export command see Exporting data on p 20 The display shows The export of the data from the 1 Insert a standard USB stick in no export device has failed the USB port of the device All existing electroporation 2 Repeat the export command protocols have already been see Exporting data on p 20 saved on the USB stick The display shows The export of data has failed 1 Perform electroporation no protocol No exportable protocols are 2 Repeat the export command see Exporting data on p 20 23 Eppendorf Eporator Operating manual 6 2 2 Device error Symptom Cause Remedy message The display shows Device error 1 Quit the error message with the Hardware error Exit key 2 Perform the electroporation again gt If the error message appears again switch the device off and on The display shows Device error 1 Quit the error message with the internal error Exit key 2 Perform the electroporation again gt Ifthe error message appears again switch the device off and on Eppendorf Eporator Operating manual 7 Maintenance 7 1 Cleaning DANGER Electric shock as a result of penet
9. DANGER Will lead to severe injuries or death WARNING May lead to severe injuries or death CAUTION May lead to light to moderate injuries NOTICE May lead to material damage 1 3 Symbols used Depiction Meaning gt You are requested to perform an action 1 Perform these actions in the sequence described 2 List Text Terms and key names from the software Ci References useful information Eppendorf Eporator Operating manual 1 4 Glossary A Arcing If electrical voltage is applied between two parallel electrodes a current flows in an evenly distributed layer If the voltage exceeds a critical value this layer contracts to a narrow circuit with high current density an electric arc The electrode material melts at this location and explosive evaporation occurs The cuvette can be destroyed under these conditions E Electrical field Ratio of potential difference between two electrodes in V and the distance strength between these electrodes electrode gap in cm However this only applies if the electrical field is homogeneous as with parallel plate electrodes for example in Eppendorf electroporation cuvettes T Time constant Time during which the voltage decreases to the value U e Eppendorf Eporator Operating manual 2 Product description 2 1 Main illustration Fig 1 Main Illustration 1 Operating controls Cuvette carrier In the cuvette s
10. Management Ce 13 01 2012 ce ce ce Hamburg Date Se 5 Eppendorf AG Barkhausenweg 1 22339 Hamburg Germany H GE ce C e CE T C T C qe Ci T Te T Te T Te C t KE ee E t Te ce Te C C E e c E CE 0015 033 509 02 c ce ce SA d A c g C ce ce c H goxo bogozC Ce er ek Ki ce ve C LE CE Le CE e SS e Sy ce C nr es ern ce e ce EN g eS l V j 1 Li Evaluate your operating manual www eppendorf com manualfeedback eppendorf Your local distributor www eppendorf com worldwide Eppendorf AG 22331 Hamburg Germany Tel 49 40 53801 0 Fax 49 40 538 01 556 E mail eppendorf eppendorf com Eppendorf North America Inc 102 Motor Parkway Hauppauge N Y 11788 5178 USA Tel 1 516 334 7500 Toll free phone 1 800 645 3050 Fax 1 516 334 7506 E mail info eppendorf com Application Support Europe Tel 49 1803 666 789 Preis je nach Tarif im Ausland 9 ct min aus dem dt Festnetz Mobilfunkh chstpreis 42 ct min support eppendorf com North America Tel 1 800 645 3050 E mail techserv eppendorf com Asia Pacific Tel 60 3 8023 6869 E mail support_asiapacific eppendorf com
11. de of operation 3 3 Information on product liability In the following cases the protection provided by the device may be impaired The liability for the function of the device passes to the operator if The device is not used in accordance with the operating manual The device is used outside of the range of application described in the preceding chapters The owner has made unauthorized modifications to the device 3 4 Safety instructions on the device Depiction Meaning WARNING A Follow the operating manual Eppendorf Eporator Operating manual 3 5 Warnings for intended use WARNING Damage to health from toxic radioactive or aggressive chemicals as well as infectious liquids and pathogenic germs gt Observe the national regulations for handling these substances the biological security level of your laboratory the material safety data sheets and the manufacturer s application notes gt Wear personal protective equipment PPE WARNING Lethal voltages inside the device Ensure that the housing is always closed and undamaged so that no parts inside the device can be contacted by accident gt Do not remove the housing of the device gt Do not allow any liquids to enter the inside of the housing gt Do not allow the device to be opened by anyone except service personnel who have been specifically authorized by Eppendorf WARNING Risk of electrical shock fro
12. e Hold down the desired program key for at least 2 seconds A signal tone sounds Voltage stored appears in the display The control LED over the pressed program key illuminates blue The voltage is saved under the corresponding program number 1 to 2 19 20 Eppendorf Eporator Operating manual 5 7 Enhanced settings Additional settings can be carried out in the menu The date and time can be defined in the device in order to track exported data The following settings are available Fig 6 Menu display 1 Data export 2 Date The data export is described in the chapter Set date Export data see p 20 3 Time 4 Signal tone Set time Set signal tone The display switches between vol very quiet vol2 quiet vol loud vol4 very loud and vol OFF signal tone switched off Open menu B 1 Press menu enter key Switch between parameters B 2 Press menu enter key The selected parameter blinks in the display Change the value of the parameter O 3 Press the arrow keys Exit the menu 4 Press the exit key The changed parameters are automatically saved 5 8 Exporting data Using the USB port at the rear of the device you can save the last 50 experiments on a USB stick in separate TXT files see Main illustration on p 7 The files are named according to the corresponding sample number This format is well suited for additional editing in a text editor or Microsoft E
13. e holder and carefully transfer the sample to the corresponding medium without bubbles 5 4 Regeneration of the cells Example for the bacterium E coli 1 After the electroporation immediately place about 1 mL fresh medium without selection chemicals on the cells A rich medium is best suited for this e g the SOC medium for E coli 2 Carefully resuspend cells and transfer them to a new tube 3 Incubate cells at optimal growth temperature e g 37 C for E coli for one hour at light vibration e g with the Eppendorf Thermomixer comfort 5 5 Determination of the transformation efficiency After the recovery period the cells should be plated with a selection medium To determine the efficiency streak different cell concentrations and use this information to calculate the number of transformers ug DNA 5 6 Programs A program contains a saved voltage setting for quick access to frequently used settings 5 6 1 Loading program The programs 1 and 2 are stored with the following parameters at delivery Program key P1 1 700 V e g for E coli electroporation in 1 mm electroporation cuvettes Program key P2 2 500 V e g for E coli electroporation in 2 mm electroporation cuvettes Eppendorf Eporator Operating manual Press desired program key The control LED above the pressed program key illuminates blue the voltage is displayed 9 Saving program 1 2 Use the arrow keys to set the voltag
14. haft 3 RS232 port Only for Technical Service USB port For inserting a USB stick 5 Mains switch Mains connection socket Eppendorf Eporator Operating manual 2 2 Delivery package Number Description 1 Eppendorf Eporator 1 Mains cable 1 Cuvette carrier 5 Electroporation cuvette 1 mm width of slit 5 Electroporation cuvette 2 mm width of slit 1 Eporator operating manual 2 3 Features The Eporator is inserted for the electroporation It contains a capacitor that is discharged during electroporation using a resistor thus generating an exponential discharge curve A voltage between 200 V and 2 500 V can be set The exponential pulse generated by the Eporator is transferred to a disposable electroporation cuvette that contains the biological sample Unlike devices from other manufacturers the Eporator features an integrated cuvette carrier with cuvette holder The construction of the Eporator minimizes the risk of short circuits This also applies in the case of prohibited high salt concentrations or maximum voltage Even in the most unlikely situation of an electric arc in the cuvette no bacterial suspension can escape from the cuvette and contaminate the device The Eporator is easy to operate None of the device components require user maintenance The experimental electroporation data can be saved on a USB stick and evaluated on a PC Application protocols for the
15. jintormali oni es 27 9 1 Oto Ae 71 1 10 27 9 2 dsp e ed bn hw 27 10 Transport storage and disposal 2 28 10 1 Sofage 28 10 2 Decontamination before shipment eeeeee eee 28 110 3 Transport 1 1 5 28 10 4 Disposals s EE 29 Eppendorf Eporator Operating manual 1 Operating instructions 1 1 this manual gt Please read this operating manual completely before using the device for the first time gt Please view this operating manual as part of the product and keep it somewhere easily accessible gt When passing the device on to third parties be sure to include this operating manual gt If this manual is lost please request another one The current version can be found on our website www eppendorf com 1 2 Danger symbols and danger levels 1 2 1 Hazard symbols Toxic substances Electric shock Hazard point Material damage p g 1 2 2 Degrees of danger The degree of danger is a part of a safety note and distinguishes the possible results of non observance from each other
16. leshooting 6 1 General errors Many factors can contribute to a low transformation efficiency The set voltage Specific voltage parameters exist for each microorganism Some cells die during electroporation If the field strength is too high or too low a low transformation efficiency is achieved The expected survival rate varies between 20 and 80 of inserted cells The electroporation of E colirequires an impulse of approximately 5 ms and field strengths between 12 kV cm and 19 kV cm You should check the transformation efficiency at different voltages in order to optimize conditions Application protocols for the electroporation of various bacteria and yeast strains can be found on the Eppendorf home page www eppendorf com The cells Generally cells are transformed most efficiently when they are in an early to medium log phase Different growth conditions can improve the transformation efficiency see Growth and preparation of cells on p 15 If too many cells are killed the electroporation conditions for the strain must be optimized and the DNA preparation and cell preparation for toxic or organic substances must be inspected see DNA preparation on p 14 After electroporation cells especially E coli must be immediately transferred to a rich medium in order to obtain good results Even a small delay in completing this step can lead to a significantly lower transformation efficiency see Regeneration of the cells on p 18
17. m damage to the device or mains cable gt The device may only be switched on if the device and mains cable are undamaged gt Only use devices which have been professionally installed or repaired WARNING Danger from using an incorrect power supply gt Only connect the device to voltage supplies which correspond with the electrical requirements on the name plate gt Only use sockets with a protective grounding conductor and a suitable mains cable CAUTION Poor safety due to incorrect accessories and spare parts The use of accessories and spare parts other than those recommended by Eppendorf may impair the safety functioning and precision of the device Eppendorf cannot be held liable for any damage resulting from the use of non recommended accessories and spare parts or from the improper use of such equipment gt Only use accessories and original spare parts recommended by Eppendorf NOTICE Damage to device due to infiltration of liquids When electroporation is performed in electroporation cuvettes that do not have lids liquid can enter the device gt Only use electroporation cuvettes with lids 11 12 Eppendorf Eporator Operating manual 4 Installation 4 1 Preparing installation Retain the transport carton and packing material for subsequent safe transport or storage gt Check all parts for any transport damage 4 2 Selecting the location Please select the loca
18. on times can lead to DNA degradation due to the DNases in the cell suspension DNA concentration The concentration of DNA can significantly influence the transformation efficiency Frequency and efficiency The frequency is defined as the number of transformed cells per surviving cells The efficiency is defined as transformed cells per ug DNA You can obtain a high frequency by using high DNA concentrations High efficiency is achieved by using high cell concentration Reducing the DNA concentration helps prevent co transformations of the same cell Eppendorf Eporator Operating manual 5 2 2 Electroporation medium Sensitivity of the cells The cells are sensitive to external influences because electroporation creates temporary pores in the cell membrane Electrolysis of the medium During electroporation the electrolysis of the medium significantly influences the characteristics of the medium e g the pH value Many cells can die if fresh medium is not immediately added after electroporation for the recovery ofthe cells see Regeneration of the cells on p 18 lonic strength of the medium The ionic strength of the medium must be taken into account during electroporation of cells In order to keep the resistance of the medium as high as possible salts must be removed from the cell and DNA preparation lons remaining in the cell suspension often come from the culture medium Higher transformation efficiency can be obtained b
19. poration 5 3 1 Switching on the device gt Press the mains power switch on the rear of the device to switch on the device 5 3 2 Inserting cuvette To increase the efficiency of the electroporation the electroporation cuvette can be cooled prior to filling Remove residual liquids from the cuvette before further use The integrated cuvette carrier is located at the front of the device to the right of the control panel Proceed as follows 1 Remove cuvette carrier from the device The cuvette mount is now freely accessible for the insertion of the electroporation cuvette Eppendorf electroporation cuvettes are sterilized with gamma rays and individually packaged They are available in 3 different sizes Width of slit and filling quantity are noted on the cuvettes 1mm up to 100 uL filling quantity 2 mm up to 400 UL filling quantity 4 mm up to 800 UL filling quantity 2 Remove the electroporation cuvette from the individual packaging Remove the lid of the electroporation cuvette 4 Fill the electroporation cuvette with the sample The slit between the plate electrodes must be filled without bubbles Di Eppendorf electroporation cuvettes can be labeled using a matte window 5 Place the locking lid on the electroporation cuvette NOTICE Damage to device due to infiltration of liquids When electroporation is performed in electroporation cuvettes that do not have lids liquid can en
20. ration of liquid gt Switch off the device and disconnect it from the power supply before any maintenance or cleaning work is carried out gt Do not allow any liquids to enter the inside of the housing gt Do you use any spray disinfectants on the housing gt Only reconnect the device to the power supply once it is completely dry NOTICE Damage from using aggressive chemicals Do not use any aggressive chemicals on the device or its accessories such as strong and weak bases strong acids acetone formaldehyde halogenated hydrocarbons or phenol gt Ifthe device becomes contaminated with aggressive chemicals clean it immediately with a mild cleaning agent gt Weta cloth with a mild cleaning fluid and demineralized water and remove the contamination on the outside of the device 7 2 Replacing fuses The fuse holder is located between the mains connection socket and the mains power switch 1 Disconnect the power plug 2 Press the upper and lower end of the plastic springs 1 together and pull the fuse holder 2 fully out 3 Replace faulty fuses and reinsert the fuse holder Make sure that the guiding rail 3 is positioned correctly 25 26 Eppendorf Eporator Operating manual 8 Technical data 8 1 Power supply Mains power connection 100 V to 240 V 10 50 Hz to 60 Hz Automatic adjustment to the voltage Power consumption 20 W Cha
21. rging time lt 10s 8 2 Ambient conditions Environment For indoor use only Ambient temperature 5 C to 40 C Relative humidity 10 to95 Atmospheric pressure 79 5 kPa to 106 kPa Degree of contamination 2 8 3 Weight dimensions Weight 3 2 kg Dimensions Width 19 cm 7 48 in Height 12 5 cm 4 92 in Depth 27 5 cm 10 83 in 8 4 Interfaces USB USB 2 0 RS232 Only for Technical Service 8 5 Operating mode Power on time 10 120 max Eppendorf Eporator Operating manual K Pr Pr 9 Ordering information 9 1 Eporator Order no Order No North Description international America 4309 000 019 4309 000 027 Eppendorf Eporator 9 2 Accessories Order no Order No Description international North America Electroporation cuvettes 50 pieces sterile 4307 000 569 940001005 Gap vvidth 1 mm 100 UL 4307 000 593 940001013 Gap vvidth 2 mm 400 UL 4307 000 623 940001021 Gap vvidth 4 mm 800 UL Cuvette stand 4308 078 006 940001102 for 16 cuvettes 4309 900 010 4309 900 010 Operating Manual Eppendorf Eporator 28 Eppendorf Eporator Operating manual 10 Transport storage and disposal 10 1 Storage Air temperature Rel humidity Air pressure In transport packaging 25 to 55 C 10 to 95 70 to 106 kPa VVithout transport 5
22. s sige dd d en dd N v B aes aie 18 5 4 Regeneration of the cells aa 18 5 5 Determination of the transformation efficiencyY aaa 18 Eppendorf Eporator Operating manual 5 6 Programs sen ee 0 18 5 6 1 Loading program nennen nennen 18 s ndrrue Gerdi enden brag a 19 5 7 3 arena desde nd ai Adare doh tdi EH 20 5 8 Exporting data cen d d shu is n se ee E ha os 20 6 Troubleshooting enis spa ee NEEN va mann 22 6 1 General errors anne de peddle tienes so 22 6 2 EITOFIMESSAGBS u 7 77 11 1 23 6 2 1 Operational emer er hinein 23 T eu ao SEELEN E E O 90 24 7 Maintenance 1 ns kama padi dd 25 KS We ul Le 1 EE 25 7 2 711 0 25 8 Technical data cica 2 2 nen 26 8 1 POWer SUPPLY da eses ee 26 8 2 Ambient conditions aa 26 8 3 Weight dimensions dd EES 26 8 4 3 3 5 1 1 1 111 26 8 5 Op ratingjimodes is aa 2 2 dd ey awh Gan Rs Deren g 26 9 Ordering
23. ter the device gt Only use electroporation cuvettes with lids 16 Eppendorf Eporator Operating manual 6 Place the electroporation cuvette in the cuvette mount with the plastic nose pointed towards the back Fig 4 on p 17 Fig 4 Insert the electroporation cuvette Operate the cuvette carrier with two hands to prevent the cuvette carrier from tipping 7 Slide the carrier in the cuvette shaft of the Eporator until it engages Fig 5 on p 17 Fig 5 Insert the cuvette carrier In the display the actual parameters of the last run disappear and a cuvette symbol appears in the lower line 17 18 Eppendorf Eporator Operating manual 5 3 3 Electroporation O 1 Set a voltage between 200 V and 2 500 V using the arrow keys After switching on the device the last set voltage is always displayed The most frequently used voltages can be saved and accessed using the program keys see Programs on p 18 2 Press the Start key to start the electroporation process Charge and a progress bar appear in the display during loading Asignal tone sounds after the discharge After the electroporation the actual voltage act the discharge time of the performed electroporation and a voltage symbol appear in the display 3 Remove cuvette holder from the device The cuvette symbol and voltage symbol disappear 4 Remove electroporation cuvette from the cuvett
24. tion for the Eporator according to the following criteria Mains connection 230 V 120 V 100 V according to the name plate This is located on the rear side of the device At least 10 cm away from adjacent devices and walls Solid bench with stable horizontal and even work surface 4 3 Installing the instrument WARNING Danger from using an incorrect power supply gt Only connect the device to voltage supplies which correspond with the electrical requirements on the name plate gt Only use sockets with a protective grounding conductor and a suitable mains cable 1 Connect the provided mains cable to the mains connection socket of the Eporator and the power supply 2 Switch on the Eporator at the mains power switch Eppendorf Eporator Operating manual 5 Operation 5 1 Before using the Eporator for the first time familiarize yourself with the operating controls and the Control element overview display Fig 2 Operating controls 1 P1 program key with control LED Press load voltage value Hold down gt 2 s save current voltage value P2 program key with control LED Press load voltage value Hold down gt 2 s save current voltage value 3 Press the Start key Arrow keys Start electroporation Set voltage 5 Exit key menu enter key Exit the menu Select menu parameter 13 14 Eppendorf Eporator Operating manual
25. to 45 C 10 to 95 70 to 106 kPa packaging 10 2 Decontamination before shipment If you are shipping the device to the authorized Technical Service for repairs or to your authorized dealer for disposal please note the follovving VVARNINGI Risk to health from contaminated device 1 Follow the instructions in the decontamination certificate This can be found PDF file on our homepage www eppendorf com decontamination 2 Decontaminate all the parts you want to dispatch 3 Enclose the fully completed decontamination certificate for returned goods incl the serial number of the device vvith the dispatch 10 3 Transport gt Only transport the device in the original packaging Air temperature Rel humidity Air pressure General 25 C to 60 C 10 to 95 Vo 30 kPa to 106 kPa transportation Air freight 40 C to 55 C 10 to 95 Vo 30 kPa to 106 kPa Eppendorf Eporator Operating manual 10 4 Disposal In case the product is to be disposed of the relevant legal regulations are to be observed Information on the disposal of electrical and electronic devices in the European Community Within the European Community the disposal of electrical devices is regulated by national regulations based on EU Directive 2002 96 EC pertaining to waste electrical and electronic equipment WEEE According to these regulations any devices supplied after August 13 2005
26. xcel The data sets of an electroporation contain the following information Sample number sample of the experiment For each experiment the device automatically assigns a four digit sample number that is counted from 0001 upwards Eppendorf Eporator Operating manual Date date of the experiment Time time of the experiment Set voltage set of the experiment Voltage that vvas selected for the corresponding experiment using the arrovv keys Actual voltage act of the experiment Voltage that vvas actually applied to the electroporation cuvette in the corresponding experiment Time constant of the discharge curve tc of the experiment Time constant of the discharge curve of the corresponding experiment Software version sw of the device Serial number serial no of the device Pate and time can be specified in the enhanced settings see p 20 Insert USB stick 1 Insert a standard USB stick in the USB port at the rear of the device see Main illustration on p 7 Open menu 2 Press menu enter key The menu is displayed and cursor blinks in the export display exp OFF Activate parameter O 3 Press one of the arrow keys The selection item exp ON is displayed Export data 4 Press the Start key The data transfer is started The main screen is displayed after the export is completed 21 22 Eppendorf Eporator Operating manual 6 Troub
27. y removing salts from the DNA solution and cell preparations Generally the lowest possible ionic solution that cells can withstand is preferred 5 2 3 Growth and preparation of cells Growth phase of cells For optimum electroporation efficiency use bacterial e g E coli or yeast cells in the exponential growth phase Preparation of cells Thoroughly wash the cells in order to remove the growth medium that affects electrocompetence Concentration of cells Use a final concentration of cells of about 1 3 x 1011 cells mL If this value is exceeded the homogenity of the electrical field can be affected Requirements for an electroporation Each bacteria strain and yeast strain has optimal conditions that must be determined empirically These conditions include The cell volume The quantity of the specific plasmid The used field strength E For E coli a field strength of 12 19 kV cm is generally required to reach a maximum transformation efficiency The field strengths are calculated from the voltage used and the distance of the electrodes E V cm 5 2 4 Temperature Cooling electroporation cuvettes Electroporation of microorganisms produces the best results at low temperatures 0 4 C Cool the electroporation cuvettes to 0 C before electroporation Remove residual moisture from the electroporation cuvette before inserting it in the Eporator 15 Eppendorf Eporator Operating manual 5 3 Performing electro
Download Pdf Manuals
Related Search