UVP Hybridizer HB-1000 Скачать руководство пользователя страница 24 | Manualshive

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Hybridization Ovens

Page 24

81-0169-01 Rev Q

Protocol 3:

Chemiluminescence Detection: HRP-Tagged, Alkaline Phosphatase (AP) Probes

or Antibody Conjugates

Equipment

•

Clear plastic cling-wrap or Clear transparent sheet protector

•

UVP EC3 or AC1 Darkroom with Cooled CCD camera

•

Pipette

Reagents for Chemiluminescence

•

ECL™ (or other) detection reagent 1

•

ECL™ (or other if required) detection reagent 2

•

Membrane following hybridization.

Procedure

1.  Mix equal volumes of detection reagents 1 and 2
2.  Pipette the mixture over the surface of the membrane and leave at room temperature for 1 minute.
3.  Drain the sample and wrap it transfer to Darkroom
4.  Close the darkroom and image using CCD camera.

References

Ross, J; Nucleic Acid Hybridization: Essential Techniques; 1998, John Wiley and Sons, ISBN 0-471-97125-1
Sambrook, J; Molecular cloning: a Laboratory manual; 1987, Cold Spring Harbor Laboratory Press; ISBN 0-87969-

309-6
Current Protocols in Molecular Biology; 1987, John Wiley and Sons; ISBN 0-471-50338-X

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Содержание Hybridizer HB-1000

Страница 1: ...ybridizer Multidizer or HybriLinker systems to familiarize yourself with operation procedures _____________________________________________________________________________________ UVP LLC 2066 W 11th Street Upland CA 91786 Tel 909 946 3197 800 452 6788 Fax 909 946 3597 Web Site www uvp com Ultra Violet Products Ltd Unit 1 Trinity Hall Farm Estate Nuffield Road Cambridge CB4 1TG UK Tel 44 0 1223 42...

Страница 2: ...roduction 3 Hybridization Oven Specifications 4 Setup Instructions 6 Operating Instructions 8 Care and Cleaning 12 Changing the UV Wavelength of the Crosslinker 13 Replacement Parts 14 Accessories 16 Maintenance Repair and Technical Assistance 17 Warranty 18 Hybridization Techniques 19 ...

Страница 3: ...ironment is calibrated at 68 0 C At this temperature the accuracy is 0 3 C An internal circulating fan assists in uniformly distributing heating throughout the chamber Two different hybridizer bottle sizes are available for use within the hybridizer using the standard carousel provided with the unit The rotary carousel wheel can hold up to twenty 20 35 x 150 mm bottles or ten 10 35 x 300 mm bottle...

Страница 4: ...arate power supplies for independent operation Part Numbers Volts Hz 95 0031 01 115V 60Hz 95 0031 02 230V 50Hz 95 0031 03 100V 50Hz Specifications Unit Net Weight 60 lbs Dimensions Exterior 17 5W x 24 6H x 15D in 445 x 625 x 381 mm Hybridization Oven Temperature Ambient 10 C to 99 9 C Temperature Stability Inside Oven 0 3 C to 68 C Inside Bottles 0 1 C to 68 C Temperature Uniformity Inside Oven 0 ...

Страница 5: ...ons Exterior 17 5W x 16H x 15D in 445 x 406 x 381 mm Upper Chamber Temperature Ambient 10 C to 80 C Temperature Stability Inside Oven 0 3 C to 68 C Inside Bottles 0 1 C to 68 C Temperature Uniformity Inside Oven 0 1 C to 68 C Inside Bottles 0 1 C to 68 C Heating Element 500 Watts Shaker Tray Speed 54 106 cycles min Orbital Motion Tray 30 rpm 2 degrees angle Carousel Rotation 12 20 rpm Lower Chambe...

Страница 6: ...ntly push bottle into clips see Pic 3 Insert one side at a time for large bottles NOTE ALWAYS USE AN EVEN NUMBER OF BOTTLES AND LOAD BOTTLES OPPOSITE EACH OTHER TO BALANCE THE BOTTLE HOLDER THIS PREVENTS EXCESSIVE WEAR OF THE BEARINGS AND DRIVE MOTOR Bottle Care Hybridization bottles are made of lead free borosilicate glass which provides superior long term reliability Each bottle includes a polyp...

Страница 7: ...he inside of the coupling sleeve engage the slots in the motor drive shaft of the chamber Rotate the sleeve counter clockwise 30 until locked in place To remove the tray reverse the process To install the Orbital Motion Tray open the chamber door remove the carousel connect the power cable to the orbital tray and to the power port inside the unit and place the tray in the oven so the switch is acc...

Страница 8: ... panel To adjust the temperature setting to the necessary degree press either the UP or DOWN buttons below the temperature control display to raise or lower the displayed degree displayed in degrees Celsius This setting is adjustable from ambient 10 C to 99 9 C See Setting the Temperature Controls for more detail To turn on the motor for the bottle holder press the motor control power switch to th...

Страница 9: ...e memory when the display reverts to the normal mode Calibrating the Temperature Reading The hybridizer is calibrated at the UVP factory UVP recommends temperature recalibration be performed at the UVP factory as recalibration by the user may void the warranty Obtain an RGA Returned Goods Authorization number from UVP prior to returning a unit to the factory Call UVP s customer service department ...

Страница 10: ...formation label Note For 230V models or those requiring special power cord connectors ensure that the proper configuration of the male connector or the plug has been properly connected to the power cord 4 Turn the On Off switch to the On position NOTE WHEN TURNED ON THE CROSSLINKER DEFAULTS TO THE LAST USED UV EXPOSURE SETTING 5 The last UV exposure setting will now be displayed on the LCD and the...

Страница 11: ... displayed must be multiplied by 100 If the settings are correct press ENTER on the touchpad Press START on the touchpad After a slight delay to energize the UV tubes the LCD will begin to count down The unit will stop automatically at the end of the exposure cycle and will beep five times Exposure is now complete User Set UV Time Exposure Setting your own UV time exposure can be achieved as follo...

Страница 12: ...e free operation To ensure correct operation Wipe ANY water from inside and outside the unit with a soft cloth or sponge Use soap and water with a soft cloth or sponge to clean the unit Do not allow chemicals to remain on the unit s surfaces Never clean the unit with abrasive pads or cleaners Never clean the unit with acetone or chloroform Clean the UV sensor regularly with a soft cloth and alcoho...

Страница 13: ... the Crosslinker by holding down STOP on the tactile touchpad and then turning the power to the unit ON 6 A sequence of tones will be heard from the Crosslinker and a 180 second count will display on the LCD once the calibration procedure has begun 7 No calibration occurs during this 180 second period This time sequence allows for the UV tube sources to warm up and stabilize 8 Upon completion of t...

Страница 14: ...mpression Latch 1 22 0110 01 Power Cord 115V 1 58 0085 01 Power Cord 230V UK 1 58 0085 02 Power Cord 230V Euro 1 58 0085 03 Replacement Parts for HL 2000 HybriLinker Crosslinker DESCRIPTION QTY PART NO Borosilicate Glass Bottle with Cap and O Ring Large 35 x 300mm 1 07 0194 02 Cap and O Ring for all Bottle sizes 1 88 0004 01 Bottle Holder Carousel 2 76 0070 01 Carousel Drive Shaft Rod 1 76 0064 03...

Страница 15: ... Drip Tray for Lower Chamber 2 10 0292 02 Hood Acrylic for Upper Chamber 1 10 0317 01 Roller Assembly 2 76 0309 01 Roller Insert and Cam Assembly 1 76 0309 02 Reciprocating Shaker Tray 10 W x 6 D 1 98 0067 02 Drip Tray for Upper Chamber 1 10 0318 01 Fuses 15 Amp 250V 5 x 20 mm Slo Blo 2 56 0044 04 for all voltages Main Switch 4 53 0165 01 Knob Motor Control 2 72 0030 01 Adjustable Rubber Foot 4 72...

Страница 16: ...for HM 4000 upper chamber 76 0089 01 Capacity 4 35 x 300mm 8 35 x 150mm or 8 50mm Conical Other Equipment Rocker Tray for all models 12 5W x 10D in 318 x 254 mm 98 0067 03 Removable plate is manufactured of stainless steel Orbital Motion Tray for all models 10 3 W x 8 2 D 98 0067 04 For use of the orbital tray outside the ovens order the following Orbital Motion Tray external stand alone 100 115V ...

Страница 17: ...ervice at the following numbers UVP LLC In the US toll free 800 452 6788 or 909 946 3197 Fax 909 946 3597 2066 W 11th Street Upland CA 91786 USA Ultra Violet Products Ltd In Europe UK call 44 0 1223 420022 Fax 44 0 1223 420561 Ultra Violet Products Ltd Unit 1 Trinity Hall Farm Estate Nuffield Road Cambridge CB4 1TG UK NOTE A Returned Goods Authorization RGA number must be obtained from UVP Custome...

Страница 18: ...ess for any purpose and all other warranties expressed or implied except those expressly set forth herein are deemed waived and excluded UVP s duty under the warranty is limited to replacement and or repair of the defective part at the option of UVP LLC UVP shall not be held liable for any expenses or damages incurred by the purchaser except expressly set forth herein and in no event shall UVP be ...

Страница 19: ...ms of Nucleic Hybridization Hybridization occurs with a process called nucleation whereby the two separate nucleic acid strands come into close proximity of each other A duplex region is formed where a minimum of three bases of one strand complements to those on the second strand If the remainder of the strands are complementary the two strands will anneal or zipper together very quickly The rate ...

Страница 20: ...olution Unacceptable high background Use less probe Hybridize at lower salt higher temperature Wash with lower salt higher temperature Incubate with very low salt change nuclease solution Use a smaller probe or a different probe Clean probe of contaminants prior to use 4 Aqueous or Denaturing Hybridization Buffer If hybridization takes place in an aqueous salt environment of 0 8 to 1 2M salt the T...

Страница 21: ...eaction buffer 10X 50mM MgCl2 10mM 2 Mercaptoethanol 500 mM Tris HCl pH 7 5 Tagged nucleotide fluorescene 11 dUTP Template DNA in water 5ng mL Procedure 1 Pipette 10 mL of template DNA plus 10 mL of water into a microcentrifuge tube and cap tightly Cover cap with a cap lock or bend a paper clip in half and secure over the microcentrifuge tube 2 Place the tube into the boiling water bath for 5 minu...

Страница 22: ... Bath Bucket of ice Gloves Plexiglas shield UVP Minidizer HybriCycler or Hybridizer Hybridization Oven Procedure 1 Add 15 mL of prehybridization solution to each hybridization bottle containing the blot Remove bubbles between the glass and blot Cap the blots and close the Hybridizer 2 Incubate the blot at 42 C for 1 hour 3 Remove prehybridization solution and replace with 10 mL of hybridization so...

Страница 23: ... wash dry blot on filter paper for 10 minutes This is a good time to quickly pass your hand held radioisotope reader beta or gamma counter over your blot to get a general idea as to the exposure time you will need for the x ray film Hot blots are 20 minutes to 2 hours Not so hot blots can be left overnight 8 hours 6 Tape the blot to a cardboard backing 7 Cover with plastic wrap to prevent the blot...

Страница 24: ...brane following hybridization Procedure 1 Mix equal volumes of detection reagents 1 and 2 2 Pipette the mixture over the surface of the membrane and leave at room temperature for 1 minute 3 Drain the sample and wrap it transfer to Darkroom 4 Close the darkroom and image using CCD camera References Ross J Nucleic Acid Hybridization Essential Techniques 1998 John Wiley and Sons ISBN 0 471 97125 1 Sa...

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