Typhoon™ FLA 9000 biomolecular imager
Contents
1. 473 nm blue LD laser 532 nm green SHG laser 635 nm red LD laser 685 nm optional near IR LD laser and 785 nm optional near IR LD laser 3H 11C 14C 125 18F 32D 33D 355 99MTC and other sources of ionizing radiation 5 orders of magnitude 16 bit 40 x 46 cm 10 25 50 100 200 um and prescan 1000 um IP Phosphorimaging LPB 510LP LPG 575LP LPR 665LP BPB1 530DF20 and BPG1 570DF20 BPFR700 R715 BPFR800 R810 DBR1 530DF20 665LP and DGR1 570DF20 665LP 900 x 400 x 800 mm 97 kg 50 60 Hz 15 C 10 30 C 20 to 70 no condensation 100 240 VAC 10 Approx 0 3 kVA Optimal choice of filter stage laser and PMT Filters are easily accessed and exchanged without tools to attain optimal imaging conditions The system accommodates up to four computer controlled filter positions at any time Custom filters can be easily installed by the user Stages Fig 4 give the correct positioning and stability for optimal imaging of a range of sample types Samples that can be scanned include agarose and polyacrylamide gels membranes DIGE gels radioisotope labeled samples using a phosphorimaging plate as well as microplates and glass slides with the titer plate TP plug in The system can simultaneously scan two DIGE gels each measuring up to 21 5 x 27 5 cm with the Low Fluorescent Glass Plate stage The stages are easily removed from the system for cleaning The system can be equipp2. demonstrate detection in the red wavelength region with a linear signal response Arrow indicates the limit of detection LOD The linear dynamic range DR was 2 1 orders of magnitude and the linearity of the response L was R2 0 995 Scanning is rapid and detection is sensitive for laser induced fluorescence radioisotopic imaging by storage phosphor and digitization A fast 1000 um prescan function gives a rapid overview of the sample for selecting the correct settings At a pixel size of 200 um a 10 x 15 cm sample is scanned in two minutes The system provides a linear signal response over five orders of magnitude This together with digitization of the image with up to 16 bit resolution provides a suitable basis for the precise quantitation of proteins DNA and other labeled molecules Lasers can be exchanged in the field to accommodate new applications and fluorophores The system can house up to four lasers simultaneously from a choice of five laser excitation wavelengths 473 532 635 685 and 785 nm Typhoon FLA 9000 2500 1250 Transferrin 630 310 160 78 39 19 5 pg Sample Transferrin 45000 Membrane Hybond LFP S 40000 Detection Primary antibody ie Rabbit anti human 53000 transferrin 30000 Secondary antibody 25000 Anti rabbit IRDye 680 Imaging Excitation Emission filter 20000 Fa Typhoon 685 nm BPFR700 R715 15000 r LOD 19 5 pg transferrin 10000
3. 73 685 nm laser upgrade 1 28 9610 22 685 nm laser BPFR700 filter PMT2 installation and testing 785 nm laser upgrade 1 28 9610 25 785 nm laser BPFR800 filter PMT2 installation and testing 685 785 nm laser upgrade 1 28 9610 32 685 nm and 785 nm lasers BPFR700 filter PMT2 BPFR800 filter installation and testing Related literature Code no Typhoon FLA 7000 biomolecular imager Data file 28 9610 73 Minimum computer requirement OS Windows XP SP3 32 bit or Windows Vista Business SP1 32 bit RAM more than 1 GB Processor Intel Core 2 Duo processors Hard disk more than 80 GB USB Ports USB 2 0 Optical drive DVD ROM or Super Multi Drive Monitor 1280 x 1024 pixel resolution or higher Please contact your local sales representative for the latest recommended computer configuration 03 2010 28 9610 72 AB 7 For local office contact information visit www gelifesciences com contact www gelifesciences com quantitative_imaging GE Healthcare Bio Sciences AB Bj rkgatan 30 751 84 Uppsala imagination at work GE imagination at work and GE monogram are trademarks of General Electric Company Amersham Cy CyDye DeCyder Deep Purple ECL ECL Advance ECL Plex Ettan Hybond ImageQuant and Typhoon are trademarks of GE Healthcare companies 2 D DIGE 2 D Fluorescence Difference Gel Electrophoresis 2 D DIGE technology is covered by US patent numbers 6 043 025 6 127 134 and 6 426 1
4. IgG Cy3 ECL Plex goat anti mouse IgG Cy5 Imaging Excitation Emission filter Cy3 532 Nnm BPG1 570DF20 Cy5 635 nm LPR 665LP LOD B tubulin in 160 ng CHO cell lysate Cy5 ERK 1 2 in 78 ng CHO cell lysate Cy3 Fig 3 Multiplex detection of endogenous proteins by Amersham ECL Plex Western blotting Tubulin and ERK1 2 were targeted in a dilution series of CHO cell lysate by using mouse anti tubulin and rabbit anti MAP kinase ERK 1 2 primary antibodies Secondary antibodies were ECL Plex anti mouse Cy5 red and anti rabbit Cy3 green Imaging was performed with Typhoon FLA 9000 using separate detection channels Arrows indicate the limits of detection LOD in each detection channel The minimal crosstalk and low background mean that it is possible to reliably quantitate specific signals relative to a housekeeping protein Fig 4 A The Phosphor Stage B Multi Stage C Low Fluorescent Glass Plate Stage and D Fluor Stage are designed to accommodate a variety of sample formats and imaging modes 2 03 2010 28 9610 72 AB Table 1 Typhoon FLA 9000 specifications Detection modes Excitation wavelengths Radioisotopes Dynamic range Bit depth Scanning area Pixel sizes Standard filters Optional filters Dimensions W x H x D Weight Line frequency Temperature Humidity Supply voltage Power consumption Technical features Fluorescence phosphorimaging digitization and chemiluminescence
5. T 70000 Label CyDye DIGE fluor 60000 Cy3 minimal dye vA Gel 12 acrylamide 50000 Tris glycine cae f 2 4 Imaging Excitation Emission filter ey Cy3 532nm BPG1 570DF20 30000 LOD 0 2 ng carbonic anhydrase amp L R 0 998 T 4 DR 3 3 orders of magnitude 10000 ot O 100 200 300 400 500 Amount of carbonic anhydrase ng Fig 8 Different concentrations of carbonic anhydrase were labeled with CyDye DIGE fluor Cy3 minimal dye and subjected to 1 D electrophoresis The gel was imaged with Typhoon FLA 9000 The detection limit LOD was 0 2 ng carbonic anhydrase and the linear DR was 3 3 orders of magnitude Arrow indicates the LOD Chemiluminescence The light detection path is scanned across the sample without illumination The emitted chemiluminescence from each scanned point is collected and transformed to an electrical signal by a PMT The electrical signal is then converted into digital information by A D conversion for image display and analysis Data storage Data are stored either in linear 16 bit grayscale TIFF TIF file format or in square root encoded 16 bit TIFF GEL file format The GEL format encoding provides higher dynamic resolution than TIF at lower signal levels to exploit the low signal detection capability of the phosphorimaging technology Image analysis Designed for seamless data transfer and quantitative gel and blot analysis we provide image analysis software
6. 4 4 pett Gya 4 4 4 4 4 4 4 4 4 Protein Stains Deep Purple Total Protein Stain 4444 SYPRO Ruby NanoOrange solutions Pro Q Diamond phosphorylated proteins H H Jy Pro Q Sapphire 532 Histidine tagged proteins os eared rs ELISA AttoPhos ttit iri Fluorescence Nucleic acids Cy3 and Cy5 et Alexa Fluor 532 and Alexa Fluor 633 Nucleic acid stains Ethidium Bromide post stain Fi 444 ti Vistra Green SYBR Gold SYBR Green and II PicoGreen RiboGreen Chemifluoresence enzyme catalyzed Amersham ECL Plus Western blotting ECF AlkPhos direct ECF HHH Ht h DDAO Phosphate Other applications Cy2 Cy3 Cy5 Fluorescein FAM FITC Alexa Fluor 488 TET HEX ROX TAMRA Green fluoresecent protein Chemiluminescence 4 Amersham ECL Amersham ECL Plus Amersham ECL Advance Superior performance High performance Good performance Acceptable performance Not compatible Ratings are based on overall system performance including model specific features versatility and sensitivity limit of detection Blank fields indicate that data are not available Multiplex experiments e g 2 D DIGE and Amersham ECL Plex cannot be performed on Typhoon F
7. 90 and equivalent patents and patent applications in other countries and exclusively licensed from Carnegie Mellon University DeCyder This release of DeCyder version 2 software is provided by GE Healthcare to the customer under a non exclusive license and is subject to terms and conditions set out in the 2 D Differential Gel Electrophoresis Technology Access Agreement Customer has no rights to copy or duplicate or amend the Software without the prior written approval of GE Healthcare Deep Purple Total Protein Stain Deep Purple Total Protein Stain is exclusively licensed to GE Healthcare from Fluorotechnics Pty Ltd Deep Purple Total Protein Stain may only be used for applications in life science research Deep Purple is covered under a granted patent in New Zealand entitled Fluorescent Compounds patent number 522291 and equivalent patents and patent applications in other countries ECL Advance ECL Advance contains Lumigen TMA 6 substrate and is sold under exclusive license from Lumigen Inc ECL Plus contains Lumigen PS3 substrate and is sold under exclusive license from Lumigen Inc CyDye This product or portions thereof is manufactured under an exclusive license from Carnegie Mellon University under US patent number 5 268 486 and equivalent patents in the US and other countries Cy3 UTP or Cy5 UTP Cy3 5 dCTP or Cy5 5 dCTP Cy3 CTP or Cy5 CTP These products are manufactured for GE Healthcare UK Limited by Perkin Elmer Life Scienc
8. GE Healthcare Data file 28 9610 72 AB Imaging systems software and accessories Tyophoon FLA 9000 biomolecular Imager Typhoon FLA 9000 Fig 1 is a versatile laser scanner for biomolecular imaging applications including sensitive and quantitative measurements of radioisotopic labels by storage phosphor chemifluorescent Western blots and multiplex fluorescence as well as digitization of colorimetric stains e g Coomassie Blue and silver stained gels The system supports both 2 D Difference Gel Electrophoresis DIGE and Amersham ECL Plex Western blotting systems Typhoon FLA 9000 delivers e Versatility image radioisotope multifluorescent chemifluorescent and colorimetric labeled samples e High resolution and quantitation a pixel resolution of up to 10 um and a linear signal response over five orders of magnitude provides precise quantitation in gels blots tissue sections and arrays e High sample throughput A scanning area of 40 x 46 cm enables simultaneous imaging of up to 20 gels or blots measuring 10 x 8 cm in size This facilitates comparisons among blots and reduces workload and waiting time e Flexibility optimized performance for new applications by adapting the system with stages detectors filters and lasers e 2 D DIGE imaging simultaneously image two 2 D DIGE gels for differential expression studies e Visible and infrared fluorescence imaging optional near infrared excit
9. L R 0 990 DR 2 1 orders of magnitude 2000 r4 0 0 1 2 a Amount of transferrin ng Odyssey Transferrin 2500 1250 630 310 160 78 39 19 5 pg ga e a err Sample Transferrin 45000 Membrane Hybond LFP S 40000 Detection Primary antibody a Rabbit anti human 35000 transferrin G 30000 Secondary antibody 55000 Anti rabbit IRDye 680 ue Imaging Excitation Emission filter D 20000 Odyssey 685nm 700 S 15000 i ap a 2 10000 DR 2 1 orders of magnitude 5000 Wes o O 1 2 3 Amount of transferrin ng Fig 6 A two fold dilution series of transferrin starting at 2 5 ng was subjected to Western blotting and detected with a rabbit anti transferrin primary antibody and anti rabbit IRDye 680 secondary antibody Imaging by Typhoon FLA 9000 showed similar performance for detection in the near infrared wavelength region compared to the Odyssey Infrared Imaging System from LI COR Arrows indicate the LOD The experiments were performed at GE Healthcare laboratories according to the manufacturers instructions Optimal detection of chemifluorescent Western blots Laser scanning systems are not optimal for imaging chemiluminescence Typhoon FLA 9000 does however perform well with Amersham ECL Plus by imaging its stable chemifluorescent signal which is emitted upon excitation by the 473 nm laser This provides a means to obtain optimal imaging performan
10. LA 7000 CyDye DIGE Fluors and Amersham ECL Plex conjugates can be imaged in single probe experiments on Typhoon FLA 7000 i e experiments where there is only one dye or conjugate on the gel or membrane 6 03 2010 28 9610 72 AB Ordering information System Quantity Code no Typhoon FLA 9000 1 28 9558 08 Includes 473 nm 532 nm and 635 nm lasers filter tray IP filter LPB filter LPG filter LPR filter BPB1 filter BPG1 filter Fluor Stage Membrane Weight Phosphor Stage Low Fluorescent Glass Plate Stage Multi Stage and TP plug in Fluorescent plate for digitization capture software USB cable mains cables EU and USA User Manual and Getting Started Guide Upgrades and accessories Quantity Code no Fluor Stage Set 1 28 9589 04 Fluor Stage Membrane Weight Multi Stage Set 1 28 9564 19 Multi Stage TP plug in BAS IP MS 2040 E 1 28 9564 74 Phosphorimaging plate 20 x 40 cm multipurpose BAS IP MS 2025 E 1 28 9564 75 Phosphorimaging plate 20 x 25cm multipurpose BAS IP MS 3543 E 1 28 9564 76 Phosphorimaging plate 35 x 43 cm multipurpose BAS IP SR 2040 E 1 28 9564 77 Phosphorimaging plate 20 x 40 cm high resolution BAS IP SR 2025 E 1 28 9564 78 Phosphorimaging plate 20 x 25 cm high resolution BAS IP TR 2040 E 1 28 9564 81 Phosphorimaging plate 20 x 40 cm for Tritium detection BAS IP TR 2025 E 1 28 9564 82 Phosphorimaging plate 20 x 25 cm for Tritium detection FLA Image Eraser il 28 9564
11. ation for imaging IRDye and other infrared dyes Fig 1 Typhoon FLA 9000 is a high performance biomolecular imager for sensitive and quantitative measurements Typhoon FLA 9000 is a variable mode laser scanner with modular access to the optical components and excitation sources Fig 2 providing both versatile and flexible imaging for precise quantitation of proteins nucleic acids and other biomolecules The system provides several imaging modes such as fluorescence filmless autoradiography and digitization of colorimetrically stained gels e g Coomassie Blue and silver stain Chemiluminescence imaging is possible but for detection of low abundance proteins by chemiluminescence the ImageQuant imager series is recommended a us gt i Fig 2 Filters are easily exchanged by the user The system is optimized for 2 D DIGE imaging for differential protein expression studies Amersham ECL Plus chemifluorescent imaging for quantitative protein detection by Western blotting and multifluorescent Amersham ECL Plex imaging for precise quantitation of two or more proteins in the same blot Fig 3 CHO cell lysate n 5000 2500 1250 630 310 160 tota 78 protein Sample CHO cell lysate Membrane Hybond LFP Target protein B tubulin Cy 5 red ERK 1 2 Cy3 green Detection Primary antibodies Mouse anti tubulin and rabbit anti MAP kinase ERK1 2 Secondary antibodies ECL Plex goat anti rabbit
12. ce from a chemifluorescent reagent 03 2010 28 9610 72 AB 3 Ettan DIGE system Ettan DIGE system is an integrated solution for accurate quantitation of changes in protein expression Typhoon FLA 9000 is a fully optimized part of Ettan DIGE system with DeCyder 2D Differential Analysis Software Fig 7 The strengths of Typhoon FLA 9000 high sensitivity and broad dynamic range for measuring low and high abundant proteins in one scan Fig 8 make it highly suited for 2 D DIGE applications enabling the user to detect and accurately quantitate subtle changes in protein expression By generating overlaid multichannel images for each gel with minimal crosstalk Typhoon FLA 9000 exploits the multiplexing potential of CyDye DIGE fluors to remove experimental variation between gels Images are analyzed using DeCyder 2D to accurately and confidently measure very small differences in protein abundance Imaging For the detection of radioactivity fluorescence and chemifluorescence emitted light is collected and transformed to an electrical signal by a photomultiplier tube PMT The electrical signal is then converted into digital information by A D conversion for image display and analysis Detection of radioactivity Samples containing radioactive probes are exposed to a storage phosphor screen Light is emitted from the screen in proportion to the amount of radioactivity in the sample upon laser induced stimulation Fluoresc
13. ed for dual simultaneous fluorescence detection by the addition of a second photomultiplier tube PMT Each PMT is selected for optimal response to the detected emission wavelength The standard bialkali PMT 1 is suitable for phosphorimaging and dyes excited by blue e g Cy2 green e g Cy3 and red e g Cy5 or Alexa Fluor 633 light Fig 5 whereas the optional multialkali PMT 2 is optimal for dyes excited by far red and infrared light such as IRDye680 or IRDye800 In experiments using a secondary antibody labeled with IRDye 680 the performance of Typhoon FLA 9000 in terms of sensitivity dynamic range and linearity of response was shown to be similar to that of the Odyssey Infrared Imaging System from LI COR Fig 6 Transferrin 5000 2500 1250 630 310 160 78 39 19 5 pg Sample Transferrin 50000 Membrane Hybond LFP x Detection Primary antibody gt 40000 Rabbit anti human 30000 transferrin Secondary antibody gt 20000 Anti rabbit Alexa Fluor 633 10000 wo ape aeons 2 5 Imaging Excitation Emission filter Wr Alexa 633 635nm LPR 665LP 0 LOD 39 pg transferrin O 1 2 3 4 5 6 L R 0 995 Amount of transferrin ng DR 2 1 orders of magnitude Fig 5 Atwo fold dilution series of transferrin starting at 5 ng was subjected to Western blotting and detected with a rabbit anti transferrin primary antibody and anti rabbit Alexa Fluor 633 secondary antibody Results
14. ence Upon excitation light is emitted from a fluorescently labeled sample in proportion to the amount of labeled compound in the sample Multiple fluorescent wavelengths can be detected with minimal crosstalk for comparative expression experiments See Table 3 for emission filters Chemifluorescence Upon excitation light is emitted from a fluorescent product generated in an enzyme catalyzed reaction in proportion to the amount of labeled compound in the sample Digitization Excitation light passes through the sample and excites a fluorescent plate The emitted light from the plate passes through the sample again and is collected and converted to an electrical signal The method is suitable for documentation of colorometrically stained gels 4 03 2010 28 9610 72 AB Sample CHO cell lysate expressing monoclonal antibody MAb grown under various culture conditions Gel Large precast DIGE Gel within glass cassette Imaging Excitation Emission filter Cy2 473 NM BPB1 530DF30 Cy3 532 nm BPG1 570DF20 Cy5 635 nm LPR 665LP Fig 7 CHO cells expressing MAb were grown in different culture media 2 D DIGE was used to analyze the expression of host cell proteins as part of efforts to improve MAb process understanding Image analysis was performed using DeCyder 2D software Carbonic anhydrase 409 6 2048 1024 512 256 128 64 3 2 16 08 0 4 0 2 ng Saan lt Sample Carbonic anhydrase
15. es under US patent numbers 5047519 and 5151507 The cyanine dyes in the product are manufactured under an exclusive license from Carnegie Mellon University under US patent numbers 5 268 486 and equivalent patents in the US and other countries The purchase of CyDye products includes a limited license to use the CyDye products for internal research and development but not for any commercial purposes A license to use the CyDye products for commercial purposes is subject to a separate license agreement with GE Healthcare Commercial use shall include 1 Sale lease license or other transfer of the material or any material derived or produced from it 2 Sale lease license or other grant of rights to use this material or any material derived or produced from it 3 Use of this material to perform services for a fee for third parties including contract research and drug screening If you require a commercial license to use this material and do not have one return this material unopened to GE Healthcare Bio Sciences AB Bjorkgatan 30 SE 751 84 Uppsala Sweden and any money paid for the material will be refunded All third party trademarks are the property of their respective owners Odyssey is a trademark of LI COR Biosciences 2009 2010 General Electric Company All rights reserved First published Oct 2009 All goods and services are sold subject to the terms and conditions of sale of the company within GE Healthcare which supplies t
16. for use with Typhoon FLA 9000 Table 2 Table 2 Image analysis software Software Analysis ImageQuant TL 1 D gel electrophoresis dot blots arrays colony counting and user defined gel analysis DeCyder 2D Differential high resolution 2 D DIGE analysis including Extended Data Analysis ImageMaster 2D 2 D gels including Platinum single stain and 2 D DIGE Table 3 Emission filters Filter type Wavelength Detection range nm examples IP BP390 Phosphorimaging LPB 510LP gt 510 Cy2 ECL Plus SYBR Green FAM FITC Alexa Fluor 488 SYPRO Ruby SYPRO Orange GFP BPB1 530DF30 515 to 545 Cy2 DIGE Fluor ECL Plex Cy2 LPG 575LP 575 Cy3 Deep Purple HEX Alexa Fluor 532 and 555 SYPRO Red BPG1 570DF20 560 to 580 Cy3 DIGE Fluor ECL Plex Cy3 LPR 665 LP gt 665 Cy5 Alexa Fluor 633 TOTO 3 DID Cy5 DIGE Fluor ECL Plex Cy5 BPFR700 R715 713 to 726 Alexa Fluor 700 IRDye680 IRDye 700 BPFR800 R810 814 to 826 Alexa Fluor 790 IRDye800 03 2010 28 9610 72 AB 5 Imager performance Typhoon Typhoon Typhoon Typhoon 9400 9410 FLA 9000 Trio Trio FLA 7000 Storage Phosphor 3H u C 125 18F 32P 33D 355 9MTC MER and other sources of ionizing radiation Macroarray radiolabeled t Fluorescence Proteins CyDye DIGE Fluors Cy2 Cy3 4 4 4 4 4 4 4 4 4 Cyd 4 4 4 4 4 4 Fe ECL Plex Fluors Cy2 Cys Fr 4
17. hem A copy of these terms and conditions is available on request Contact your local GE Healthcare representative for the most current information GE Healthcare UK Limited Amersham Place Little Chalfont Buckinghamshire HP7 9NA UK GE Healthcare Europe GmbH Munzinger Strasse 5 D 79111 Freiburg Germany GE Healthcare Bio Sciences Corp 800 Centennial Avenue P O Box 1327 Piscataway NJ 08855 1327 USA GE Healthcare Japan Corporation Sanken Building 3 25 1 Hyakunincho Shinjuku ku Tokyo 169 0073 Japan 28 9610 72 AB 03 2010
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