Imaging ChemiDoc™ MP Imaging System Confidence in every step of your imaging workflow. ChemiDoc MP Imaging System Because results always matter. Bring a new level of capability and efficiency to your experiments with an imaging system designed for multitasking. The ChemiDoc MP Imager is a unique imaging system that enables stain-free operation and lets you visualize proteins at every stage of your blotting experiment. Its flexibility and sensitivity are complemented by simple, intuitive operation that integrates seamlessly into your workflow. bio-rad.com Superior Sensitivity Get quantitative, reproducible data without relying on outmoded film processes. The ChemiDoc MP Imaging System offers advanced detection technology that creates optimal exposure for even the faintest bands. Rely on it for fast, super-sensitive chemiluminescence and fluorescence detection and for colorimetric gel and blot documentation. Signal-to-noise ratio of transferrin. 10 sec exposure. 180.0 170 160.0 140.0 Signal-to-noise ratio Fig. 1. Sensitivity comparison of the ChemiDoc MP System versus X-ray film using blots of serial dilution of transferrin. A, the ChemiDoc Imager delivers superior dynamic range and comparable limit of detection to film. B, a 10-second exposure on film reveals a more limited dynamic range than the ChemiDoc MP System. Saturated pixels are highlighted in red. 131 121 120.0 114 95 100.0 93 85 80 60 60 40 48 30 29 28 28 27 27 26 20 0 30 20 15 ChemiDoc MP Imaging System 10 7.5 5.0 Film Fig. 1B Film Sample load, ng ChemiDoc MP Imaging System 24 3.8 2.5 Sample load, ng Fig. 1A Bio-Rad Laboratories, Inc. 58 47 34 23 26 22 1.9 1.3 0.63 32 19 19 0.31 0.15 17 14 0.08 SUPERIOR SENSITIVITY Fig. 2. A calibrated luminescent target was used to test light collection efficiency and determine overall system sensitivity. A, data from lower limits of detection are graphed to show overall signal-to-noise ratio. The ChemiDoc MP System delivers top-of-the-class chemiluminescence sensitivity against leading multiplexing imagers on the market. B, images of lower limits of detection from a calibrated luminescent target. The ChemiDoc MP System delivers excellent image quality and limit of detection. Signal-to-noise ratio of LEDs from a calibrated luminescent test plate. 5 min exposure. 100 90 90.7 89.8 80 Signal-to-noise ratio 70 60 50 40 30 20 22.1 21.6 20.3 20.2 10 5.9 6.6 6.2 7.5 1.9 2.0 3.6 4.1 0 0.7 0.9 12 34 LED well ChemiDoc MP Imaging System Competitor 1 Competitor 2 Competitor 3 Fig. 2A Fig. 2B ChemiDoc MP Competitor 2 12 34 Competitor 1 12 34 Competitor 3 ChemiDoc MP Imaging System bio-rad.com Exceptional Image Quality With patented focus calibration technology, images are always in focus at any zoom level. Exceptional dynamic range enables visualization of faint and intense bands on same blot or gel. With Image Lab™ Software you can edit and analyze images on the spot without exporting to other programs. Fig. 3. 1-D Coomassie-stained gel. Fig. 4. Chemiluminescent ELISA arrays. Quansys Biosciences Q-Plex Array has 16 distinct capture antibodies bound to each well of a 96-well plate. Fig. 5. Fluorescent multiplex blot with DyLight 488, DyLight 549, and DyLight 649 conjugates. Fig. 3 Fig. 5 Bio-Rad Laboratories, Inc. Fig. 4 EXCEPTIONAL IMAGE QUALITY Minimal cross-talk between blue and green channels. Fluors detected in green channel Cross-talk Correct Fluors detected in blue channel Correct Cross-talk FAM A488 Cy3 A546 FAM A488 Cy3 A546 2.3 1.1% 100% 100% 100% 100% 0.1% 0.0% FAM A488 Cy3 A546 FAM A488 Cy3 A546 12.4% 7.6% 100% 100% 100% 100% 0.2% 0.0% FAM A488 Cy3 A546 FAM A488 Cy3 A546 32.5% 21.3% 100% 100% 100% 100% 9.7% 6.1% ChemiDoc MP Competitor 1 Fig. 6A. Determination of cross-talk using same multiplex gel. Blue fluors (FAM and Alexa Fluor 488) are detected in the green channel and green fluors (Cy3 and Alexa Fluor 546) in the blue channel using the ChemiDoc MP System and a competitor’s system. For competitor 1, the bold percentage values indicate high cross-talk. Fig. 6A Competitor 2 Fig. 6B Cross-talk FAM A488 Correct Cy3 A546 Correct FAM A488 Fig. 6B. Visualization of cross-talk signals recorded with a second competitor’s instrument. The green and blue channel images show high cross-talk values and are indicated in bold. The blue fluors (FAM and Alexa Fluor 488) are detected in the green channel and the green fluors (Cy3 and Alexa Fluor 546) are detected in the blue channel. Cross-talk Cy3 A546 ChemiDoc MP Imaging System bio-rad.com Unmatched Application Versatility The ChemiDoc MP Imager is the only system you need when your experiments include a variety of sample types or require different detection methods. It is the perfect imager to accompany your protein and DNA electrophoresis runs as well as your western blotting experiments. And it delivers quantitative, reproducible results every time. Fig. 7. Multiplex image file allows multichannel and individual channel view. Image of multiplexed fluorescent blot with Alexa Fluor 488, Alexa Fluor 555, and Alexa Fluor 647. Bio-Rad Laboratories, Inc. UNMATCHED APPLICATION VERSATILITY Fig. 8. Multiple applications of the ChemiDoc MP Imager. DIGE Criterion™ Gel EtBr-stained wide mini ReadyAgarose™ Gel GelRed-stained mini ReadyAgarose Gel Silver-stained 2-D Criterion Gel Coomassie-stained Mini-PROTEAN® TGX™ Gel SYPRO Ruby-stained large-format 2-D gel Flamingo™-stained Mini-PROTEAN TGX Gel Criterion Stain Free™ TGX Gel SYBR® Green-stained wide mini ReadyAgarose Gel ChemiDoc MP Imaging System bio-rad.com Ease of Use The ChemiDoc MP System is designed for productivity. With little or no training, users can acquire publication-quality images in seconds. The system is precalibrated to provide the precise focus for any zoom setting or sample height; automated hands-free operation ensures consistent, reproducible, and high-throughput performance. Fig. 9A. The application selected automatically determines the excitation sources and emission filters. Fig. 9B. The ChemiDoc MP System is always in focus at any zoom level — no more out-of-focus gel or blot images. Bio-Rad Laboratories, Inc. EASE OF USE Autosetup Autofocus Autoexposure Autooverlay Autoanalysis Autoreports Autoprint ChemiDoc MP Imaging System bio-rad.com Stain-Free Enabled Stain-free technology, offered only by Bio-Rad, eliminates extra steps and unproductive delays in your western blotting experiments. Stain-free technology is the keystone of the Bio-Rad® V3 Western Workflow™, a portfolio of products that enables researchers to visualize, verify, and validate results at each step of their western blotting experiment. V3 Stain-free gel Stain-free blot Total protein quantification Protein of interest probed with Alexa Fluor 649 Visualize The ChemiDoc MP System will activate and provide immediate visualization of protein separation in all lanes with stain-free gels before blot transfer. Bio-Rad Laboratories, Inc. Verify The ChemiDoc MP System paired with stain-free technology enables instant verification of protein transfer before blot detection. Validate Flexible Image Lab Software tools normalize for protein of interest using total protein stain from stain-free blot and provide quantitative blot results. STAIN-FREE ENABLED V3 Western Workflow The V3 Western Workflow streamlines the western blotting protocol, incorporating stain-free in-gel chemistry to allow rapid fluorescent detection of proteins for gels and blots as well as the use of total protein normalization as a loading control. This improved workflow saves time and increases accuracy and reliability throughout the western blotting process. Workflow Benefit Separate Proteins Run gels in as little as 15 min Speed with flexibility: TGX Stain-Free™ Gel chemistry available in precast and handcast formats 1 ■■ Visualize Protein Separation Visualize separation for all lanes in 1 min 2 ■■ Coomassie-like performance with no background variability and no staining/destaining Stain-free image of pretransferred gel Transfer Efficient and uniform protein transfer in as little as 3 min 3 ■■ Throughput: transfer 4 mini gels at once Verify Transfer Efficiency Quickly assess transfer efficiency 4 ■■ Verify quality of transfer for all lanes in 2 min Stain-free image of blot Antibody Incubation and Blot Detection ~5 hr Validate Western Blot Data by Normalization and Analysis Use stain-free blot image as total protein loading control ■■ 5 No need to strip and reprobe Use the entire protein sample in one lane (no need to rely on housekeeping proteins) ■■ Detect protein of interest Normalize protein of interest with stain-free image of blot from step 4 ■■ Reliable and accurate quantitation ChemiDoc MP Imaging System bio-rad.com Total Protein Normalization Stain-free gel chemistry makes it possible to use total protein levels as a loading control rather than the housekeeping proteins used in traditional western blotting protocols. This negates the need to strip and reprobe the blot and avoids the attendant errors that can be introduced in this step. 1 Relative signal intensity of protein bands Using total protein normalization produces a much greater linear dynamic range for measuring target protein levels. Housekeeping proteins such as b-actin, b-tubulin, or GAPDH are often very abundant in biological samples, which results in their signal being oversaturated compared to target proteins. Normalizing results to a total protein measurement corrects this problem, allowing a meaningful comparison even with low-abundance targets, and leads to far greater quantitative accuracy in measuring proteins of interest. MORE RELIABLE to quantitate the protein load 6 Stain-free total protein versus housekeeping proteins 5 b-actin GAPDH 4 b-tubulin Quantitative response 3 2 1 0 0 2 Linearity comparison of stain-free total protein measurement and immunodetection of three housekeeping proteins in 10–50 µg of HeLa cell lysate. Stain-free 10 20 30 40 HeLa lysate load, µg 50 60 AVOID FALSE FINDINGS caused by housekeeping protein signal saturation 50 HeLa lysate load, µg 40 30 20 False Findings: Kinase Protein Levels Normalized against Actin 10 2.5 Akt Erk b-actin Signal intensity (artificial units) MEK 2 1.5 1 0.5 0 10 HeLa lysate load, µg 40 Truth Revealed: Kinase Protein Levels Normalized against Total Protein Stain-free blot image Signal intensity (artificial units) 2.5 2 1.5 1 0.5 0 10 HeLa lysate load, µg Bio-Rad Laboratories, Inc. 40 Comparison of kinase protein levels normalized by stain-free total protein or actin loading controls. 10–50 µg of HeLa cell lysate from the same sample were loaded onto a stain-free gel to probe MEK ( ■ ), Akt ( ■ ), and Erk ( ■ ). There should be no changes in the kinase protein levels when data are normalized. Expected value ( ). TOTAL PROTEIN NORMALIZATION Research Papers Using Stain-Free Technology as Loading Controls in Western Blots Methodology: How stain-free total protein normalization improves western blotting Stain-free total protein staining is a superior loading control to b-actin for western blots. Gilda JE, Gomes AV (2013). Anal Biochem 440, 186–188. V3 stain-free workflow for a practical, convenient, and reliable total protein loading control in Western blotting. Posch A, Kohn J, Oh K, Hammond M, Liu N (2013). J Vis Exp, video ID 50948. Cancer Biology BRCA2 is epistatic to the RAD51 paralogs in response to DNA damage. Jensen RB, Ozes A, Kim T, Estep A, Kowalczykowski SC (2013). DNA Repair (Amst) 12, 306–311. Differential network analysis applied to preoperative breast cancer chemotherapy response. Warsow G, Struckmann S, Kerkhoff C, Reimer T, Engel N, Fuellen G (2013). PLoS One, article ID 0081784. Neurobiology Proteomic analysis of gliosomes from mouse brain: identification and investigation of glial membrane proteins. Carney KE, Milanese M, van Nierop P, Li KW, Oliet SH, Smit AB, Bonanno G, Verheijen MH (2014). J Proteome Res [published online ahead of print Nov 4, 2014]. Accessed Nov 10, 2014. Hippocampal extracellular matrix levels and stochasticity in synaptic protein expression increase with age and are associated with age-dependent cognitive decline. Vegh MJ, Rausell A, Loos M, Heldring CM, Jurkowski W, van Nierop P, Paliukovich I, Li KW, Del Sol A, Smit AB, Spijker S, van Kesteren RE (2014). Mol Cell Proteomics 13, 2975–2985. A trifluoromethyl analogue of celecoxib exerts beneficial effects in neuroinflammation. Penta AD, Chiba A, Alloza I, Wyssenbach A, Yamamura T, Villoslada P, Miyake S, Vandenbroeck K (2013). PLoS One 8, e83119. Skeletal Muscle Identification of the immunoproteasome as a novel regulator of skeletal muscle differentiation. Cui Z, Hwang SM, Gomes AV (2014). Mol Cell Biol 34, 96–109. Immunology Activation of the P2X7 receptor induces the rapid shedding of CD23 from human and murine B cells. Pupovac A, Geraghty NJ, Watson D, Sluyter R (2014). Immunol Cell Biol [published online ahead of print Aug 26, 2014]. Accessed Nov 10, 2014. Metabolomics Quantification of protein copy number in yeast: The NAD+ metabolome. Mei SC and Brenner C (2014). PLoS One 9, e106496. ChemiDoc MP Imaging System bio-rad.com Instrument Tour Optimized emission and excitation filters for discrete detection of multiplexed visible fluorescence. Red Automatic detection of red fluorescence using dyes such as Cy5, Cy5.5, Alexa Fluor 647, Alexa Fluor 680, DyLight 649, DyLight 680, IRDye 680. Green Automatic detection of green fluorescence using dyes such as Cy3, Flamingo, Krypton, Pro-Q Diamond, Alexa Fluor 546, DyLight 549, Rhodamine. Blue Automatic detection of blue fluorescence using dyes such as Cy2, Coomassie Fluor Orange, Alexa Fluor 488, DyLight 488, Pro-Q Emerald 488, Qdot 523, Qdot 605, Qdot 625, Qdot 705. Bio-Rad Laboratories, Inc. INSTRUMENT TOUR Cooled CCD with autofocus for all zoom levels Unmatched sensitivity and image quality Multiplexed fluorescent imaging Multicolor LEDs optimized for quantitative western blot imaging Sample tray with UV, white, and blue imaging Flexible transillumination source for easy and uniform imaging of gels and blots 6-position automated filter wheel Broad application flexibility Touch-pad operation For positioning and band excision Go to bio-rad.com/chemidocmp to see the complete ChemiDoc MP video. ChemiDoc MP Imaging System Specifications Ordering Information Automation Capabilities Workflow automated selectionApplication driven; user selected or recalled by a protocol Catalog # Workflow automated executionControlled by a protocol via application-specific setup for image area, illumination source, filter, analysis, and reporting Workflow reproducibility100% repeatability via recallable protocols; from image capture to quantitative analysis and reports Autofocus (patent pending)Precalibrated focus for any zoom setting or sample height Image flat fielding* Dynamic; precalibrated and optimized for every application Autoexposure2 user-defined modes (intense or faint bands) Hardware specifications Maximum sample size (L x W) 28 x 36 cm Maximum image area (L x W) 26 x 35 cm Maximum image area for 25 x 26 cm standard UV-excited gels (L x W) Excitation sourceTrans-UV (302 nm included; 254 nm and 365 nm available as options) and epi-white; optional trans-white conversion screen, XcitaBlue™ Conversion Screen, and epi-red, -green, and -blue LED modules Illumination control8 modes available. Trans-UV, epi-white, and no illumination for chemiluminescence are standard; epi-red, epi-green, epi-blue, transwhite, and XcitaBlue Conversion Screens are optional Description 170-8280 ChemiDoc MP Imaging System with Image Lab Software, PC or Mac, includes darkroom, UV transilluminator, epi-white illumination, camera, power supply, cables, Image Lab Software Accessories 170-8283 Red LED Molecular Kit, pkg of 2 epi-red LED modules and 1 red emission filter, for use with applications requiring red fluorophore detection 170-8284 Green LED Molecular Kit, pkg of 2 epi-green LED modules and 1 green emission filter, for use with applications requiring green fluorophore detection 170-8285 Blue LED Molecular Kit, pkg of 2 epi-blue LED modules and 1 blue emission filter, for use with applications requiring blue fluorophore detection 170-8289 White Light Conversion Screen, for viewing Coomassie, silver stain, and other colorimetric gel stains 170-8182 XcitaBlue Conversion Screen, includes view goggles; blue conversion screen for viewing SYBR® Green, SYBR® Safe, GFP, Flamingo, and other fluorescent gel stains 170-8183 XcitaBlue Conversion Screen and Filter, includes view goggles and SYBR® Safe Filter (170-8075, 560DF50); blue conversion screen for viewing SYBR® Green, SYBR® Safe, and other fluorescent gel stains 170-8083 Filter 520DF30 62 mm, for SYBR® Green/GFP/SYBR® Gold/fluorescein 170-8098 254 nm UV Lamps, pkg of 6 170-6887 365 nm UV Lamps, pkg of 6 170-8097 Standard 302 nm UV Lamps, pkg of 6 170-8089 Mitsubishi Thermal Printer 170-7581 Mitsubishi Thermal Printer Paper, 4 rolls 170-8184 Gel Alignment Templates, pkg of 3 Detector Supercooled CCD Image resolution 4 megapixels Software 170-9690** Image Lab Software, PC or Mac, for automated image capture, optimization, and 1-D data analysis Pixel size (H x V) 6.45 x 6.45 µm ** Included with the imaging system. Cooling system Peltier Camera cooling temperature –30°C absolute and regulated Filter holder6 positions (5 for filters, 1 without filter for chemiluminescence) Emission filters 1 included (standard), 4 optional Dynamic range >4.0 orders of magnitude Pixel density (gray levels) 65,535 Instrument size (L x W x H) 36 x 60 x 96 cm Instrument weight 32 kg Operating Ranges Operating voltage 110/115/230 VAC nominal Operating temperature 10–28°C (21°C recommended) Operating humidity <70% noncondensing * U.S. patent 5,951,838. Scan this QR code to learn more about the ChemiDoc MP Imaging System, or visit biorad.com/chemidocmp. Alexa Fluor, Coomassie Fluor Orange, Pro-Q, Qdot, SYBR, and SYPRO are trademarks of Life Technologies Corporation. Coomassie is a trademark of BASF Aktiengesellschaft. Cy is a trademark of GE Healthcare Group Companies. DyLight and Krypton are trademarks of Thermo Fisher Scientific. GelRed is a trademark of Biotium, Inc. IRDye is a trademark of LI-COR Biosciences. Mac is a trademark of Apple Inc. Mitsubishi is a trademark of Mitsubishi Companies. Q-Plex is a trademark of Quansys Biosciences. Bio-Rad Laboratories, Inc. is licensed by Life Technologies Corporation to sell SYPRO products for research use only, under U.S. patent 5,616,502. Bio-Rad Laboratories, Inc. 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