Widefield (epifluorescence)
DANTE - Olympus IX81 widefield (Porter B047A)
Light Sources:
- Excelitas X-Cite XylisÌý365nm, modelÌýXT720S. The is a high-intensity LED, with selectable intensity in 1% increments, 5-100%. Excitation bandwidth from UV (365nm)Ìýto Near-IR (770nm) with peaks atÌý365, 430, 475, 545, 635, and 735nm, matching optimally with common fluorophores. Total wavelength range 360-770nm. Xylis awarded by Core Facility Assistance Grant (Orth) from CU Research and Innovation Office.
- Halogen for brightfield and phase-contrast.
Standard filter configurations
This system utilizes the finest optical filters from Semrock and Chroma and employs separate excitation and emission wheels for maximum flexibility and customizableÌýoptical settings. This system employsÌýfluor-matched single dichroic beamsplitters, and GFP/RFP dual and Ìýbeam splitters; the multi-band beam splitters allow more rapid multi-channel imaging. These components together provide maximum brightness and fast switching between wavelengths to provide the best results.Ìý
Excitation filters:
- 325-375 (nm) (Chroma 49000); DAPI, Hoechst, Alexa350, 405, etc.
- 426-450 (nm) (Semrock); CFP, cerulean, etc.
- 450-490 (nm) (Chroma 49002); Alexa488, FITC, EGFP, etc.
- 459-481 (nm) (Semrock Brightline GFP/DsRed- 2X2M-C-000); Alexa488, FITC, EGFP, etc.
- 483-512 (nm) (Omega XF104-2); YFP, Alexa532, Venus, etc.
- 560-580(nm) (Chroma 41034); Alexa555, 568, 594, TxRed, mRFP, mCherry, TdTomato, etc.
- 546-566 (nm) (Semrock Brightline GFP/DsRed- 2X2M-C-000); Alexa555, 568, 594, TxRed, mRFP, mCherry, TdTomato, etc.
- 618-648 (nm) (Semrock Brightline 4040A); Alexa647, Cy5, etc.
- 672.5-747.5 (nm) (Chroma 49007); Alexa680/700/750, Cy7, etc.
Emission filters:Ìý
- 435-485 (nm) (Chroma 49000); DAPI, Hoechst, Alexa350, 405, etc.
- 472-494 (nm) (Semrock); CFP, cerulean, etc.
- 502.5-527.5 (Chroma 49002); Alexa488, FITC, EGFP, etc.
- 499-529 (nm) (Semrock Brightline GFP/DsRed- 2X2M-C-000); Alexa488, FITC, EGFP, etc.
- 527.5-562.5 (nm) (Omega XF104-2); YFP, Alexa532, Venus, etc.
- 590-650 (Chroma 41034); Alexa555, 568, 594, TxRed, mRFP, mCherry, TdTomato, etc.
- 580.5-653.5 (nm) (Semrock Brightline GFP/DsRed- 2X2M-C-000); Alexa555, 568, 594, TxRed, mRFP, mCherry, TdTomato, etc.
- 672-712 (nm) (Semrock Brightline 4040A); Alexa647, Cy5, etc.
- 765-855 (nm) (Chroma 49007); Alexa680/700/750, Cy7, etc.
Special Dichroic mirrors
- GFP/DsRed dual-edge dichroic (Semrock BrightLine GFP/DsRed- 2X2M-C-000)
- DAPI/GFP/RFP/CY5/Cy7 penta-edge dichroic (Semrock BrightLine FF409/493/573/652/759-Di01 25x36)
Other imaging
- Brightfield
- Phase-contrast (PH2 20x andÌý40x objectives)
Standard objectives:
- 4x 0.13NA UPlanFl N UIS2 WD 17 (mm)
- 10x 0.40NA UPlanApo Ph1 WD 3.1 (mm)
- 20x 0.70NAÌýUPlanApo Ph2 WD 0.65 (mm)
- 40x 0.75NA UPlanFl N Ph2 WD 0.51 (mm)
- 60x 1.35NA UPlanSapo OIL WD 0.15 (mm)
- 100x 1.40NAÌýUPlanSapo OIL WD 0.13 (mm)
Available upon request:Ìý
- 60X 1.20NAÌýUPLSAPO WATER WD 0.28 (mm)
- Inquire
- . sCMOS. 12/16 bit. Camera awarded by Core Facility Assistance Grant (Orth) from CU Research and Innovation Office.
- 3000 x 3000, 3.76 x 3.76 micron pixels
- Digital binning 2x2, 4x4, sub-array
- HDR mode, 8X analog gain modeÌý
- Peak QE >80% 420 - 520 (nm)
- Wavelength range >30% QE, 360 - 750 (nm)
- High-speed large area montaging via , with advanced stitching in up to 7 wavelengths (inquire).Ìý
- Slide scanning for up toÌý4 standard 25x75mmÌýslides per run;Ìýentirety ofÌýslide(s), multiple areas across all slides,ÌýviaÌýPrior stage insert. IncludesÌýpre-defined settings for 1, 2, 3, or 4 slides in diffferent orientations. Pre-defined slide scanning or largeÌý50x75mm slide format,Ìývia Prior stage insert.
- Intelligent Shading Correction uses proprietary correction that virtually eliminates 'tiling' in large area imaging; User selectable.
- Photobleaching prevention mode pulses LED at chosen exposure time only when focusing or moving your sample, minimizing light exposure; User selectable.
- Stage and well-plate navigator interface withÌýpre-programmed plate formats for 6, 12, 24, 48, 96 wells and more.Ìý
- Advanced 2D and 3D deconvolution;ÌýGraphical Experiment Manager GUI; Debluring;ÌýFluorescence unmixing; Image analysis. More!!
- Save your own custom settings including optical configurations, scan areas, acquisition settings,Ìýfor routine precision.
- Live cell imaging with InVivo Sci. stage-top humidified chamber, ; 1x35mm, multi-well plates, pre-mixed 5% CO2, balance air (please contact Dr. Orth); Oustanding phase contrast (20x and 40x); precise repeatability over time viaÌýÌýZ-drive; FRET (CFP --> YFP);ÌýGFP/mCherry dual dichroic or Blue/Green/Red/FarRed/Near-IR penta beam splitters provide for rapid multicolor live sample and large area imaging.
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Software updated awarded by Core Facility Assistance Grant (Orth) from CU Research and Innovation Office.
Contact Director, James.Orth@olorado.edu
PEPPER - Nikon TiU widefield with RGB color brightfield (Porter B047A)
Light Sources:
- Halogen illumination for brightfield
Standard filter configurations:
Semrock BrightLine quad Sedat
- DAPI, Hoechst, etc.
- Alexa488, FITC, GFP, etc.
- TRITC, Alexa555, 568, 594, mRFP, mCherry, TdTomato, etc.
- Alexa647, Cy5, etc.
Standard objectives:
- 4x 0.13NA Plan FluorÌýWD 17.2 (mm)
- 10x 0.45NA Plan Apo DIC N1ÌýWD 4 (mm)
- 20x 0.50NA Plan FluorÌýWD 2.1 (mm)
- 40xÌý0.60NA Plan FluorÌýWD 1.8Ìý(mm)
- 100x 1.30NA Plan Fluor Ph3 OIL WD 0.20 (mm)
Optional objectives:
- 2x 0.10NA Plan Apo WD 8.5Ìý(mm)
- More (please inquire)
- Ìýfor fluorescence
- Ìýfor color brightfield (e.g. eosin, hematoxylin, crystal violet. etc.). This camera uses a CMOS chip with 5.9 megapixels.Ìý
Ìýmotorized stage. Sample holders for standard slide, 35 and 60 mm dishes, and multiwell plates.ÌýBrightfield color RGB imaging.ÌýMulti-point imaging.ÌýMontaging with stitching.ÌýLive cell imaging with TOKAI-HIT Cell Incubator; 1 X 35 (mm) dish or chamber slide (coverslip bottom).ÌýRapid 4 color imaging via quad Sedat filter set (Semrock BrightLine).
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