Laboratories for Molecular Medicine
Room 111
70 Ship Street
Providence, Rhode Island 02903
Zeiss LSM 880 Confocal Laser Scanning Microscope
Laboratories for Molecular Medicine, Room 133
The Zeiss LSM 880 confocal laser scanning microscope is designed for imaging fluorescence, either in fixed or live samples. A pinhole removes the out-of-focus light and allows investigators to acquire thin optical sections at various focal planes. Stacks of images can be acquired for 3D visualization.
Scanhead. The system has a Quasar module scanhead - 32 channel GaAsP detector and two flanking multialkali PMTs. The 32-channel spectral detector allows unmixing of multiple dyes on-the-fly with a scan of a single frame. Two additional channels can be acquired with the flanking detectors optimized for imaging in deep blue or far red spectra. Both the spectral GaAsP and far-red MA PMT are liquid-cooled keeping the noise very low and thus increasing sensitivity of the system.
Lasers. The system has a unique set of lasers. Besides the typical Argon laser (458/488/514 nm for CFP/GFP/YFP), diode 561 nm laser and a He-Ne 633 laser, the system is also equipped with the HeNe 594 nm laser (mCherry, mPlum), and pulsed high-power 405 nm and 440 nm lasers that can be used for ablation and photoactivation.
Objectives. This LSM 880 has a set of premium objectives for a broad range of applications:
LD C-Apochromat 40x/1.1 W Corr UV-VIS-IR. This objective has a unique working distance of ~600 um for imaging large volumes, e.g. cleared specimen.
Plan-APOCHROMAT 100x/1.4 Oil Ph3. An objective for fluorescence and phase contrast imaging
Plan-Apochromat 40x/1.3 and 63x/1.4 Oil DIC UVVIS-IR. Both lenses are routinely used for high-resolution imaging and are corrected over a broad range of spectral lines from UV to IR.
Finally, two highly corrected air (no immersion) objectives Plan-Apochromat 20x/0.8 WD=0.55 M27 and Plan-Apochromat 10x/0.45 WD=2.0 M27.
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The Zeiss LSM 710 Confocal Laser Scanning Microscope
Laboratories for Molecular Medicine, Room 111. NOTE: the LSM710 will be moved to SFH later this year
The Zeiss LSM 710 confocal laser scanning microscope is designed for imaging fluorescence, either in fixed or live samples. A pinhole removes the out-of-focus light and allows investigators to acquire thin optical sections at various focal planes. Stacks of images can be acquired for 3D visualization.
The confocal module is mounted on a Zeiss Axio Observer Z1 inverted microscope with high quality objectives (listed below). A heated stage is available for imaging live samples at 37°C. The microscope has multiple lasers for excitation at 405, 458, 488, 514, 561, 594, and 633 nm, and is equipped with a 34-channel QUASAR detector, which can be set to collect emitted light of any wavelength within the visible spectrum. The QUASAR detector is capable of acquiring spectral information and its flexibility allows imaging of a broad range of fluorescent indicators, including the Alexa-Fluors, Calcium-Green, Cy2, Cy3, Cy5, DAPI, Fluorescein (FITC), FM1-43, Fluo-3, Fluo-4, GFP, GFP variants (BFP, CFP, EGFP, YFP), Hoechst 33258, Rhodamine, Texas Red, TRITC, and others. Images are acquired with ZEN 2008 software and can be stored on a CD, DVD, or a flash drive via a USB connection. The LSM images can be opened using a freely downloadable ZEN viewer.
Objectives on the 710:
|
Mag
|
Correction
|
NA
|
DIC
|
WD (mm)
|
Coverslip #
|
|---|---|---|---|---|---|
|
10x
|
Plan-Apochromat
|
0.45
|
-
|
2.0
|
0.17mm #1 1/2
|
|
20x
|
Plan-Apochromat
|
0.8
|
DIC
|
0.55
|
0.17mm #1 1/2
|
|
40x W
|
C-Apochromat
|
1.2
|
DIC
|
0.28
|
0.17mm #1 1/2
|
|
63x Oil
|
Plan-Apochromat
|
1.40
|
DIC
|
0.11
|
0.17mm #1 1/2
|
Theoretical XY Objective Resolution*: based on: Dxy=0.61*wavelength/NA
|
Mag
|
NA
|
~460 nm (Dapi)
|
~535 nm (Fitc)
|
~620 nm (Rho)
|
~720 nm (Cy5)
|
|---|---|---|---|---|---|
|
10x
|
0.45
|
625 nm
|
725 nm
|
840 nm
|
975 nm
|
|
20x
|
0.8
|
350 nm
|
410 nm
|
475 nm
|
550 nm
|
|
40x W
|
1.2
|
235 nm
|
270 nm
|
315 nm
|
365 nm
|
|
63x Oil
|
1.40
|
200 nm
|
230 nm
|
270 nm
|
315 nm
|
Theoretical Z Objective Resolution*: based on: Dz=wavelength/(NA*NA)
|
Mag
|
NA
|
~460 nm (Dapi)
|
~535 nm (Fitc)
|
~620 nm (Rho)
|
~720 nm (Cy5)
|
|---|---|---|---|---|---|
|
10x
|
0.45
|
2270 nm
|
2640 nm
|
3060 nm
|
3555 nm
|
|
20x
|
0.8
|
720 nm
|
835 nm
|
970 nm
|
1125 nm
|
|
40x W
|
1.2
|
320 nm
|
370 nm
|
430 nm
|
500 nm
|
|
63x Oil
|
1.40
|
235 nm
|
270 nm
|
315 nm
|
370 nm
|
Nikon Ti2-E Fluorescence Microscope for high-content imaging
Laboratories for Molecular Medicine, Room 111 (installed Oct. 2018)
The Nikon Ti2-E fluorecence microscope is a high-content imaging system, based on an inverted microscope and equipped for imaging standard slides, culture dishes, and microplates (e.g. 96-well plates). The microscope includes 'Perfect Focus' for autofocus applications, a Photometrics Prime 95B sCMOS camera for imaging fluorescence, a 16 megapixel Nikon DS-Ri2 color camera for imaging histology slides, a motorized stage and NIS-Elements High Content (HC) software for multiwell plate acquisition.
Objectives:
- 4x Plan Apochromat Lambda, N.A. 0.2, W.D. 20 mm, F.O.V. 25 mm
- 10x Plan Fluor Phase Contrast DL, N.A. 0.3, W.D. 16 mm, F.O.V. 25 mm
- 20x Super Plan Fluor ELWD, N.A. 0.45, W.D. 6.9-8.2 mm, F.O.V. 22 mm, DIC, Correction Collar 0-2 mm
- 40x Super Plan Fluor ELWD, N.A. 0.6, W.D. 2.8-3.6 mm, F.O.V. 22 mm, DIC, Correction Collar 0-2 mm
- 60x Oil Plan Apochromat Lambda Phase Contrast DM, N.A. 1.4, W.D. 0.13 mm, F.O.V. 25 mm, Ph3
Zeiss Axiovert 200M Fluorescence Microscope
Laboratories for Molecular Medicine, Room 111
The Zeiss Axiovert 200M Fluorescence Microscope is a motorized inverted microscope with a Zeiss Mrc5 color camera and a Roper CoolSnap HQ monochrome camera controlled Axiovision 4.8.2 software. This microscope is well equipped for transmitted light microscopy and fluorescence microscopy.
Objectives:
|
Mag
|
Correction
|
NA
|
Phase
|
DIC
|
WD (mm)
|
Coverslip #
|
|---|---|---|---|---|---|---|
|
5x
|
Plan-NEOFLUAR
|
0.15
|
Ph1
|
-
|
13.6
|
0.17mm #1 1/2
|
|
10x
|
Plan-NEOFLUAR
|
0.3
|
Ph1
|
DIC II
|
5.5
|
0.17mm #1 1/2
|
|
20x
|
Plan-NEOFLUAR
|
0.5
|
Ph2
|
DIC II
|
2.0
|
0.17mm #1 1/2
|
|
40x
|
Plan-NEOFLUAR
|
0.75
|
Ph2
|
DIC II
|
0.5
|
0.17mm #1 1/2
|
|
40x Oil
|
Plan-NEOFLUAR
|
1.3
|
Ph3
|
DIC III
|
0.2
|
0.17mm #1 1/2
|
|
100x Oil
|
Plan-NEOFLUAR
|
1.3
|
Ph3
|
DIC III
|
0.2
|
0.17mm #1 1/2
|
Theoretical XY Objective Resolution: based on: Dxy=0.61*wavelength/NA
|
Mag
|
NA
|
DAPI
|
FITC
|
RHO
|
CY5
|
|---|---|---|---|---|---|
|
5x
|
0.15
|
1870 nm
|
2175 nm
|
2520 nm
|
2930 nm
|
|
10x
|
0.3
|
935 nm
|
1090 nm
|
1260 nm
|
1465 nm
|
|
20x
|
0.5
|
560 nm
|
650 nm
|
755 nm
|
880 nm
|
|
40x
|
0.75
|
375 nm
|
435 nm
|
505 nm
|
585 nm
|
|
40x Oil
|
1.3
|
215 nm
|
250 nm
|
290 nm
|
340 nm
|
|
100x Oil
|
1.3
|
215 nm
|
250 nm
|
290 nm
|
340 nm
|
Transmitted light microscopy: The Axiovert 200M has DIC and Phase Contrast on all objectives. The high resolution color camera can be used for imaging fixed samples (e.g. histological preparations) and for time-lapse recordings of living cells. A stage heater is available for imaging living cells at a set temperature.
Fluorescence microscopy: The filter sets allow imaging of a wide variety of fluorescent indicators such as Alexa-Fluors, Calcium-Green, Cy2, Cy3, Cy5, DAPI, Fluorescein (FITC), FM1-43, Fluo-3, Fluo-4, GFP, CFP, YFP, Hoechst 33258, Rhodamine, Texas Red, TRITC, and others. The stage heater and the sensitive monochrome camera make this microscope particularly well suited for imaging dynamic patterns of fluorescence in living cells.
Filter sets on the Axiovert 200M:
1) DIC: Analyzer for DIC
2) DAPI: Chroma set 31000V2, Excitation 325-375 nm, Emission 435-485 nm (bandpass filter).
3) FITC: Chroma set 41001, Excitation 460-500 nm, Emission 510-560 nm (bandpass filter).
4) RHOD: Chroma set 41002c, Excitation 530-560 nm, Emission 590-650 nm (bandpass filter).
5) Cy5: Chroma set 41024, Excitation 590-650 nm, Emission 665 nm and above (long pass filter).
Also available upon request:
6) GFP: Chroma set 41017, Excitation 450-490 nm, Emission 500-550 nm (bandpass filter).
7) CFP: Chroma set 31044v2, Excitation 426-446 nm, Emission 460-500 nm (bandpass filter).
8) YFP: Chroma set 41028, Excitation 490-510 nm, Emission 520-550 nm (bandpass filter).
9) CFP/YFP FRET: Chroma set 31052, Excitation 426-446 nm, Emission 520-550 nm.
Image Analysis
Laboratories for Molecular Medicine, Room 111
MetaMorph software (version 7.0) is available for image analysis. This versatile software package allows for deconvolution (digital removal of out-of-focus light), 3-D reconstruction, brightness measurements, cell counting, colocalization analysis, fluorescence / brightfield overlay, FRET analysis, morphometry, motion analysis, particle tracking, and timelapse measurements.