Services

The Ross Fluorescence Imaging Center is an open access light microscopy core facility at the Johs Hopkins University East Baltimore campus. 

Available services include: 

                            FV3000RS was funded by NIH shared instrumentation grant  1S10OD025244-01 to Prof. Brian O'Rourke, with Prof. Mark Donowitz, co-P.I. - most projects are through our NIDDK P30 Center.

                               Please cite the grant number in your acknowledgement section!

20180711Wed: Our two main instruments are confocal microscopes. With the arrival of our Olympus FV3000RS confocal microscope, I am posting my recommended XY pixel and Z step size to get optimal image quality from our confocals + GPU deconvolution (SP8 -> HyVolution2 or FV3000RS w/Olympus GPU deconvolution).

McNamara 20180711W Recommended Confocal Microscope Pixel Sizes

XY resolution: d = 0.6 * Lambda / NA … pixel size = d/3

Z resolution = 3 * XY resolution

Leica SP8 = ACCM Confocal Microscope (iLab Organizer)

* 405, 488, 552, 638 nm lasers

* HyD1, PMT2, HyD3 ... I normally use the HyD's in photon counting mode (16-bit acquisition), 10 line accumulation (more or less if needed or usable, respectively). Emission band at least 10 nm from laser line (more if close to coverglass or slide).

Objective lens               XY Pixel size    Z step        WD (working distance)

20x / 0.75NA dry               120 nm        360 nm       0.62 mm

63x / 1.40 NA oil                 50 nm         150 nm      0.14 mm

   * On our SP8, I suggest 600Hz for maximum flexibility of Zoom (0.75 to 10+), and higher scan speed if possible (SP8 max is 1800 Hz, zoom 7.5 or more). Faster scan speed and more line accumulation => generally better fluorophore photophysics than same 'total dwell time'. That is, 600 Hz * 10 line accumulation

Olympus FV3000RS confocal Microscope

* 405nm, 445nm, 488nm, 514nm, 561nm, 640nm and 730nm lasers.  
* 4 GaAsP high sensitivity, TRU Spectral detectors
* 2 external GaAs red-shifted high sensitivity detectors

Standard (dry or oil) objective lenses

Objective lens  mag   NA      XY Pixel size         Z step        WD

PLAN APO         2X 0.08         1250 nm          3750 nm    6.2 mm

UPLAN S-APO 10X 0.4             250 nm            750 nm    3.1 mm

U PLAN S-APO 20X 0.75          120 nm            360 nm    0.6 mm

U PLAN S-APO 60X 1.35 oil       50 nm             150 nm    0.15 mm

Silicone Oil objective lenses

Objective lens                        XY Pixel size        Z step        WD

UPLSAPO N 30X 1.05 SI           100 nm           300 nm      0.8 mm

UPLSAPO     40x 1.25 SI             80 nm           240 nm      0.3 mm

UPLSAPO   100x 1.35 SI             50 nm           150 nm      0.2 mm

 

Why these pixel and Z settings? Yes.

Some answers might (or not) be in 'my' CPHG Unit (which awesomely is open access) at https://currentprotocols.onlinelibrary.wiley.com/doi/abs/10.1002/cphg.42 

If you are like Sammy Hager, and cannot drive 1.4NA with 50nmXY and 150nmZ ( https://www.youtube.com/watch?v=RvV3nn_de2k )  that's nice. But hopefully you have a better reason than the microscope company default settings of 512x512 pixels and 400 Hz.

NIRF: Near Infrared Fluorescence

Our new Olympus FV3000RS confocal microscope will have a 730nm laser and two NIR GaAs PMTs (Gallium Arsenide photomultiplier tubes):

 730nm lasers.  
* 4 GaAsP high sensitivity, TRU Spectral detectors
* 2 external GaAs red-shifted high sensitivity detectors

GaAs PMTs

 

 

 

 

 

 

 

 

 

 

 

 

Figure from:

Wang Y, Wang K, Wen W, Qiu P, Wang K 2016 Comparison of Signal Detection of GaAsP and GaAs PMTs for Multiphoton Microscopy at the 1700-nm window. IEEE Photonics J 8: 6803406 

https://ieeexplore.ieee.org/document/7471455

 

 

We are not the only light microscopy core facility at Johns Hopkins University. We will try to keep the links below up to date - if you find a non-functional link, or would like us to add a link, please contact George McNamara, gmcnamara@jhmi.edu  with information.

We note there are several Zeiss LSM 880 or 980 AiryScan or AiryScan2 confocal microscopes at the East Baltimore campus, including:

* ICE = Institute for Cellular Engineering (Miller Bldg)

* MicFac (Physiology Bldg)

* Wilmer Eye Institute (see Wilmer web page)

? (maybe) Neuroscience (PCTB?)

We note that AiryScan and AiryScan2 data would likely benefit from SVI.nl Huygens deconvolution for AiryScan, see https://svi.nl/Huygens-Array-Detector-Software which may compute better data than what Zeiss produces (Huygens needs the raw data file format from the Zeiss microscopes). We routinely encourage our confocal microscope users to use deconvolution, either Leica SP8 --> Hyugens Essential ("HyVolution2" in the Leica SP8 LAS X software), or FV3000RS confocal and FISHscope widefield  - or our other Z-series capable microscopes -- data to the Olympus cellSens deconvolution software (which was purchased on the FV3000RS grant, so use should have that grant number in acknowledgement).

Our current user base -- mostly G.I., ACCM, and cardiology, with many other School of Medicine Departments, and Dept of Medicine Divisions, represented. We do our best to keep informed of our users needs (mostly money) and wants (mostly a lot more money). So far (Prof. Bin Wu and Dr. George McNamara's management since 5/2017) our users needs are either (a) well satisfied by ourt confocal and other conventional microscopes, or (b) they go elsewhere for super-resolution and don't tell us. 

JHU Light Microscopy Cores
JHU iLabs Organizer JHU / JHMI wide core resources

iLabs Organizer ... we  suggest you search for   confocal

https://johnshopkins.corefacilities.org/landing/42#/search
MicFac East Baltimore - Physiology Bldg

https://johnshopkins.corefacilities.org/service_center/show_external/3804

lots of equipment and staff.

* Many confocal Microscopes ... mid-2019 prices: $29/hr, see https://microscopy.jhmi.edu/Services/fees.html

* Four super-resolution microscopes. ... OMX, Nikon TIRF/dSTORM Super-Res, Lattice Light Sheet ... see URL above for rates.
MPI Core (Neurosci P30) East Baltimore - PCTB

http://neuroscience.jhu.edu/resources/3

multiphoton and confocal microscopy:

Multiphoton Imaging Core

Advanced Imaging and Physiology Resources For Common Use

NINDS Institutional Center Core Grant To Support Neuroscience Research: Multiphoton Imaging (MPI) Core

This is part of the Department of Neurosciences. their core's web site 

http://neuroscience.jhu.edu/resources/1

includes links to many JHU cores and mentions their department has a machine shop.

Microscopy and Imaging Core Facility (MICF)

East Baltimore -

Robert H. and Clarice Smith Building

Wilmer Eye Institute 

https://www.hopkinsmedicine.org/wilmer/research/micf 

Zeiss LSM880 AiryScan

https://www.hopkinsmedicine.org/wilmer/research/core-centers/micf/equipment/zeiss-LSM880.html

 

 
ICE East Baltimore - Miller Research Bldg

Institute for Cellular Engineering (ICE). Contact: Stewart Neifert. LSM880 with AiryScan and FastAiryScan, and LSM710

https://www.hopkinsmedicine.org/institute_cell_engineering/index.html

https://www.hopkinsmedicine.org/institute_cell_engineering/about

* G.I. Center members may be eligible for G.I. P30 Center financial contribution to help pay for your use of the ICE LSM880 (i.e. for AiryScan and FastAiryScan capabilities), please contact Prof. Donowitz or Dr. McNamara if interested.
BME BME

BME

https://www.bme.jhu.edu/research/facilities-resources/

The department has a Nikon epifluorescence microscope equipped with a CCD camera for digital imaging. This unit is a core facility, available to departmental members at large. In addition, the department has an Olympus Fluoview 300, laser-scanning confocal microscope that is shared among several departmental laboratories that competed successfully for an NIH shared instrumentation grant. The confocal microscope is equipped for simultaneous electrophysiological and patch-clamp recording, and has custom lasers permitting CFP/YFP FRET imaging.
MRB  

MRB Molecular Imaging Service Center and Cancer Functional Imaging Core (MRB - miller Research Bldg)

https://johnshopkins.corefacilities.org/service_center/3787/?tab=equipment      (iLab Organizer)

Multiphoton Intravital Microscope

  • The Olympus FV1000MPE multiphoton laser-scanning microscope offers bright, clear imaging deep within specimens. The microscope is equipped with a wide variety of regular and water immersion objectives for in vivo and ex vivo imaging (see table below). In addition, our facility also carries a special 20 X MicroProbe objective that allows high-resolution imaging in vivo. This MicroProbe objective has a unique design employing a characteristic long, narrow tip that is optimized for insertion into an animal’s body to allow observation of various fluorescence signatures. The tunable pulsed laser with a range of 690 nm to 1040 nm and 3 standard Photo Multiplier Tube (PMT) light detectors (2 have spectral discriminations) enables the detection of fluorescence markers such as most Alexa dyes, Fluorescent proteins, DAPI, Rhodamine, Calcein, Fluo-3, Fluo-4, Indo-1, FITC, Hoechst, and second harmonic signal generating and auto-fluorescing molecules such as collagen I and NADH. The specialized microscope software and motorized x-y stage allows for users to take z-stacks, time series, and tile images of true 12 bit acquisition with up to 4096 by 4096 pixels. Application: a>Imaging neural networks in the brain. b> Photoreceptors in the retina. c> Cancer cell and tissue structures in the various vital organs of small animals such as mice or rats. d> SHG Imaging for Collagen 1 fibers. Animals and specimens that are still radioactive at a low dose will be allowed on this microscope to integrate multiphoton microscopy into multimodal imaging studies using PET and SPECT.
Cell Imaging Core Facility The Sidney Kimmel Comprehensive Cancer Center

https://www.hopkinsmedicine.org/kimmel_cancer_center/research_clinical_trials/research/shared_resources/cell_imaging.html

https://johnshopkins.corefacilities.org/service_center/3000/?tab=equipment   (iLab Organizer)

  • Nikon C1si True Spectral Imaging Confocal Laser Scanning Microscope System. One of the advantages of conventional Confocal microscopy is the ability to optically section tissue or cells therefore eliminating out-of-focus glare. The C1si also has the capability of acquiring 32 channels of fluorescent spectra over a 320 nm wide wavelength in a single pass using spectral imaging.
  • Nikon E-600 is an upright microscope with a Nikon DS-Fi2 camera connected. The E-600 scope uses Elements for fluorescent microscopy and for brightfield microscopy.
  • E-800 is an upright Microscope. This also has a CCD camera attached to it for imaging of fluorescent slides using Elements software.
  • Leica LMD 7000 Microscope. Used for laser capture microdissection.
  • Nikon SMZ 1500 fluorescent stereoscopic zoom microscope is suitable for quickly assessing the presence of fluorescent proteins, such as GFP in live cells. In addition, the low magnification capability permits photography of whole tissue mounts using Elements and Act-1 software.
  • TE-200 + CCD camera - Inverted Microscope. Also has CCD Camera attached for imaging of culture flasks/plates using Elements software.
  • Nikon TE2000E is an inverted microscope. 
  Homewood - IIC

http://pages.jh.edu/~iic/resources/instrumentation.html

​IIC Light Microscopy (20180501 list)

• Zeiss LSM 800 Confocal with AiryScan supper-resolution module, motorized stages and 405, 488, 561 and 633nm lasers  and Pecon incubator(acquired with funds from NIH SIG award #1S10OD020152-01A1, McCaffery PI)

• Lavision BioTECH Ultramicroscope II lightsheet microscope equipped with 405, 488, 561, 640, and 785nm lasers and okoLab incubator (acquired with funds from NIH SIG award #1S10OD020152-01A1, McCaffery PI)

• Two Zeiss LSM 700 confocals with Pecon cage incubators for live imaging. Lasers include 405, 488, 555, and 633 nm.

• Zeiss LSM 780 confocal microscope with Pecon cage incubator;

• Zeiss AxioObserver Yokogawa CSU-X1 spinning disk confocal equipped with dual Evolve EMCCDs, 405/488/561and 633nm lasers, FRAP module, and stage incubator for live imaging. (acquired in part with NIH S10 award #1S10OD010712-01A1, PI McCaffery)

• Zeiss LSM 510 Multiphoton and Confocal equipped with Confocor 3 Fluorescence Correlation Spectroscopy module an

• Zeiss AxioObserver Epifluorescence SIM microscope equipped with Apotome 2 and Hamamatsu Orca Flash 4.0 CMOS camera. (acquired in part with NIH S10 award #1S10OD010712-01A1, PI McCaffery)

• 3i Marianas Live Cell Imaging Workstation with In Vivo Scientific cage incubator equipped with dual Cascade II 512 EM cameras for simultaneous 2-channel acquistion and TIRF (acquired in part with NIH S10 award #1S10OD010712-01A1, PI McCaffery)

• Zeiss Axiovert 200 LM with In Vivo Scientific cage incubator equipped with excitation and emission filter wheels; and Cooke Sensicam camera; also equipped for microinjection and micromanipulation (acquired in part with funds from NIH SIG award #1S10RR019409-01, McCaffery PI)

• Zeiss Axiovert 100 LM equipped with a CoolSnap digital monochrome camera; and and I-Pentamax 512 digital camera (acquired in part with funds from NIH SIG award #1S10RR019409-01, McCaffery PI; and NIH R01GM060979-06, Wendland PI)

• Zeiss Axioplan II LM equipped with a Cooke Sensicam digital monochrome camera; and a Q-Imaging color camera (This system is excellent for polarized microscopy and color photomicrography.)

• Zeiss AxioZoom V16 Fluorescence Stereoscope equipped with Axiocam MRm camera.

Note: accessories not listed here.

 

 
Carnegie embryology Carnegie Institute for Science - Department of Embryology (next to JHU Homewood campus)

https://emb.carnegiescience.edu

One Leica STED nanoscope and lots more confocals.
     
HI^2 BME & more

Hopkins Imaging Initiative (BME centric, radiology, image processing, some microscopy)

http://imaging.jhu.edu

Welcome to the Hopkins Imaging Initiative (HI^2), a collaborative resource for all of the imaging research at Johns Hopkins University. HI^2 consists of Hopkins researchers who focus on image acquisition, analysis, and application. 

Some of our current goals:

Connect researchers who develop and utilize imaging technologies
Organize seminars to showcase faculty and student research
Provide advice for incoming students
Compile a database of imaging resources
Evaluate classes, media, and resources
Create an informational forum to initiate innovations in imaging 


This resource was primarily created by the students of the Johns Hopkins Training Program in Translational Imaging.
If you are interested in receiving more information about events involving the Hopkins Imaging Initiative, we invite you to Join the Initiative   http://imaging.jhu.edu/join-the-initiative  
   

More cores (for various applications) and links

https://www.hopkinsmedicine.org/research/resources/offices-policies/ora/handbook/corefacilities.html

GCRC "web notes"   http://www.hopkinsmedicine.org/webnotes/core_facilities  (links to monthly calendar)

 
    Other Baltimore Area Academic Confocal Core's
UMd SOM UMd SOM http://www.medschool.umaryland.edu/cibr/confocal 
NIH NIA IRP CIF Baltimore Bayview NIH NIA IRP CIP Baltimore

https://www.nia.nih.gov/research/labs/irp-core-confocal-microscopy-faciltiy

IRP Core: Confocal Microscopy Faciltiy
Fred Indig, Ph.D., Facility Head

The Confocal Imaging Facility (CIF) provides state-of-the-art equipment, training, and image processing capabilities to assist all researchers of the NIA IRP in experiments involving light and confocal microscopy.

The following major equipment is available:

Zeiss LSM 880 confocal with Airyscan, 355 nm UV laser and including full live cell capabilities
Zeiss 710 confocal including full live cell capabilities
Visitech VT-iSIM super-resolution with heated stage Zeiss Axioscope/MCID system to digitize histological (colorimetric) specimens
The CIF serves as an imaging resource for NIA IRP, and in addition to performing imaging and training researchers in confocal microscopy, we provide consultation in cell biology and imaging experimental design, optimization and analysis and advice on microscopy, including non-CIF systems.

 

NIH NIDA Baltimore Bayview

 NIH NIDA Baltimore Bayview

https://irp.drugabuse.gov/organization/core-facilities/histology/equipment/

NIDA = National Institute on Drug Abuse

Histology Core – Equipment and Services

Olympus MVX10 - macroview zoom microscope

Olympus VS120  - Virtual Slide Scanner

TissueCyte 1000 - two-photon serial tomography scope

LaVision Biotech Ultramicroscope II - light sheet fluorescence microscope.

Equipment

ACCM_IncuCyte_S3

ACCM Incucyte S3 Live-Cell Imaging and Analysis System Live cell imaging system inside 37 C incubator - short to long term...

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. FISHscope

FISHscope Main application: single molecule RNA fluorescence in situ hybridization (smFISH). FISHscope Quick Tips web page...

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q Andor Revolution X1 Spinning Disk confocal microscope -...

Andor Revolution X1 Spinning Disk confocal inverted microscope - Legacy instrument Pease note: this microscope has been...

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Olympus FV1000MP Multiphoton Excitation Fluorescence...

Olympus FV1000MP Multiphoton Excitation Fluorescence Microscope (MPEF) ... also capable of SHG (ex. 840 nm --> 420 nm) ***...

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Li-Cor Odyssey CLx

Li-Cor Odyssey CLx fluorescence scanner. This instrument is hosted by Prof. Donowitz lab and use is intended for G.I. Center...

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