Leadership
Co-Core Director:Teresa P. DiLorenzo, Ph.D.
Location: Forchheimer Building, Room 403
Co-Core Director: Dirk Homann, MD,
Location: Icahn School of Medicine at Mount Sinai
Overview
The new Immuno-Technology Core (ITC) has been conceived as an education, coordination
and service core that offers access to advanced cellular, imaging and
immunomodulation technology platforms operated across both Einstein and Mount
Sinai. Specifically, the ITC supports experimental design, execution and
analysis for investigations that seek to employ a tailored approach to mass
cytometry; multiplexed tissue imaging, automated tissue staining and whole
slide image acquisition; biophotonic in
vivo imaging; and immunomodulatory T cell targeting through synTAC
technology.
Objectives
To meet the complex
demands for tailored assay development, performance and interpretation, the ITC
will harness the combined technology infrastructure at Einstein and Mount Sinai
for the following principal objectives:
- To raise awareness about select cellular, imaging, and
immunomodulation technology platforms; to provide consultation and advice
about the potential and limitations of respective technologies; to assist
investigators with experimental planning, design and considerations about expected
financial investments; and to facilitate interactions with managers,
application scientists and data analysis specialists working with each
technology platform.
- To provide support services required for the effective and
efficient implementation of tailored experimental protocols including specific
reagent selection, testing and validation; protocol adjustment and
optimization; and integration with established assay workflows.
- To facilitate access and provide practical support for the usage of
advanced technology platforms:
Cellular
(Mount Sinai): High Dimensional Mass
Cytometry / Cytometry by Time Of Flight (CyTOF)
Tissue
imaging (Mount Sinai):
A) MICSSS platform (multiplexed
immunohistochemical consecutive staining on a single slide)
B) Automated tissue staining (Leica BOND RX Automated IHC
Research Stainer)
C) Whole slide image acquisition (Hamamatsu
NanoZoomer S60 Digital Slide Scanner)
In vivo imaging (Einstein
and Mount Sinai): IVIS Spectrum In
Vivo Imaging System
Immunomodulation
(Einstein): synTac platform (artificial
immunological synapse for T cell activation; unique technology developed at
Einstein)
ITC Technology Platforms
I. High Dimensional Mass Cytometry / CyTOF
Mass cytometry combines the
high-throughput single cell analysis capabilities of flow cytometry with the
specificity and resolution of mass spectrometry for a broad and detailed
characterization of heterogeneous cell samples according to ~50 discrete
parameters at the single-cell level. Sample processing, staining, acquisition
and basic QA/QC are performed and administered through the Mount Sinai Human Immune Monitoring Center (HIMC).
Due to the
fundamentally modular nature of the assay technology, specific experimental
strategies need to be precisely tailored according to the nature of biological
samples, preferred composition of antibody staining samples, sample processing
protocols, data analysis requirements, and potential need for training. Thus, unless noted otherwise, overall cost
estimates can be developed only in detailed discussions with interested
investigators and typically align with the similarly tailored fee schedules
implemented by HIMC.
The ITC provides individualized consultation
about specific experimental design, execution and analysis that is free of
charge. To initiate a consultation, please email ITC Co-Director Dr. Homann (dirk.homann<a>mssm.edu).
Specific topics covered in the consultation include:
Biological samples
(single cell preparations):
Tissue origin
(PBMC, pancreas, other)
Tissue availability
and quantity
Processing
requirements
Quantity of samples
Composition of
antibody staining panels:
>500
metal-conjugated antibodies available through the HIMC.
Multiple novel
pancreatic cell-specific metal-conjugated antibodies available through the ITC.
Commercial
metal-conjugated antibodies.
Custom metal
conjugations of investigator-provided antibodies (ITC service provided at
$450/100 µg antibody)
Sample processing
protocols:
Cell culture (e.g.,
immune cell stimulation)
Bar coding
Cell surface
staining
Intracellular
staining
Sample acquisition
Data analysis:
Basic data
processing and QA/QC
Data storage and
sharing
Advanced data
analysis strategies and platforms
Investigator
training
Overall expense
considerations and cost estimates
II. Tissue Imaging
The ITC provides individualized
access to three tissue staining platforms covering a multiplexed tissue
staining approach, automated tissue staining, and whole slide image
acquisition.
II.A. MICSSS
technology platform: Multiplex Immunohistochemical Consecutive
Staining on a Single Slide, originally developed at Mount Sinai, is a chromogen-based
IHC staining assay independent of proprietary reagents or equipment. It is
performed with formalin-fixed, paraffin-embedded (FFPE) tissue sections using up
to 10 iterative cycles of staining, revelation, scanning and bleaching of
labile chromogenic substrate. The method can be implemented with standard
staining protocols (thus benefitting from previously determined specificity and
sensitivity parameters); works regardless of antibody isotype or species; is
impervious to staining order for the majority of antigens evaluated; is not
limited by antibody crossreactivity, photobleaching or autofluorescence; relies
on brightfield images that present more detail about the microanatomical
organization of the tissue section; and permits prolonged slide storage for
future use as new markers become available. The
fee schedule for MICSSS stains performed by ITC personnel are dependent on the
number of slides, the availability or need for adjustment of MICSSS staining
protocols, the number of iterative staining cycles, and image acquisition modes
and aligns with current HIMC fee schedules. To
initiate a free consultation, please email ITC Co-Director Dr. Homann (dirk.homann<a>mssm.edu);
specific topics covered in the consultation include:
Survey of
MICSSS-adapted staining protocols (including reagents and antibody targets)
Development/adaptation
of MICSSS-adapted staining protocols
Development/adaptation
of MICSSS staining protocols compatible with the autostainer (see item II.B.)
Investigator
training
Overall expense
considerations and cost estimates
II.B. Automated tissue
staining: the ITC coordinates access and
usage of a fully automated Leica BOND RX
Automated IHC Research Stainer that can stain up to 30 slides per run while
testing multiple protocols simultaneously (including both IHC and IF staining).
The instrument offers speed, flexibility and consistency to process numerous
slides in timely manner with high-quality results, and permits a streamlined
approach to optimize new and adjusted staining protocols. To initiate a consultation, please email
ITC Co-Director Dr. Homann (dirk.homann<a>mssm.edu). All consultation and
training is free of charge; pricing for
autostainer usage is contingent on the nature of staining protocols, number of
slides, and usage modality (ITC personnel-assisted or unassisted), and complies
with current HIMC fee schedules. Specific ITC-provided services include:
Tissue staining
protocol and reagent review
Development/adaptation
of standard staining protocols compatible with the autostainer
MICSSS stain
development and/or execution by ITC personnel
Investigator
training
Overall expense
considerations and cost estimates
II.C.
Whole slide image acquisition: the ITC provides access to a Hamamatsu NanoZoomer S60 Digital
Slide Scanner that uses NDP.view2 viewer software (downloadable free of charge
at https://www.hamamatsu.com/us/en/product/type/U12388-01/index.html).
The instrument is a next-generation, high-throughput digital scanner that delivers
rapid and high-quality (20x and 40x) image acquisition of whole slides in both
brightfield and/or four parameter fluorescence modes. Pricing is tailored according to magnification and type/number of image
acquisition modes and corresponds to the current HIMC fee schedule. Additional
services facilitated by the ITC include:
Tissue staining
protocol review
Investigator
training (scanner & software)
Development/application
of image analysis strategies
Overall expense
considerations and cost estimates
III. In vivo imaging
The
ITC will coordinate access and usage of two Perkin Elmer IVIS Spectrum
biophotonic imaging systems, one located in Einstein’s In Vivo Imaging System Facility
and the other in Mount Sinai’s Translational and Molecular Imaging Institute.
The IVIS Spectrum enables longitudinal three-dimensional intravital imaging of
small animals or tissues expressing or containing bioluminescent or fluorescent
probes across the blue to near-infrared wavelength region. The instrument is
equipped with a high-sensitivity charge-coupled device camera which is ideal
for imaging single or multiple probes or reporters. Please schedule a free consultation
with ITC Co-Director Dr. DiLorenzo (teresa.dilorenzo<a>einsteinmed.org) regarding
instrument usage and animal housing for longitudinal imaging studies.
IV. synTac
technology platform
ES-DRC member and Einstein investigator Dr. Steven Almo
recently developed innovative reagents to manipulate disease-relevant T cells
under a variety of conditions, including type 1 diabetes. Termed synTacs (for
“artificial immunological synapse for T
cell activation”),
these inventive reagents are soluble, precision biologics in which
non-exchangeable single-chain peptide-class I MHC (pMHC) is covalently linked
to a modulatory domain (e.g., PD-L1,
a ligand for the T cell-inhibitory checkpoint receptor PD-1) in a manner that
recapitulates the proximity, orientation, and overall organization experienced
at the immunological synapse. This strategy not only allows for the explicit
targeting of disease-relevant T cells, but also greatly reduces the devastating
side effects associated with the global immune modulation directed by all
biologics currently in the clinic. Given the emergent, early-stage, and
cutting-edge nature of the synTac technology, ES-DRC investigators utilizing
these reagents will initially do so in collaboration with ITC Co-Director Dr.
DiLorenzo and/or Dr. Almo as appropriate, in order to maximize the likelihood
of success of the research. Please schedule a free consultation with ITC
Co-Director Dr. DiLorenzo (teresa.dilorenzo<a>einsteinmed.org) regarding
the synTac technology.