The University of Toronto (the “University”) has a requirement for a fluorescence widefield microscope with the ability to facilitate linear unmixing by phasor-based, intensity-independent unmixing. The purpose of this Advance Contract Award Notice (ACAN) is to signal the University’s intention to award a contract for the above-noted goods and/or services to the following pre-selected supplier:
Leica Microsystems Canada Inc.
71 Four Valley Drive
Concord Ontario L4K 4V8
Canada
Note: The product is to be purchased from the pre-selected supplier’s exclusive distributor, VWR International.
Prior to awarding this contract, the University is providing other potential suppliers with the opportunity to demonstrate that they are capable of satisfying the requirements as per this ACAN, by submitting a Statement of Capabilities during the posting period.
If other potential suppliers submit a Statement of Capabilities that meet the University’s requirements as set out in the ACAN, the University may proceed to a competitive procurement process.
If no other supplier submits a Statement of Capabilities on or before the closing date/time, a contract may be awarded to the above-noted pre-selected supplier.
- Background
The Structural Genomics Consortium (SGC) at the University of Toronto, is a public-private partnership that seeks to accelerate drug discovery through collaborative protein-based science. Through discovery of novel small molecule chemical probes and ligands, SGC goal is to provide tools to the academic community that can be used to better understand the function of understudied proteins. The ligand discovery at SGC requires development of protein assays that reflect the activity and abundance of the protein target. Ligand discovery and characterization includes studies of protein target localization changes, downstream signaling markers and measuring changes in posttranslational modifications. Microscopy provides the means to simultaneously assess the compound impact on the target protein localization, abundance, activation state and monitor cell viability thus facilitating the evaluation of compounds in a wide variety of cells. The proposal outlines protein targeting methods, all of which will rely on imaging: tracking the small molecule inhibitor effects on protein activity, localization and complexes, and compound elicited protein degradation monitoring as well as screening compound libraries for the degrader activity in live cells, which makes fast and longitudinal imaging with low exposure necessary. The microscope will also be used by other researchers at the SGC who image protein binding by fluorescently labeled macrocycles.
The following studies at SGC require microscopy imaging:
- We are developing small molecule inhibitor compounds and their fluorescent derivatives to study the target protein biology. The first step of this development process is to thoroughly characterize the compound properties and off target effects. Thus, we need to simultaneously evaluate target protein localization, its know binding partners and cellular compartment markers if they are known for the target protein or perform the reduced Cell Paint assay that includes cytoskeleton, DNA, endoplasmic reticulum, and mitochondria staining. Flexibility in wavelengths is of utmost importance as by using the fluorescently tagged compounds and HaloTag containing protein target we will be using non-standard fluorophores and will need to differentiate clearly between spectrally adjacent signals. While traditional systems with excitation/emission filters can achieve the separation of select chromophores, only a fully spectral system would give us the flexibility to differentiate between spectral fingerprints and the capture of the whole emission range to maximize signal to noise. Continuous collection of emitted light without the cutoff from emission filters will enable reliable data collection. Photon efficiency is relevant to our application, as fluorophore tagged compound signals are dim (low quantum yield) in comparison to commercial dyes. To enable subcellular resolution computational clearing and magnifications of at least 63x are a must. Computational clearing must be integrated into the workflow, fast, and computationally inexpensive (< 30 s image). The instrument must also be upgradable to a confocal unit to support continuous state-of-the-art imaging.
- We are also developing Proteolysis Targeting chimera compounds (PROTACs) and molecular glues. The target proteins will be tagged with fluorescent proteins containing either destabilization sequence or dimerization domains. These reporter systems will be stably integrated and engineered for protein degradation kinetic studies. As the compound screening will need to take into account cell toxicity, markers of apoptosis, cytoskeletal integrity will be included in the assays to evaluate the spatial information in live cells. To this end, the microscope must be able to perform fast imaging of 96-well plates for screening in both brightfield (phase contrast) and fluorescence with a designated plate imaging mode, with as little excitation light exposure as possible to minimize phototoxicity, with flexibility in terms of chromophores, and with the ability to automize 63x imaging in glass-bottom 96-well plates using an automated dispensing of immersion liquid to enable time-lapsed imaging. The microscope must be upgradable to contain a cell incubation chamber which enables both temperature, humidity, oxygen, and CO2 control as well as protect the sample from light (contained imaging chamber).
For this work an intuitive software interface is a requirement. The trainees performing this work must be able to acquire images with minimal onboarding and training to facilitate integration into their workflow (< 30 minute training required). Compatibility with the similar systems from other SGC sites in Germany and UK is of importance to enable the comparison of compound evaluation results and adaptation of protocols.
Therefore, the University of Toronto intends to purchase a Mica Microhub with Fluosync detector with the minimum specifications and requirements described below to address these research needs. The items will be in new, working condition, and need to be delivered within 90 days of the order.
- Minimum Specifications and Requirements
- Technical Specifications
- Instrument – Fluorescence Widefield Microscope with Linear Unmixing Ability
A filter-free widefield fluorescent microscope with linear spectral unmixing.
- A system for fully automated imaging, which is an inverted imaging system for fluorescence axis widefield imaging
- Widefield detection unit for FluoSync with the ability to detect at least up to 4 labels simultaneously with a spectral detection range between 420 - 705 nm, consisting of 5 MP CMOS detectors, automatically aligned with 0.3 px precision and at least 12 Bit resolution as well as a system frame rate of 10 fps when imaging 4 fluorophores simultaneously for fast imaging, quantum efficiency of ~ 61% at 450 nm, 64% at 500 nm, 67% at 536 nm, 46% at 700 nm
- Transmitted light axis with a S40 condensor
- Automated modulation contrast unit
- Hardware autofocus for drift correction
- Software autofocus for image based focusing
- Sample finder to generate in-focus overview
- A precise scanning stage with control in x, y, z (Travel range 127 x 83 mm, Resolution < 0.02 μm, Reproducibility < 1 μm, Max. speed 37 mm/sec)
- Imaging capability in standard plastic tissue culture plates (6, 12, 24, 96, and 384-well plates) up to at least 20x with automated selection of wells
- At least a 4-fold LED light source (365 nm with 100 mW direct LLG output, 470 nm with 170 mW direct LLG output, 560 nm with 170 mW direct LLG output, 625 nm with 170 mW direct LLG output)
- Objective turret with 1.6x and 10x semi-apochromate and space for a total of 6 additional objectives
- Objective HC PL APO 20x/0.75 CS2 Plan apochromatic objective with superior color correction Optimized for confocal scanning applications For use with 0.17 mm coverglass, no immersion. Free working distance: 0.62 mm
- Objective HC PL APO 63x/1.20 W motCORR CS2 - Plan apochromatic corrected water immersion objective with high numerical aperture, optimized for Mica. FWD: 220 μm. - Motorized correction collar for precise and fast adjustment of coverglass thickness from 0.14-0.18 mm, water immersion within the temperature range 20-40°C.- MotCORR connector on the objective- Compatible with Type 2A watercap and AutoImmersion watercap with sensor A
- Objective turret with 1.6x and 10x semi-apochromate and space for a total of 6 additional objectives
- Real-time control to ensure synchronization of all system components
- MotCorr board to enable use of MotCorr objectives plus SmartCorr
- Provide a light-controlled environment
- The microscope must be upgradable to a live cell system with temperature, CO2, O2 and humidity regulation
- The microscope must be upgradable to a confocal imaging system
- Software
- LAS X Mica Base license
- Mica workstation and analysis package
- Thunder functionality to enable at least 3 computational clearing modes (instant computational clearing, small volume computational clearing, large volume computational clearing)
- Smart move I2C
- “Learn and Results” license packages to improve sample processing
- Sample finder, navigator, and objective collision prevention
- Accessories
- Anti-vibration table
- Okolab K-frame standard and chamber slide
- Okolab Click-in 35 mm petri dish
- Okolab click-in 4x slide
- Mica Auto-immersion: feedback-loop controlled auto-immersion for water immersion objectives including pump, tubing, water reservoir
- Computer
- At least a HP Z32 4K-UHD Display, 31.5 inch
- At least a Win10 IoT Enterprise 2019 operating system
- At least Intel XEONW-2123 3.6 4C CPU (CPU)
- At least 128GB (4X32GB) DDR4 2666 ECC REG RAM (main memory)
- At least NVIDIA Quadro RTX 5000 16GB (4)DP+USB or equivalent (Graphics Board)
- At least 480GB SATA Enterprise SSD (System Drive)
- At least 2x1.9TB M.2 SSD (Temp Drive and Data Drive)
- At least 9.5 DVDWR 1ST ODD (DVD writer)
- At least HP 5/5/5 Warranty – provided by HP manufacturer for the PC and its
- Keyboard, mouse
- Delivery, Installation, Testing and Training
- Service installation by a factory trained and certified Field Service Engineer
- Microscopy Training Basic with 4 hours of training by a Leica Application Specialist
- Follow-up support resources such as access to the remote application support
team and web material
- Warranty and Service
- One (1) year of full parts and labour warranty must be included for each of the core items (microscope, light source, detectors, cameras, stage) as well as supporting infrastructure (command/control computers, etc.) commencing upon successful installation and acceptance of the equipment.
- Provide access to discounted and favourable prices on parts, labour and service agreements during the contract term.
- The Warranty Extension coverage for 4 years that starts at the end of the Standard Warranty and includes the following: 1 annual preventative maintenance visit (PM), unlimited onsite repairs, including replacement for service parts, components, or modules necessary, for repairs, prioritized on-site response time for remedial services (critical repairs), unlimited access to Technical Support Center (standard working hours), firmware and software updates when available.
- Technical Support
a. The supplier should have the capacity and available resources to provide timely technical support in instrumental and applications development work on these tools. This will include sharing of technical information, collaboration with the University as well as appropriate application, consulting, and design expertise.
- Evaluation Criteria:
The supplier’s Statement of Capabilities will be evaluated on a pass or fail basis based on their ability to meet the minimum specifications and requirements outlined above. If the specifications set above are not met, please provide an explanation, including recommendations as to why the different specification or performance features meets or exceeds the University’s requirements.
- Contract Term:
The contract term will be for one (1) year.
- Limited Tendering Justification:
Leica's microscope contains FluoSync, a filter-free system, which addresses the limitations of linear unmixing by facilitating phasor-based, intensity-independent unmixing. Other microscopes use excitation and emission filters. Linear unmixing allows flexibility in wavelengths, which is of utmost importance as we will need to deviate from commonly used fluorophores as well as the ability to differentiate clearly between spectrally close fluorophores. This filter-free system also makes it the most flexible option for our custom compounds conjugated to fluorophores and protein labeling fluorophores/dyes.
The MICA system's approach for spectral unmixing simplifies the spectral imaging workflow in comparison to other systems which need designated filter combinations and circumvents the extensive calibration necessary in linear unmixing. As opposed to other microscopes, it enables unmixing solely based on the spectrum rather than intensity, beneficially utilizing crosstalk and effectively mitigating errors associated with linear unmixing. It uses a single image exposure through an automated spectral unmixing process, thereby enhancing efficiency in multicolour fluorescent imaging, as opposed to collecting images by cycling through different emission filters as other instruments. Employing emission and excitation filters that are spaced closely together can achieve separation between spectrally similar fluorophores but at the cost of signal intensity as well as necessitating designated filter combinations for each pair. The spectral unmixing approach of the MICA system on the other hand utilizes the entire signal from the sample instead of cutting of signal with emission filters, thus offering a more sensitive method. Images generated from phasor unmixing retain fine details while minimizing noise, which is crucial for subcellular imaging.
In contrast to traditional sequential imaging methods, FluoSync enables rapid, simultaneous multi-channel acquisition. With simultaneous four-channel imaging, it is ideally suited for scanning large number of wells with cells exposed to screening compounds and imaging fast dynamic processes in live cells. The Mica platform has a designated mode for the imaging of well plates and a scanning setup to obtain an overview of the whole slide fast. This contrasts with other systems, which take hours to image plates. Moreover, FluoSync simplifies the experimental workflow by eliminating the need to manage multiple filter cube sets.
- University Contact:
All questions/responses related to this ACAN must be sent to the following contact:
Philip Clements
Procurement Officer
Philip.clements@utoronto.ca
- Process of Submitting a Statement of Capabilities
Interested suppliers may submit a Statement of Capabilities clearly demonstrating how they meet the requirements of the ACAN, by sending their response via e-mail to the University contact before the closing date/time.
Statement of Capabilities received by the closing date/time will be evaluated and considered solely for the purpose of determining if a competitive procurement process is warranted.
The University will notify the suppliers who have submitted a Statement of Capabilities in response to this ACAN of its final decision.
