McMaster University has a requirement for an automated microscope with spinning disk confocal and widefield microscopy with multi-mode detection in one system for high content imaging. The system must have automated water immersion objectives with bubble detection functions for high resolution high magnification, automated water immersion imaging for microbiological applications. The system must have 6-line laser light sources for confocal microscopy (405, 445, 470, 520, 555, and 640 nm). The spinning disk confocal microscope must be capable of 4 channel fluorescence imaging with the 6-user interchangeable fluorescence filter cubes including DAPI, GFP, and TRITC. Up to 4 widefield microscopy fluorescence color channels the following color cube and LEDs (DAPI filter cube with 365 nm LED; GFP filter cube with 465 nm LED; TRITC filter cube with 554 nm LED) with the option to add or remove or upgrade to additional color channels as needed is required. It must have UV/Vis Absorbance, Fluorescence (Top/Bottom), Luminescence also available. The flexible plate reader must have a quadruple grating monochromator and variable bandwidths from 9 nm to 50 nm in 1 nm increments for fluorescence to improve sensitivity over a fixed bandpass with narrow Stokes shift fluorophores.
The system must be able to control temperature to 45°C with a 4-zone temperature control system, with the ability to set temperature gradient to prevent condensation on plate lids for condensation free incubated kinetics and microbiological growth curves. Temperature control must be highly uniform, ±0.5°C at 37°C across an entire 96-well plate. It must also include a gas controller for monitoring and controlling O2 and CO2 levels to support live cell assays. Gas controller must use 100% CO2 gas bottles and automatically maintain 5% CO2 in incubation chamber. The system must be able to accommodate compatibility for microscope slides, 6-1536-well microplates, 60-100 mm Petri dishes, cell culture dishes, T25 cell culture flasks and counting chambers.
Automatic imaging across different sized microplates or slides is required of the water immersion system. The system must have a hit-picking function to automatically identify samples of interest to increase acquisition rates and decrease file sizes. It must include a laser autofocus capability for faster focusing, especially with dim samples, in the presence of debris, or multicell samples.
It is required that the system has small footprint and weight to minimize lab bench needs not exceeding 28” w x 18.5” h x 21” d). To meet McMasters needs, an automated objective turret with 6-objective capacity for confocal and inverted microscope with the ability to change objectives during an imaging assay without human intervention is required. Due to the diversity of specimens and cell types to be imaged, it must have a 6 objective capacity and be compatible with 1.25x to 60x magnification objectives to cover broad imaging applications. This includes the following long working distance objectives with correction collars to accommodate a variety of labware: 4x, 20x, and 40x Plan Fluorite Phase Objective and 60x Water Olympus Plan Apochromat Objective. There is also a need for a syringe pump-based dual reagent injectors, able to automatically dispense reagents in all detection modes and plate types for fast kinetics, flash luminescence etc. The injectors must have straight tips for direct centre dispensing and available angled tip options that can be added in the future by a user for use with gentle dispensing in cell-based assays. McMaster University requires that this system have a joystick for controlling stage movement and focus and is capable of full automation with autofocus, autoexposure, auto-LED intensity adjustments for walkaway experimental automation. Must come standard with linear, orbital and double orbital shaking capabilities.
The system must include data analysis software for quantitative and qualitative analysis and image analysis (2 licenses) that does not require programming knowledge and with a graphical interface. The software must include image processing (background flattening, deconvolution, image stitching, Z-projection, digital phase contrast), region of interest capability (where a lower magnification image can be taken, and a region of interest can be drawn by the user as a square, poly-line, ellipse, and the instrument will automatically capture at a higher magnification), recording movies of live samples, automatic cell counting, sub-population analysis, dual masking where intensity between different masks can be calculated, spot counting where small objects (tertiary masks) can be counted within other masks, and histograms and scatterplots can be generated, kinetic object tracking and 3D rendering capabilities for confocal and widefield z-stack image sets. To handle these large datasets and analysis it must include a high-performance computer system. Comprehensive installation and training by a field application specialist on the instrument operation, data acquisition, data analysis etc. must be included.
