The Wistar Molecular Screening Facility maintains libraries and equipment for high-throughput screening of small molecule and nucleic acid (e.g. cDNA, miRNA, siRNA, shRNA) libraries. The facility has the following equipment available for use by its users:
- Liquid handling robotics to transfer compounds/nucleic acids from library stock plates to assay plates.
- Automated dispensers to rapidly fill assay plates with bulk reagents (e.g. cells, proteins, buffers, etc.).
- Plate reader with absorbance, luminescence, ultraluminesence, fluorescence, fluorescence polarization, time-resolved fluorescence, and alpha screen detection capability.
- Automated plate washers.
- Tissue culture hood and incubators.
- Multichannel pipettes.
- Plate Sealer and Bar Code Labeler.
Laboratory personnel will provide users with training in the use of the walk-up equipment (e.g. bulk reagent dispensers, plate washers, and plate readers). Users must be trained by facility staff prior to using the instrumentation in an unassisted mode. To schedule a training session, please make an appointment with David C. Schultz prior to training on the instrumentation in the Facility. All training time on equipment is charged as assisted use.
|Automated pipetting workstations|
The laboratory is equipped with a Perkin Elmer Janus 96/384 Modular Dispensing Tool (MDT), automated liquid pipetting workstation for compound/reagent transfer and library management. The Janus MDT is equipped with a 384 well head for transferring volumes from 0.5 ul to 25µl with disposable tips and a 96 well head with capabilities of transferring volumes upto 235 ul. The MDT workstation is integrated with a plate stack unit with capacity for up to 50 microplates.
The laboratory is also equipped with a Perkin Elmer Janus Verispan 8 channel independent pipetting workstation for use in reformatting libraries (e.g. dilution series, Hit-Picking) and inserting controls on assay plates. The Verispan 8-tip is designed for use with fixed washable tips, disposable tips in 20 ul, 200 ul, and 1 ml sizes, and disposable filter tips in 25 ul and 175 ul sizes, or both.
|Bulk reagent dispensers|
|The Biotek Microflo uses a peristaltic pump mechanism to dispense liquids in as many as 8 channels, with each nozzle fed by a separate feeding tube. The entire tubing assembly is removable and can be autoclaved when sterility during dispensing is essential. The Microflo can be programmed to fill specific plate columns as desired by the user, and is capable of dispensing 2.0 to 2000 µL in increments of 1 µL. The priming/dead volume for this automated dispenser is about 6-10mL. In addition the Microflo can be integrated with Biostak plate stacker unit that can accommodate up to 30 plates.|
|The Biotek µFill|
|This is a syringe-based, automated plate dispenser with a 16-channel manifold with a volume range of 5uL to 1500uL for 384-well plates and 10-3000uL for 96-well plates. The syringe and tubing assembly is removable and can be autoclaved when sterility during dispensing is essential. The priming volume for this liquid dispenser is about 13-20mL. The uFill can also be integrated with Biostak plate stacker unit that can accommodate up to 30 plates.|
|The ELx405 is an automated plate washer with a 96 channel head that can be used with 96- and 384-well plates. This plate washer also offers a sonicating wash bath for the pins to help minimize clogging and increase the cleaning capabilities. The Elx405 offers the flexibility of integration with the BioStak plate stacker, as necessary.|
|The laboratory has a PerkinElmer EnVision Xcite Multilabel plate reader with a 40-plate stacking carousel. In addition, the facility has access to a Pekin Elmer Top Count and Wallac Victor 2 .|
The EnVision Xcite plate reader is capable of standard multi-mode detection, including:
- Fluorescence Intensity (FI)
- Ultra Luminescence
- Time-Resolved Fluorescence
- Fluorescence Polarization (FP)
- AlphaScreen (Amplified Luminescence Proximity HomogeneousAssay)
A wide selection of filters and dichroic mirrors make most dyes and dye-pairs accessible to the reader. Certain filters and mirrors are designed for fluorescence polarization (FP) assays. These are indicated (*) in the tables below.
|Umbelliferone 355||(λ: 355 nm, bandwidth: 40 nm)|
|Fura2, BFP 380||(λ: 380 nm, bandwith: 10 nm)|
|Photometric 405||(λ: 405 nm, bandwidth: 8 nm)|
|Photometric 450||(λ: 450 nm, bandwidth: 8 nm)|
|FITC 485||(λ: 485 nm, bandwidth: 14 nm)|
|Photometric 560||(λ: 560 nm, bandwidth: 8 nm)|
|*FITC FP 480||(λ: 480 nm, bandwidth: 30 nm)|
|*Cy5 FP 620||(λ: 620 nm, bandwidth: nm)|
|Umbelliferone 460||(λ: 460 nm, bandwidth: 25 nm)|
|Fura2, BFP 510||(λ: 510 nm, bandwith: 10 nm)|
|FITC 535||(λ: 535 nm, bandwidth: 25 nm)|
|Rhodamine 590||(λ: 590 nm, bandwidth: 20 nm)|
|*FITC FP S-pol 535||(λ: 535 nm, bandwidth: 40 nm)|
|*FITC FP P-pol 535||(λ: 535 nm, bandwidth: 40 nm)|
|*Cy5 FP S-pol 688||(λ: 688 nm, bandwidth: 45 nm)|
|*Cy5 FP P-pol 688||(λ: 688 nm, bandwidth: 45 nm)|
|AlphaScreen||(λ: 570 nm, bandwidth: 100 nm)|
|General Dual||BS50/BS50 (beam splitter)|
The Wistar screening facility has a tissue culture suite approved for work with BSL2 level pathogens. It includes:
- A 6ft. tissue culture hood, which can house rapid dispense plate fillers (e.g. uFill) to dispense tissue culture cells into assay plates.
- 2 CO2 regulated tissue culture incubators.
- A benchtop centrifuge fitted with microtiter plate holders and an inverted phase-contrast microscope for use while passaging and preparing cells for assays.
The BSL2 classification of this facility enables utilization of retroviruses (e.g. lentivirus) produced from cDNA and shRNA libraries.