| Technology - Screening by Imaging |
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ICCB-Longwood High Throughput Microscopy Facility Information
High Content Screening Microscopes
Image Analysis
Other Information about Screening by Imaging:
Designing an Imaging-Based
Screen
Below is a list of items to consider for designing an imaging based
screen:
- Assay plates: Use black, clear-bottom (plastic or glass), tissue
culture treated multi-well plates. Many researchers use the Corning
Costar 384-well plate (Fisher Catalog number 07-200-655).
- Seeding density: Try a variety of cell densities under the conditions
you plan to use. A good range to begin with is 2,000 to 10,000
cells per well.
- Transfection efficiency of your expression vector: Make sure
that greater than 50% of your cells are expressing your protein
of choice before attempting a screen.
- Imaging live cells: This is possible, but remember that the
microscope stage is not heated and your cells will be at room
temperature for 40-90 minutes during imaging. Use phenol red-free
medium when doing live cell screening.
- After processing cells for imaging fill your wells with PBS
to prevent the cells from drying out and to reduce background
noise.
- Please make sure that the fluorophores that you select do not
conflict with the filter sets on the screening scope. See details
below for each individual microscope.
Training requirements
for Microscope usage
- Access to the imaging microscopes can be arranged through Melody Tsui melody_tsui"AT"hms.harvard.edu.
- Microscope usage training by the Screening Staff is
required prior to signing up for time.
Storage guidelines for imaging data
Screeners keep their visual screening data generated by the imaging scopes
on the HMS-ICCB-L Network. Since this network space is limited, screeners
must move their data to the repository on a regular basis. See
the data
storage guidelines (PDF document) for more instructions.
Notice (posted April 2008) : As of July 1st 2008 the HMS Research IT Group (RITG) will be charging individual microscope users for file storage on the ICCBL servers. The rate is very reasonable, $1 per GB per year, but for imaging screens, obviously, this can add up.
Every three months ICCBL will be billing microscope screeners for the storage space that they are using at the end of each quarter (March 31st, June 30th, September 30th and December 31st). To minimize storage charges, image data that has already been analyzed can be archived to tape. RITG and ICCBL have put in place an archiving system which is free. There is no charge for archived file storage and the files should be accessible with 48-72 hours notice.
ICCBL will remind screeners via email, approximately two weeks before the billing date that they should archive any files that are no longer in use. ICCBL staff (microscopy specialists) will help screeners with the archiving process.
At the beginning of each quarter, ICCBL will invoice the 33-digit code or PO number that you have provided on the ‘ICCBL Screening Facility Fee – Billing information form’ for your current file storage use.
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Schedule microscope time
- You may sign up no more than two weeks in advance and usage is limited to 12 hours per week unless approved by the screening staff.
- You may not sign up to use two instruments simultaneously.
- Time slots of 8, or 12 hours are ideal. If you only require a block of one or two hours, we recommend that you schedule for the morning or late afternoon.
- To modify or cancel your reservation, contact the screening staff at iccb_screen"AT"hms.harvard.edu.
- Please provide your email and a phone number where you may be reached.
Go to Instrument Sign-ups to Schedule time for the following instruments and workstatiouns:
cellWoRx
Image Xpress Micro
Olympus Fluoropoint
AutoScope
Image Analysis 1
Image Analysis 2
Image Analysis 3
Image Analysis 4
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