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Transwell Plating for better Imaging

We have developed a solution for this problem.  We plate our cells on the basal surface of the transwell insert in the following manner.

1. A bio-compatible piece of tubing is cut into a small cylinder of suitable geometry as to provide a shallow well and fluid barrier when placed over the distal end of the insert.

2. A plug made of silicon is placed into the tubular portion of the insert to prevent leakage through the membrane.

3. The insert is inverted (membrane side up) and cells are poured into the well defined by the cylindrical tubing surrounding over the membrane.

4. The cells are returned to the incubator and allowed to plate in this inverted orientation for 45 minutes.

5. The plug and tubing are removed from the insert and the insert is returned to the tray with appropriate media and allowed to divide until confluent.  We have had much success with this plating technique.

6. When the cells are confluent the insert is placed into a Bioptechs Delta T4 Transwell Adapter which is then lowered into a Delta T dish on the microscope.   The cells are now facing down for easy imaging through the coverglass bottomed self-heating Delta T dish.  See Bioptechs web site for details on equipment and contact Bioptechs directly for additional information regarding membrane insert micro-observation accessories. 

7. Cells on the membrane can be perfused on either the apical or basal surface during microscopy if necessary.

Dan Focht
Bioptechs
3560 Beck Road
Butler, PA 16002
dan@bioptechs.com
www.bioptechs.com


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Copyright 2001-2014  Dr. M. Chua, Program in Molecular Biology & Biotechnology, School of Medicine, University of North Carolina, Chapel Hill, NC 27599
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Last Updated: 2014-07-23