User:Igor R. Kuznetsov/Notebook/RBL-2H3/2010/07/07

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U937 Delta ST on VCAM-1

Cells from Alex on 07/01 (one week in growth media). Coverslips: base-treated as per Andrew's protocol, then coated with Recombinant Human VCAM-1/CD106 Fc Chimera (R&D Systems #P19320) for 30 minutes at room temperature, and gently washed 2 times with Hepes buffer. Plated cells resuspended in Hepes on the coverslips for 60 minutes at room temperature, then washed with Hepes two times to remove unattached cells.

Sample 1: Sells resuspended in Hepes on untreated coverslip. Cells are unattached and globular in shape.

Image:070710_01.jpg


Sample 2: Untreated coverslip coated with VCAM, after 1 hour and 2 gentle washes with Hepes buffer. Only a few cells can be detected: most have been washed off. Cells appear to be attached, but not spreading.

Image:070710_02.jpg


Sample 3: Base-cleaned coverslip coated with VCAM, after 1 hour and 2 gentle washes with Hepes buffer. Plenty of cells can be detected. Cells appear to be attached, but only a few (about 1:100) are spreading. Those that are, spread into symmetric fried egg shape, while those that are not remain globular in shape.

Image:070710_03.jpg


Sample 4: Sample 3 after another hour (2 hours total). Cells that were spreading continue to do so in a symmetric fried egg shape, with plenty of cytoplasmic activity. Majority of the cells remain attached but not spreading.

Image:070710_04.jpg


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