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ADAM12 Inhibits Cell Migration on an α4β1- but Not on an α5β1-specific FN Fragment
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(B) CHO-α4/GFP cells were transfected as indicated and then analyzed in scratch wound assays, as described in the legend to Figure 2, except that 24-well dishes were coated with 2 μg/ml CS-1. Similarly, expression of ADAM12 in CHO-α4/GFP cells caused a reduction in cell motility in both transwell (Figure 2C) and scratch wound (Figure 2D) assays. Expression of ADAM12 reduced the motility of CHOB2-α4 cells in both transwell (Figure 2A) and scratch wound (Figure 2B) assays. After three washes, adherent cells were stained with crystal violet and solubilized with 1% SDS. After 45-min incubation on ice, cells were washed twice with PBS, resuspended in 0.3 ml of 2% paraformaldehyde in PBS, and analyzed on a FACSCalibur benchtop analyzer at the Flow Cytometry Core Facility at University of Virginia. Primary antibody (mouse anti-human α4, HP2/1), diluted to 20 μg/ml in blocking buffer (PBS/fetal bovine serum/bovine serum albumin), was incubated with cells for 45 min at 4°C.