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Blood, Vol. 110, Issue 7, 2475-2483, October 1, 2007
Requirement of  and ß subunit transmembrane helix separation for integrin outside-in signaling
Blood Zhu et al.
110: 2475
Supplemental materials for Zhu et al. Vol. 110, Issue 7, 2475-2483
Files in this Data Supplement:
- Figure S1. mAb LM609 efficiently blocks αVβ3-dependent adhesion (JPG, 67.3 KB)
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CHO-K1 cells transfected with either  IIb and  3 integrin (IIb3 CHO) or 3 integrin alone (3 CHO (V3)) were allowed to adhere to fibrinogen (10 µg/mL)-coated surfaces in the presence of either 1 mM Ca2+/1 mM Mg2+ or 5 mM EDTA and in the absence or presence of indicated function-blocking antibodies (at 10 µg/mL). After washing, the amount of bound cells was determined by measuring the lactate dehydrogenase (LDH) activity as described in “Materials and methods, Cell adhesion.” Results show that adhesion of IIb3 CHO-K1 cells was inhibited by approximately 90% with mAb 10E5 (IIb3 functional inhibitor), by approximately 10% with mAb LM609 (V3 functional inhibitor), and by 100% with 10E5 and LM609 together. For 3 CHO (V3), LM609 blocked 100% of adhesion.
- Figure S2. Spreading of 293T cell transfectants on immobilized fibrinogen (JPG, 216 KB)
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WT or mutant 968C/693C receptors were transiently transfected into 293T cells. Green fluorescent protein (GFP) was cotransfected to select visually the transfected cell area by microscopy. 293T cells were incubated in wells coated with fibrinogen at 37°C for 1 hour, and GFP signals were analyzed by fluorescence microscopy. The WT cells spread on the coated surface efficiently, whereas cells transfected with the mutant receptors did not spread. Scale bar, 10 µm. Images were acquired as in “Materials and methods, Cell spreading and microscopy.”
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