Figure 1.
An overview of non-covalently fibrin clot-bound plasma proteins.
Plasma clots were made by adding CaCl2, thrombin and aprotinin to platelet-poor citrated normal plasma, unbound proteins were washed away and bound proteins were extracted. These proteins were separated with 2D gel electrophoresis and visualized by Sypro Ruby. A) The numbers and arrows indicate the protein spots that were excised from gel and analyzed with mass spectrometry. B) The trains of spots that resemble the same protein are indicated by white ellipses. They include: fibronectin (I), α2-macroglobulin (II, III and VIII), plasminogen (IV), FXIII A chain (V), albumin (VI), α1-antitrypsin (VII), apolipoprotein J (IX), apolipoprotein E, HFREP-1 (X) and apolipoprotein A-I (XI). C) A zoomed image of the 2D gel with a lower fluorescent signal. The isoforms of α1-antitrypsin (A), apolipoprotein J (B) and apolipoprotein A-I (C) are indicated by white ellipses.
Table 1.
Mascot analysis of fibrin clot-bound proteins.
Figure 2.
Western blot analysis with specific α2-macroglobulin antibodies.
Fibrin clot-bound plasma proteins were separated with 2D gel electrophoresis and analyzed with Western blot analysis using specific α2-macroglobulin antibodies. The arrows indicate the three different α2-macroglobulin trains that were also identified as α2-macroglobulin with mass spectrometry (protein spots 2, 3 and 9 in figure 1A and table 1). The molecular mass of the protein marker is indicated in kDa.
Figure 3.
Western blot analysis for apolipoproteins in purified fibrinogen.
Fibrinogen was isolated from plasma with immunoaffinity chromatography and run on SDS-PAGE. Different apolipoproteins were detected with Western blot analysis using specific antibodies. Lane 1: protein marker, lane 2: apolipoprotein A-I (Mw = 28,900), lane 3: apolipoprotein J (Mw = 37,000), lane 4: apolipoprotein A-II (Mw = 8,700). Only the relevant section of the gel is shown.