Platelet retention test

This chapter deals with a specific test of blood-surface interaction in vitro platelet retention in a column of beads (due to platelet adhesion and aggregation). Protein adsorption precedes platelet adsorption, and thus the in vitro platelet retention test involves competitive and sequential adsorption of proteins, the outcome of which produces surfaces having widely varying degrees of platelet retention. Except in the case of thrombin (3), plasma protein absorption on these surfaces has not been studied. 

Concentration of the polymer, temperature, and evaporation conditions under argon were maintained as closely as possible, specially with casting microscope slides and glass beads, so that the surface morphology in ESCA and platelet retention tests were about the same. 

Figure 1. In vitro test for platelet retention, using plastic columns, packed with fine glass beads whose surfaces are thereafter coated with test polymer (ambient temperature = 37°C).

We tested the hypothesis that the platelet retention index should increase as polymer composition varied from 100 mol % methyl acrylate to 100 mol % styrene, the respective platelet retention indices p being about 0.25 and 0.55 for the homopolymers. The results are shown in Figure 3, wherein p increases to a maximum near 40% styrene. When the surfaces were incubated in platelet-poor plasma before contact with whole blood, the values of p were much reduced (from 0.25 to 0.05 for methyl acrylate) for copolymers containing up to 60 mol % styrene. Copolymers of higher styrene content were not rendered significantly less retentive by plasma pretreatment. 

Thus, time of water exposure prior to challenging platelets is probably important, but the matter needs further study. Our tests suggest that increasing the hydrophobic side group length increases platelet retention in these experiments surfaces were exposed to isotonic saline solution for less... 

Molecular models (2) of the hard-segment phase of segmented polyether polyurethanes tested in the in vitro bead column (4) show a high platelet retention index (p = 0.8) regardless of molecular composition. The variable p is defined as the fraction of platelets in whole citrated human blood entering a test column that are retained on the bead surfaces, averaged for... 

Parallel tests show that thrombin adsorption is minimal on well-prepared polyurethanes containing amorphous PEO, greater on PTMO, and very high on analogues of the hard segment (diisocyanate-diamine copolymers), thus paralleling the trends in platelet retention index p. This result is consistent with the postulate that protein adsorption must precede platelet adsorption. 

These polyurethanes (and their precursors or analogues) were cast from an appropriate solvent, usually DMF, onto the glass bead surfaces used in the in vitro test for platelet retention (4), or for the thrombin absorption test used previously (2). Crystals of KBr for Fourier transform infrared (FTIR) spectroscopy and glass microscopic slides for examination by XPS (ESCA) served as supports for polymers cast from the same solvents. Concentration of polymer (5 wt %), temperature of casting... 

C), and subsequent slow evaporation conditions under argon gas (40°C, 21 days) were maintained as closely identical as possible in preparing surfaces for each test (FTIR, ESC A, and platelet retention). The air side of the film was the side exposed to platelets and examined by XPS. The other side (against the glass substrate) was not exposed. 

Platelet retention in the polymer-coated bead columns was quantified by noting the percentage of platelets removed from aliquots 2 through 5 of 5 successive 1 ml effluent fractions. (Aliquot 1 was diluted with saline and was not counted.) Retention values obtained for the 14 polymers tested are shown in Table 1. These data are plotted in Fig, 2 as a function of both the alkyl side chain length and the glass transition temperatures (Tg s) of the polymers. 

ESCA examination) and on glass beads used in the Salzman "in vitro" test for platelet retention. ... 

Dion et al. (1993) evaluated the in vitro platelet retention of the new prosthetic heart valve that was designed by FII Company and Pr. Baudet, composed of Ti6A14V titanium alloy coated with DLC (obtained by CVD). The retention and adhesion of platelets was evaluated by analyzing radioactivity on the exposed wall of test or control tubes through which a blood cell suspension containing In-labelled platelets had circulated. Their results showed that on DLC/Ti6A14V, platelets adhered twice the amount that they did on the reference material (a silicone medical-grade elastomer the behaviour of which in contact with blood is the same as that observed with the National Institutes of Health-recommended polydimethyl siloxane) (Dion et al., 1993). 

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