High-resolution capillary electrophoretic separation of proteins and peptides was achieved by coating the inner wall of 75 microm ID fused-silica capillaries with 40-140 nm polystyrene particles ...which have been derivatized with alpha-omega-diamines such as ethylenediamine or 1,10-diaminodecane. A stable and irreversibly adsorbed coating was obtained upon deprotonation of the capillary surface with aqueous sodium hydroxide and subsequent flushing with a suspension of the positively charged particles. At pH 3.1, the detrimental adsorption of proteins to the capillary inner wall was suppressed efficiently because of electrostatic repulsion of the positively charged proteins from the positively charged coating which enabled protein separations with maximum efficiencies of 400000 plates per meter. A substantial improvement of separation efficiency in particle-coated capillaries was observed after in-column derivatization of amino functionalities with 2,3-epoxy-l-propanol, resulting in a more hydrophilic coating. Five basic and four acidic proteins could be separated in less than 7 min with efficiencies up to 1900000 theoretical plates per meter. Finally, coated capillaries were applied to the high-resolution analysis of protein glycoforms and bioactive peptides.
Adhesion and proliferation of bovine aortic endothelial cells on polystyrene derivatives having monoamine or diamine side chain was investigated focusing on the chemical structure of amino groups. ...Copolymers, SE8.5, is composed of polystyrene with 8.5 mol% of monoamine side chains, and SED8, which is with 8 mol% of diamine side chains, were estimated to contain almost the same amount of protonated amino groups in bulk composition at physiological pH (pH 7.4). There observed significant difference in cellular spreading of attached endothelial cells between these two types of copolymer surfaces. Spreading-% of attached cells on SED8 surfaces was approximately 1.6 times greater than that on SE8.5 6 h after seeding. This difference in cellular spreading influenced to subsequent cell growth. Cellular growth on each polymer surface was featured by parameter k, which corresponds to the 'rate constant' of cellular proliferation. While the k-value for SE8.5 decreased with decreasing seeding density as well as the case for polystyrene, SED8 maintained a high k-value even at low seeding density as 2 x 10(3) cells/cm2. These results suggest that cells may recognize the difference in the chemical structure of amine side chains of SE and SED copolymers.
Modified polystyrene resins containing sulphonate groups and tyrosyl sulphamide or tyrosyl methyl ester sulphamide groups have been investigated with respect to their potential for selective binding ...of anti-Factor VIII inhibitory antibodies from plasma. Adsorption of total immunoglobulin G and of a monoclonal antibody to Factor VIII was measured following addition of the radioiodinated proteins to normal plasma, plasma depleted of Factor VIII by adsorption on a resin coupled to anti-Factor VIM antibody, and haemophiliac plasma containing various levels of inhibitory anti-Factor VIII antibody. Depletion of anti-Factor VIII antibody from the haemophiliac plasmas by incubation with the resins was also measured by Bethesda assay. The modified resins and their corresponding unmodified ‘controls’ showed similar binding of total immunoglobulin G. However, only resins containing either sulphonate or a combination of sulphonate and tyrosyl sulphamide groups showed evidence of selective adsorption of anti-Factor VIII antibody from plasma.
Human anti‐factor VIII antibodies (anti‐FVIII) neutralize Factor VIII (FVIII) procoagulant activity. These antibodies appear in about 5–15 per cent of severely affected patients with haemophilia A ...treated with FVIII concentrates (Mannucci, 1993). In order to obtain non‐thrombogenic materials able to interact specifically with anti‐FVIII, amino acids residues that mimic part of the FVIII molecule recognized by anti‐FVIII have been grafted. Several cross‐linked polystyrenes were functionalized with sulphonate and tyrosine sulphamide groups or tyrosine derivatives sulphamide groups such as methyl ester tyrosine, or the peptides aspartic acid methyl amide tyrosine, tyrosine aspatic acid methyl amide or aspartic acid aspatic acid methyl amide tyrosine.
The in vitro removal of anti‐FVIII from haemophilic A plasma was performed on different supports. These polymers exhibit strong and selective affinity for the anti‐FVIII. The amont of adsorbed anti‐FVIII varies with the composition of the polymer and a maximum is achieved for 15–35 per cent of amino acid sulphamide groups. The influence of different chemical groups on the surface of the polymeric solid supports on the adsorption of anti‐FVIII was also studied.
