Albumin, human serum fractionated

FIGURE 12.5 Human serum tryptic digest analysis. Fractionation in the first LC dimension was performed using a C18 column at pH 10. Fractions were analyzed using NanoEase 0.3 x 150 mm Atlantis d18 column. Approximately 66 lg (400 pmole of semm albumin peptides) was injected on column. Arrow points to a selected albumin peptide illustrating a local column mass overloading. Ten-5mm wide fractions were collected in 1st LC dimension. 

A report on the binding of the anesthetic propofol to human serum albumin and to plasma presents a dataset that challenges simple notions of equilibria [70]. The unbound fraction of propofol was found to increase sharply at low drug concentrations. The authors appear to have carefully eliminated possible artifacts. Explanations based on cooperative binding modes are discussed though no clear explanation emerges. 

Most frequently, binding protein is added to the incubation mixtures as either serum or purified serum albumin. With human serum albumin, at equilibrium, the acceptor substrate will largely be protein-bound, when the bilirubin albumin molecular ratio is smaller than one (the dissociation constant of the first binding site of purified human serum albumin is approximately 7 X 10 M with 2 X 10 M for two additional binding sites) (J2). The first binding site of albumin, measured with rat serum, has a dissociation constant of about 5 X 10" M (M8). The unbound fraction will normally not exceed the very low solubility of the pigment. Addition of albumin to an alkaline solution of bilirubin, or its addition immediately after neutralization, prevents colloid formation, if the bilirubin albumin molecular ratio is smaller than one (B25). However, colloidal bilirubin, once formed, cannot be redissolved by the addition of albumin (B26). 

Fig. 62a, b, and c. Some staining techniques adapted to paper electrophoresis. Sample human serum. Run continuous electrophoresis. The fractions are labeled according to the proposed reference system (see text). The known fractions are +86.20 Albumin +86.18 oij-globulin +86.12 a2-globulin +86.07 3-globulin —86.02 Y-globulin. 

In a follow-up study, the same authors examined the applicability of the same device for relevant protein samples and investigated the main contributions to band broadening [82]. As a consequence of the small depth of the beds, zone spreading caused by Joule heating was shown to be negligible (see Sect. 3.1.1). Cross fields of up to 100 V/cm were applied for the separation of human serum albumin, ribonuclease A and bradykinin. The feasibility of fraction collection was demonstrated with four collected fractions of a whole rat plasma sample. Off-line analysis of these four isolated fractions by CE indicated the separation of serum albumins and globulins. 

Human Serum Albumin (HSA, Siga, Fraction V Powder) 3.0 g in 100 mL distilled water, prepare 3 mL aliquots and store at -20°C. 

The resolution of clinical grade human serum albumin (HSA) increases as the operating pH of the column approaches the pi of HSA.28 Eight peaks have been resolved with a neutral coated capillary. The mass spectral analysis of these fractions indicated that major component is HSA with a free Thiokol at cystein 34, a variant having cystein 34 blocked cysteine, amino-terminally... 

Plasma Protein Binding (PPB). The fraction bound to plasma proteins (%PPB) can be easily calculated from binding affinity to HSA under the assumption that binding occurs mainly to the Human Serum Albumin (HSA). Thus we can ignore differentiation of both these properties. A dataset of 94... 

Normal serum was pooled from normal donors, stored for several hours at 37 °C, and frozen in glass tubes. Rabbit antisera to the following human proteins From Hyland, antisera to albumin (list 071-107), fibrinogen (list 071-108), human fraction III-I (list 071-103), -lipoproteins (list 071-113), human serum (list 071-121), Fab (071-258), Fc (071-259). From Mann Research Labs., antisera to 7s gamma-globulins (cat. 231) and total gamma-globulins (cat. 8090). From Hoechst, Woodbury, N. Y. antiserum to pre-albumin (code 8506). 

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