Paper electrophoresis of thyroxine in tris‐maleate buffer

Myant, N. B.; Osorio, C

doi:10.1113/jphysiol.1960.sp006511

Cited by 29 publications

(16 citation statements)

References 8 publications

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“…As has recently been reported (13,14), essentially no binding of labeled T4 was noted in the area anodal to albumin when normal serum was subjected to When resin eluates containing sufficient prealbumin and TBG to provide visible spots after bromphenol blue staining were studied by paper electrophoresis at pH 7.4, prealbumin retained its localization anodal to albumin. Binding of T4 was localized, however, entirely to the TBG zone.…”

Section: Alpha Globulinmentioning

confidence: 59%

Observations Concerning the Binding of Thyroid Hormones by Human Serum Prealbumin*

Ingbar¹

1963

J. Clin. Invest.

116

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There is general agreement that thyroxine (T4)1 is the major component of the thyroid hormone in blood, where it exists almost entirely in a dissociable complex with certain plasma proteins. From the numerous studies that followed the discovery of the TU-binding (a)-globulin of plasma (TBG) in 1952, it first appeared that the binding of T. in plasma could be mainly ascribed to binding sites of high avidity but low concentration, contributed by TBG. A secondary role was ascribed to binding sites of low avidity but almost unlimited concentration, contributed by albumin. These conclusions, based almost entirely on data obtained by electrophoretic separation of serum or plasma proteins on filter paper in barbital buffer at pH 8.6, have been considered in several recent reviews (1, 2).In 1956, during efforts to purify TBG, an additional T4-binding protein was observed in subfractions of human plasma proteins. Subsequent experiments revealed that a T4-binding protein of similar electrophoretic mobility could be consistently demonstrated in unfractionated normal human serum when electrophoretic separations were performed in a buffer containing tris (hydroxymethyl)aminomethane (Tris) and maleic acid, rather than barbital. Because of its rapid anodal migration at pH 8.6, this protein was termed the thyroxine-binding prealbumin (TBPA) (3). It avidly bound T4, but failed to bind significant quantities of 3,5,3'-triiodothyronine (T3).

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Section: Alpha Globulinmentioning

confidence: 59%

Observations Concerning the Binding of Thyroid Hormones by Human Serum Prealbumin*

Ingbar¹

1963

J. Clin. Invest.

116

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“…19) The mean concentration of TBPA in euthyroid subjects was found to be 37mg per 100ml serum, somewhat higher than the value reported by Oppenheimer et al 23 It is known that there are two bands (PA-1 and PA-2) which have a faster mobility than albumin on starch gel electrophoresis. TBPA is considered to reside in the PA-1.25-27 The concentration of TBPA on polyacrylamide gel electrophoresis may be slightly higher than that on starch gel electrophoresis because it contains both PA-1 and PA-2 fractions.…”

Section: Polyacrylamide Gel Electrophoresiscontrasting

confidence: 38%

Polyacrylamide Gel Electrophoretic Study on the Effect of Estrogen on Human Thyroxine-binding Prealbumin

Sato¹,

Saito²,

Inagaki³

et al. 1967

Tohoku J. Exp. Med.

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Ethinyl estradiol was administered orally in a daily dose of 0.18-0.3mg for seven days to subjects with normal and abnormal thyroid function, and the changes of several parameters of the thyroid function during estrogen admin istration were investigated.The concentration and maximum T4 binding capacity of thyroxine-binding prealbumin (TBPA) were estimated by polyacrylamide gel electrophoresis.Estrogen These results are observed equally in subjects with intact and abnormal thy roid functions, and it is concluded that these phenomena have no relation to the pituitary-thyroid system.Many investigators have demonstrated an increase in the capacity of thyroxine-binding globulin (TBG) and in the serum PBI level during pregnancy1-4 and by administration of estrogen. 5, 6 However, no marked clinical manifestations of hyperthyroidism were observed in these instances,1 and it was supposed that free thyroxine, the biologically active form of thyroxine in the blood, remained normal even in these situations.In the present investigation, thyroxine-binding protein (TBP) was studied by the polyacrylamide gel electrophoresis and free thyroxine was determined by the method of Lee and Henry7 in subjects to whom large doses of estrogen (ethinvl estradiol) was administered.A portion of the findings was presented elsewhere in abstract form.'

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“…The partition of thyroxine at a physiological pH would therefore appear to be in reasonable agreement with the distribution of thyroxine as determined by paper electrophoresis at pH 8.6. On the other hand, if filter-paper electrophoresis is carried out at pH 7.4, no thyroxine will be found in association with prealbumin (24,25). Thus, one is led to the apparently paradoxical conclusion that the physiological distribution of thyroxine is more accurately mirrored by the results of paper electrophoresis at pH 8.6 than at pH 7.4.…”

Section: Discussionmentioning

confidence: 78%

Binding of Thyroxine by Serum Proteins Evaluated by Equilibrium Dialysis and Electrophoretic Techniques. Alterations in Non-Thyroidal Illness*

Jh¹,

Squef²,

Mi³

et al. 1963

J. Clin. Invest.

243

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Considerable evidence suggests that the concentration of free thyroxine, circulating unbound to serum proteins, is more closely relattd to a subject's thyroidal status than is the concentration of total thyroxine (1). Because of the intense binding of thyroxine by the serum proteins, the concentration of free thyroxine is very small and not measurable by conventional chemical methods. Robbins and Rall (2), on the basis of electrophoretic data and the binding characteristics of bovine serum albumin, estimated that the concentration of free thyroxine in normal subjects was in the order of 6 x 10-1" M. By equilibrium dialysis of whole serum, Sterling and Hegedus (3) arrived at a figure approximately twice this. Other techniques for assessing the relative concentrations of free thyroxine in various clinical and physiological states include measurement of the fractional rate of dialysis of I'1"-labeled thyroxine (T4-I131 ) across a semipermeable membrane from one serum compartment to another (4, 5), and simultaneous determination of red-cell uptake of I131-labeled triiodothyronine (T3-"131) and serum protein-bound iodine (PBI) (6).In the present study, we used two independent methods to provide a relative measure of the fraction of thyroxine bound to serum proteins and the concentration of free thyroxine in serum. One method is based on the principle of equilibrium dialysis of diluted serum in an aqueous buffer at pH1 7.4; the other involves determination of critical ratios by filter-paper electrophoresis of whole serum in a glycine-acetate buffer at pH 8.6. Such an electrophoretic system reveals three species of protein-binding sites: thyroxine-binding globulin (TBG), albumin, and thyroxine-binding prealbumin (TBPA). Applied to the study of sera of normal and pregnant subjects and patients with hypothyroidism and hyperthyroidism, these methods gave results that were consistent internally and also with the current view that concentrations of free thyroxine are elevated in hyperthyroidism, depressed in hypothyroidism, and normal in pregnancy. The investigation was thereupon extended to a more intensive study of thyroxine-binding by the serum proteins of patients with nonthyroidal disease. Ingbar and Freinkel (7) have noted that in patients with a variety of diseases, a reduced quantity of thyroxine was associated with TBPA, and this finding suggested to us the possibility that such alterations could influence the level of free thyroxine in serum. METHODS Equilibrium dialysisThe method of these studies is partly based on the dialysis procedures used by Sterling and Tabachnick in their investigation of thyroxine-binding by serum albumin (8). Analogous techniques have been used to study the interaction of certain drugs with thyroxine-binding proteins (9-11). T4-IP`,I tested for chromatographic purity as previously described (9), was added to the serum sample in quantities sufficient to increase the endogenous concentration of thyroxine by 1 ,ug per 100 ml. The mixture was allowed to equilibrate for 30 minutes at room tem...

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Paper electrophoresis of thyroxine in tris‐maleate buffer

Cited by 29 publications

References 8 publications

Observations Concerning the Binding of Thyroid Hormones by Human Serum Prealbumin*

Observations Concerning the Binding of Thyroid Hormones by Human Serum Prealbumin*

Polyacrylamide Gel Electrophoretic Study on the Effect of Estrogen on Human Thyroxine-binding Prealbumin

Binding of Thyroxine by Serum Proteins Evaluated by Equilibrium Dialysis and Electrophoretic Techniques. Alterations in Non-Thyroidal Illness*

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