Cortisol-binding Capacity Research Articles

Diurnal variation of prednisolone binding to serum corticosteroid-binding globulin in man.

Serum corticosteroid-binding globulin (CBG, transcortin) binding capacity for prednisolone and for cortisol, and levels of the circulating cortisol were evaluated in healthy adult subjects and in patients under long-term treatment with prednisone. Blood samples were drawn at 4-hr intervals throughout at least 24 hr; the prednisone administration was discontinued 24 hr before starting examination. Circadian fluctuations in the CBG-binding capacity for prednisolone occurred in healthy subjects: maximum binding was at 12 midnight and minimum binding at 8 a.m. The diurnal pattern of prednisolone-binding capacity was characterized by phase shifts of approximately 12 hr and 8 hr compared with those of serum cortisol and cortisol-binding capacity, respectively. There were no diurnal variations of prednisolone- and cortisol-binding capacity in the prednisone-treated patients. Moreover, the levels of both capacities, expressed as micrograms bound steroid/100 ml, were significantly reduced with respect to normal control subjects. Cyclic variations in serum CBG-binding capacity may represent an additional factor accounting for temporal differences in the action of synthetic corticoids which are bound by the transport protein. Synthetic derivatives when chronically administered may interfere not only with the adrenal secretion but also with the serum transport of glucocorticoids.

A double adsorption technique for the determination of corticosteroid-binding globulin in human serum (author's transl)

Despite the widespread use of many different estrogenic compounds in hormonal contraception and estrogen replacement, in the human a simple test for comparing the potencies of the compounds is lacking. Many of the estrogenic compounds are difficult to measure, and animal studies may not relate directly to those done on humans in terms of potency. Corticosteroid-binding globulin (CBG) in human serum has been known since 1958 to increase in response to estrogen administration, and the degree of its elevation has been shown to be closely dose-related and reasonably sensitive. Therefore, CBG represents a biological parameter that provides for the specific quantal analysis of estrogenicity in the human. In this paper a simple and reliable method for the determination of the cortisol-binding capacity of CBG (CBG-BC) in human serum is described. The procedure is based on the amount of cortisol required to saturate the CBG binding sites. All endogenous steroids are initially removed from the serum by adsorption on dextran-coated charcoal (DCC). A mixture of a known amount of unlabeled and labeled cortisol is added to the sample in order to saturate the "stripped" binding sites of CBG. The unbound cortisol is removed by DCC at 4 degrees C, and the supernatant represents the total cortisol-binding capacity of the serum sample. To distinguish between cortisol-binding to CBG and to albumin, "stripped" serum is heated for 30 min at 60 degrees C, the temperature which does not influence cortisol binding by albumin but which does inactivate CBG. The samples are subsequently equilibrated with a mixture of labeled and unlabeled cortisol. The free cortisol is removed by DCC and the bound cortisol remaining in the supernatant is designated as the albumin-bound cortisol. Thus, CBG-BC is assayed by a double adsorption technique as differential binding to native and heat-inactivated serum. The cortisol determined by this method corresponds to CBG-bound cortisol, and CBG-BC is expressed as micrograms of cortisol bound per 100 ml serum. The precision and accuracy of the method were found to be good. The values of CBG-BC obtained by this method were 14.9 +/- 1.9 (SD) microgram/100ml in adult men and 16.0 +/- 1.8 microgram/100ml in adult women. Daily serum samples in five women revealed no significant variations in CBG-BC throughout the normal menstrual cycle despite fluctuating levels of serum estradiol. CBG-BC was also found not to vary significantly with age. In pregnancy, however CBG-BC increased significantly.

The Metabolic Clearance Rates and Interconversion of Cortisol and Cortisone in Pregnant and Nonpregnant Baboons

Pregnancy alters the pattern of maternal cortisol (F) metabolism and increases the maternal serum cortisol-binding capacity (CBC) of baboons. To determine whether these changes are associated with alterations in F clearance,the metabolic clearance rate (MCR) and interconversion (p) of F and cortisone (E) were measured by continuous infusion of (3H)F and (14C)E in 9 regularly menstruating and 7 pregnant baboons (Papio papio). In nonpregnant animals, the values (X +/- SE) for MCR-E (488 +/- 48 1/day) were greater (P less than 0.001) than those for MCR-F (214 +/- 22 1/day). The p value for the conversion of E leads to F (62.8 +/- 4.7%) was greater (P less than 0.001) than that for the reaction F leads to E (41.6 +/- 3.7%), indicating that F formation is favored. Consistent with MCR-E greater than MCR-F, the per cent of F bound to proteins other than albumin (75 +/- 2) was greater (P less than 0.001) than the per cent of E bound (52 +/-3). The production rate (MCR x peripheral concentration; mug/min) of F (55.1 +/- 7.9) was greater (P less than 0.001) than that of E (28.5 +/-3.9) with essentially all of the F being secreted directly (secretion rate 51.2 +/- 7.9 mug/min). Essentially all of the E produced was derived from circulating F, vitually none being secreted directly (secretion rate 4.6 +/- 3.9 mug/min). Pregnancy did not alter the MCR-F (190 +/- 23 1/day), MCR-E 525+/- 51 1/day), per cent of F (79 +/- 3), or per cent of E (49 +/-3) bound,or F (57.2 +/- 9.2 mug/min) or E (35.5 +/- 4.9 mug/min) production rates. CBC (mug F/100 ml) was significantly (P less than 0.01) elevated (25.3 +/- 2.3, nonpregnant vs 35.1 +/- u.6, pregnant). In addition, p E leads to F was increased (75.5 +/- 1.8%) as was p F leads to E (54.3 +/- 3.7%; P less than 0.01). We have concluded that the MCR-F during pregnancy is more dependent on alterations in maternal metabolism than on the increased serum CBC characteristic of gestation. We suggest that the latter factor may be important in regulating the physiologic levels of the other steroids which bind to it.

Continuous oral low-dosage cyproterone acetate for fertility regulation in the male? — A trend analysis in 15 volunteers —

As part of the “WHO Task Force Program on Regulation of Male Fertility”, a study has been carried out to investigate the effects of 10 or 20 mg cyproterone acetate (CPA) daily over 26 weeks in 15 healthy young males. Control periods of 18 and 16 weeks, respectively, preceded and followed the phase of CPA medication. The salient observations under treatment were: A decrease in sperm density and in normal-shaped sperm with conversely an increase of pathological and immature forms, an increase in dead spermatozoa, a relative reduction of motile spermatozoa, a distinct drop in the speed of spermatozoa and a distinct decrease in the ability of the spermatozoa to migrate through the cervical mucus of ovulation quality in vitro . Androgen levels (mainly testosterone) in blood plasma dropped significantly to about 40% of the basal values, without concurrent effects on sexual behaviour; no changes in plasma LH and FSH were observed; alkaline phosphatase values in seminal plasma were found to be considerably elevated during treatment; no changes in fructose and sialic acid levels in seminal plasma were seen. CPA also increased significantly the cortisol-binding capacity of transcortin. The investigations clearly demonstrate a distinct inhibitory influence of low-dosage CPA on several parameters of male fertility. These reversible effects appear to indicate that low doses of CPA might be used for fertility control in the male, but final conclusions in this respect cannot be drawn before a large scale phase three study on this issue has been performed.

Cortisol binding capacity and oestrogen concentrations in maternal and cord plasma in pregnancies with normal and anencephalic fetuses.

The cortisol binding capacity of maternal and cord plasma samples obtained at delivery from fifteen women and their normal infants and from seven women and their anencephalic infants was measured at 4 degrees C by a gel filtration technique. The concentration of oestrogen in these samples was measured by radioimmunoassay. There was no significant difference (t test) between the cortisol binding capacity of peripheral plasma from women with normal infants (1-55 +/- 0-24 mumol/1, mean +/- SD) and from those who delivered anencephalic infants (1-35 +/- 0-30 mumol/1), nor between the cortisol binding capacity of cord plasma from anencephalic infants (0-47 +/- 0-04 mumol/1) and that of normal infants (0-37 +/- 0-10 mumol/1). However, mean oestrogen concentrations in maternal and cord plasma from the pregnancies with an anencephalic fetus were significantly lower (P less than 0-01) than in the corresponding samples from normal pregnancy. It is concluded that oestrogen concentrations in maternal and cord plasma in normal pregnancy at delivery are much greater than those required to account for the increase in plasma cortisol binding capacity. Since plasma cortisol binding capacity in pregnancy with an anencephalic fetus is not diminished, the reduced excretion of corticosteroids relative to normal pregnancy in this condition is unlikely to be due to alterations in cortisol metabolism associated with a lower plasma cortisol binding capacity.

Effect of malnutrition on cortisol-binding protein in the lens of cynomolgus monkeys.

A cortisol-binding protein exists in the lens of cynomolgus monkeys as in other species. The cortisol-binding capacity decreased markedly with a diet either moderately or severely depleted of protein, even if the diet was later changed to a normal diet during infancy. This suggests that malnutrition may cause insufficient synthesis of the important factors controlling the inactivation of steroid hormone in the lens.

Protein binding of plasma cortisol in the foetal lamb near term.

Binding of cortisol to plasma proteins was studied in the foetal lamb by equilibrium dialysis at 37 degrees C. At 122 days of pregnancy the mean level of transcortin expressed as cortisol-binding capacity was 28 +/- 6 (S.D.) ng cortisol/ml plasma. During the last 14 days of pregnancy there was a progressive increase in transcortin-binding capacity to 85 +/- 14 ng cortisol/ml plasma. A sharp increase in the concentration of both protein-bound and unbound cortisol was observed over the same period. A rise in the concentration of total cortisol from around 3 to 42 ng/ml was associated with an increase in unbound cortisol from 0-2 to a maximum of 2-1 ng/ml. The concentration of albumin-bound cortisol was approximately equal to that of unbound cortisol. The mean value for the transcortin-cortisol affinity constant was 1-15 x 10(8) l/mol. It is concluded that an increase in transcortin-binding capacity is partly responsible for the prepartum increase of corticosteroid levels observed in normal foetal lambs.

Serum cortisol binding capacity and cortisol concentration in the pregnant baboon and its fetus during gestation.

The cortisol binding capacity of serum from 11 pregnant baboons (38 samples) and from 7 baboon fetuses delivered prematurely or at term was measured after removal of endogenous steroids. Values for maternal serum collected between 60 and 120 days after mating (59.0 +/- 6.4 mug/100 ml, mean +/- SD) were greater than those for serum collected at term (42.3 +/- 4.9 mug/100 ml). The cortisol-binding capacity of fetal serum collected between 100 and 132 days' gestation was similar to that of the corresponding maternal sample, but at term was only 50% of the maternal value. The rate of clearance of cortisol from both fetal and maternal serum may therefore increase progressively during the last trimester of pregnancy. This effect is likely to be more marked in the fetus. The cortisol binding capacity of 15 serum samples from 9 non-pregnant baboons was 33.4 +/- 5.5 mug/100 ml. Mestranol2 (administered 200 mug/day im for 15 days) significantly increased the serum cortisol binding capacity. The concentration of cortisol in maternal serum from 7 pregnant baboons (10 samples) was 44.0 +/- 8.4 mug/100 ml and was independent of the state of gestation. In fetal serum the cortisol concentration was 4 mug/100 ml before 168 days' gestation and reached 49 mug/100 ml after normal delivery at term. These findings suggest that the mechanisms for production of cortisol by the fetus mature as gestation progresses. The physiological significance of the marked difference between the cortisol concentration and the cortisol binding capacity of fetal serum awaits elucidation.

Cortisol binding in plasma of men with unipolar and bipolar affective disorders.

Following an earlier report, plasma cortisol-binding capacity (CBC) was estimated in a new series of 11 men with unipolar depression and 11 men with bipolar illness admitted to hospital. The two groups were comparable except that the bipolar group tended to have more frequent episodes of illness and to have more family histories of affective disorder. There was no significant difference between the CBC values in each group.

Cortisol Binding to Proteins in Plasma in the Human Neonate and Infant

Extract: The binding of cortisol to plasma proteins was studied by a methodology allowing the determination of the apparent affinity constant (Ka) and the specific binding capacity (Ns) for transcortin in different blood compartments in normal pregnant women at term, in the umbilical cord blood, and in normal neonates and infants throughout the first year of life. Removal of endogenous steroids appeared to be necessary for these binding studies, especially in the cord plasma. Both an equilibrium dialysis and an adsorbent technique were used, inasmuch as it has been established that they yield similar results for corticosteroid-binding globulin (CBG) binding parameters.The high affinity binding of cortisol, attributed to transcortin, was found to have the highest capacity in plasma from pregnant subjects (Ns = 1.16 ± 0.19 × 10−6 M), whereas the fetal compartment (umbilical cord) exhibited low values (0.25 ± 0.03 × 10−6 M). Thus a physiologic barrier to the transplacental passage of transcortin resulted in a large concentration gradient in cortisol-specific binding sites between the fetal and maternal compartments. No significant differences were found for transcortin capacity between venous uterine plasma and peripheral plasma in three subjects studied during cesarean section. A possible physicochemical difference between maternal and fetal transcortin appeared unlikely, because in all blood compartments studied, the apparent affinity constant, Ka, was similar (5-8 × 108 M−1 at 4°), and the behavior of the specific cortisol binder in polyacrylamide gel electrophoresis was the same in all cases.After birth, although the transcortin Ka for cortisol was still in the same range, Ns remained low during the first month of life (0.23 ± 0.07 × 10−6 M); after which an increase toward normal adult values was seen, with a rather large variation between individuals from 2-12 months of age (mean 0.48 ± 0.10 × 10−6 M). Children above 1 year exhibited adult values for Ns (0.51 ± 0.03 × 10−6 M).Speculation: From the similarity of values found for transcortin Ka and electrophoretic behavior, the physicochemical properties of the glycoprotein cortisol binder are likely to be the same both in the maternal and fetal compartments, although final evidence of chemical identity is not available. Thus, the different binding capacity between these compartments might be attributed to a different concentration in circulating transcortin. This concentration gradient may play a role in the transplacental transfer of corticosteroids during pregnancy and would favor passage toward the maternal compartment. The low transcortin level in the neonatal period might be attributed to a low biosynthesis capacity of the liver in the neonate, although nothing is known about the clearance rate of transcortin in the neonatal period. The transcortin level increases toward adult values during the first year of life and this would suggest either a maturation of the liver biosynthesis enzymatic systems or a biosynthesis stimulation. A possible contribution of a pituitary factor might be suggested on the basis of experimental work in the rat, but the meaning of this increase in cortisol binding capacity in plasma remains to be elucidated.

Cortisol Binding to Proteins in Plasma in the Human Neonate and Infant

Extract: The binding of cortisol to plasma proteins was studied by a methodology allowing the determination of the apparent affinity constant (Ka) and the specific binding capacity (Ns) for transcortin in different blood compartments in normal pregnant women at term, in the umbilical cord blood, and in normal neonates and infants throughout the first year of life. Removal of endogenous steroids appeared to be necessary for these binding studies, especially in the cord plasma. Both an equilibrium dialysis and an adsorbent technique were used, inasmuch as it has been established that they yield similar results for corticosteroid-binding globulin (CBG) binding parameters. The high affinity binding of cortisol, attributed to transcortin, was found to have the highest capacity in plasma from pregnant subjects (Ns = 1.16 ± 0.19 × 10−6 M), whereas the fetal compartment (umbilical cord) exhibited low values (0.25 ± 0.03 × 10−6 M). Thus a physiologic barrier to the transplacental passage of transcortin resulted in a large concentration gradient in cortisol-specific binding sites between the fetal and maternal compartments. No significant differences were found for transcortin capacity between venous uterine plasma and peripheral plasma in three subjects studied during cesarean section. A possible physicochemical difference between maternal and fetal transcortin appeared unlikely, because in all blood compartments studied, the apparent affinity constant, Ka, was similar (5-8 × 108 M−1 at 4°), and the behavior of the specific cortisol binder in polyacrylamide gel electrophoresis was the same in all cases. After birth, although the transcortin Ka for cortisol was still in the same range, Ns remained low during the first month of life (0.23 ± 0.07 × 10−6 M); after which an increase toward normal adult values was seen, with a rather large variation between individuals from 2-12 months of age (mean 0.48 ± 0.10 × 10−6 M). Children above 1 year exhibited adult values for Ns (0.51 ± 0.03 × 10−6 M). Speculation: From the similarity of values found for transcortin Ka and electrophoretic behavior, the physicochemical properties of the glycoprotein cortisol binder are likely to be the same both in the maternal and fetal compartments, although final evidence of chemical identity is not available. Thus, the different binding capacity between these compartments might be attributed to a different concentration in circulating transcortin. This concentration gradient may play a role in the transplacental transfer of corticosteroids during pregnancy and would favor passage toward the maternal compartment. The low transcortin level in the neonatal period might be attributed to a low biosynthesis capacity of the liver in the neonate, although nothing is known about the clearance rate of transcortin in the neonatal period. The transcortin level increases toward adult values during the first year of life and this would suggest either a maturation of the liver biosynthesis enzymatic systems or a biosynthesis stimulation. A possible contribution of a pituitary factor might be suggested on the basis of experimental work in the rat, but the meaning of this increase in cortisol binding capacity in plasma remains to be elucidated.

In vitro Studies on the Metabolism of Cortisol-4-[<sup>14</sup>C] in the Lenses of Rats during Maturation

The effects of maturation on the inactivation and degradation of cortisol-4-[14C] in the lenses of rats were investigated. Inbred Wistar strain rats were used at the 4th and 10th postnatal day for the immature stage and the 60th postnatal day for the mature stage of male rats. Differences were found in the cortisol-binding capacity in the rat lens between the immature and mature stage. No change with age was found for sulphate conjugation, whereas glucuronide conjugation was found to increase with age. The most interesting fact was that degradation of cortisol-4-[14C] by lens homogenate is carried out actively in the mature stage, whereas only the side-chain loss metabolite of cortisol-4-[14C] is detectable in immature stage. These findings suggest that there is an age difference in the metabolic pattern of cortisol in the rat lens and that this might be closely related to the induction of enzymes involved in the metabolism of cortisol during maturation.

In vitro Effects of Sulphydryl Agents on the Cortisol-Binding Capacity of the Rat Lens

The authors have found that cysteine markedly increased the cortisol-binding capacity of the lens in vitro and that the oxidized form of glutathione acted as an inhibitor on the cor

The oestrogenic potency of various contraceptive steroids as determined by their effects on transcortin-binding capacity.

ABSTRACT Various contraceptive steroids were analyzed for their oestrogenic potency in the human using changes in the cortisol-binding capacity of transcortin (TC-BC) as a parameter. A linear dose-related elevation of TC-BC was recorded in response to increasing amounts of ethinyloestradiol (EE). Mestranol (EEME), given in the same galenical preparation as EE, proved to be only 64 % as potent. Megestrol acetate, dl-norgestrel, norethindrone and its acetate failed to influence TC-BC, while norethynodrel induced a significant rise. This effect is apparently due to both the inherent oestrogenic activity of norethynodrel and its contamination with EEME. The administration of combined oral contraceptives resulted in a significant elevation of TC-BC. The drugs studied were (in order to increasing total oestrogenicity) 50 μg EEME + 1 mg norethindrone, 35 μg EE + 0.25 mg d-norgestrel, 100 μg EEME + 2 mg norethindrone, 50 μg EE + 0.25 mg d-norgestrel, and 100 μg EEME + 2.5 mg norethynodrel. The observed rise in transcortin activity appears to depend primarily on the chemical structure and dosage of the oestrogen component. These findings clearly indicate that the administration of conventional oral contraceptives, even those containing low oestrogen doses, does not cause an oestrogen deficiency, but rather an oestrogen oversupply of the female organism.

Effect of Riboflavin on the Cortisol-Binding Capacity of the Rat Lens with Particular Reference to the Pathogenesis of Steroid Cataracts

Decrease of the cortisol-binding capacity (CBC) in the lenses of rats treated with dexamethasone phosphate was not prevented by the administration of either large or small doses of equimolal amount of riboflavin or flavin mononucleotide (FMN). However, a decrease of CBC was prevented, to a considerable degree, by flavin adenine dinucleotide (FAD) administration. A decrease of the CBC in the hypophysectomized rat lens was reduced by adreno-corticotropic hormone (ACTH) or FAD and this tendency was promoted by simultaneous administration of ACTH with FAD. The CBC in the lenses of unilaterally adrenalectomized rats decreased gradually with time and the decrease was smaller than in bilaterally adrenalectomized rats. The drop of CBC in the lenses of rats treated with CCl4 was partially prevented by FAD administration. These results suggest that the CBC in the lenses of rats is influenced by the pituitary-adrenal-liver axis and that the riboflavin diphosphate may play an important role in the synthesis of cortisol-binding protein in the lenses of rats after hypophysectomy, adrenalectomy or liver impairment induced by CC14 administration. The pathogenesis or clinical features in the initial stage of steroid cataracts are discussed and consideration is given to the possible role of phosphorylation insufficiency in the lens and the depression of ATP and phosphocreatine synthesis in the posterior subcapsular region.

Identification and Partial Purification of “Transcortin”-like Protein within Human Lymphocytes

Abstract We have detected and localized in situ lymphocyte proteins with antigenic determinants similar to those of the cortisol-binding globulin of human plasma. All lymphocyte fractions, obtained by differential centrifugation and ammonium sulfate fractionations exhibited cortisol-binding capacities and were recognized by transcortin-specific antibody. Inasmuch as the 40 to 65% ammonium sulfate fraction of the 105,000 x g lymphocyte cytosol accounted for over 90% of the total binding capacity of the cell, this fraction was further purified by the procedure employed for the purification of plasma transcortin. These procedures yielded a protein with about 20% the cortisol-binding capacity of transcortin, migrated more rapidly than transcortin in disc acrylamide electrophoretic systems, and formed a line of identity with transcortin when treated with transcortin-specific antibody. Fluorescein-labeled transcortin antibody applied to sectioned lymphocytes showed almost exclusive cytoplasmic localization of transcortin-like protein. These results show for the first time the existence of cortisol-binding proteins within the human lymphocyte antigenically identical with transcortin.

Biochemical studies on the pathogenesis of steroid cataracts, with particular reference to the pituitary-adrenal-liver axis

Factors having an effect on the binding and inactivation of [4- 14C] cortisol in the rat lens, such as hypophysectomy, adrenalectomy, dexamethasone phosphate administration and liver impairment were investigated. Furthermore, the effects of flavin adenine dinucleotide on the cortisol-binding capacity in the lenses of adrenalectomized rats and of dexamethasone phosphate-treated rats were also investigated. The authors found that cortisol-binding by lens protein and sulphate conjugation of cortisol in the lens are greatly influenced by the pituitary-adrenal-liver axis. A decrease in the cortisol-binding capacity in the lens of adrenalectomized rats was slightly prevented by flavin adenine dinucleotide administration. A similar result was found in the lenses of rats treated with dexamethasone phosphate, suggesting that riboflavin may influence the inactivation of steroid hormone in the lens.

Plasma cortisol-binding capacity in mental illness.

SynopsisPlasma cortisol-binding capacity (CBC) was determined in 18 psychiatric patients, including 14 who had affective disorders, and 18 controls. Men with unipolar depressive illnesses had significantly lower CBC values than men with bipolar illnesses and male controls. No other significant differences in CBC values were found and it was concluded that elevated total plasma cortisol levels in affective disorder were probably associated with increased levels of unbound cortisol. The possibility that low CBC values may be a genetic marker for certain types of affective disorder requires further investigation. The effect of certain drugs on CBC was also investigated.

Plasma cortisol, corticosterone and non-protein-bound cortisol in extra-corporeal circulation.

ABSTRACT The response of plasma cortisol, corticosterone and non-protein-bound cortisol in the extracorporeal circulation was investigated in 14 patients. The pre-perfusion levels of plasma cortisol, corticosterone and non-protein-bound cortisol were significantly elevated. During and immediately after perfusion, the levels of cortisol and corticosterone were found to decrease significantly from the pre-perfusion levels, while the percentage of non-protein-bound cortisol was shown to increase significantly. This indicates a marked decrease in cortisol binding capacity of plasma during extracorporeal circulation. Moreover in 200 plasma samples, it was demonstrated that the cortisol level increased markedly and the cortisol binding capacity decreased slightly during and shortly after major surgery without perfusion. It is concluded that stressful situations in major surgery with or without perfusion are associated with markedly increased levels of biologically active non-protein-bound cortisol. The elevated level of non-protein-bound cortisol in surgery seems to be dependent on the increase in the level of plasma cortisol as well as on the decrease in the cortisol binding capacity of plasma. Although the increased plasma cortisol plays the most important role in surgery with no perfusion, the decreased cortisol binding capacity may be the more effective factor involved during perfusion.

A new method for estimation of corticosteroid-binding globulin with hydrophobic resin

A new method is described for the estimation of corticosteroid-binding globulin by means of the hydrophobia resin adsorption method. Corticosteroid-binding globulin in the serum is saturated by incubation with cortisol, unbound and albumin-bound cortisol is removed by shaking with the resin, and cortisol remaining in the supernatant fluid is measured by fluorometry, its represents the cortisol-binding capacity of corticosteroid-binding globulin. Normal range of cortisol-binding capacity was 0.154–0.222 μg/ml at 15°.