[Neonatal screening for congenital hypothyroidism: more than 30 years of experience in the Netherlands]. / Neonatale screening op congenitale hypothyreoïdie: ruim 30 jaar ervaring in Nederland.

OBJECTIVE: To describe the Dutch neonatal screening programme for congenital hypothyroidism (CH). DESIGN: Descriptive study. METHOD: Data on neonatal screening for CH in the period 1 January 1981 through 31 December 2011 were obtained from the Department for Vaccine Supply and Prevention Programmes of the Dutch National Institute for Public Health and the Environment (RIVM), laboratories and paediatricians to whom babies with abnormal screening results were referred. The screening procedure has been amended several times. In the period 1981-1994, only T4 and TSH were measured in heel prick blood, for example. From 1995, thyroxine-binding globulin (TBG) was added to the screening protocol. RESULTS: The participation rate was 99.7%. Before 1995 the sensitivity, specificity and positive predictive value were 94%, 99.51% and 6%, respectively. From 1995 these percentages were 98%, 99.85% and 21%, respectively. The total prevalence of CH was 1:2670 (prevalence of CH of thyroidal origin was 1:3100 and CH of central origin was 1:21,600). The percentages of patients with severe CH treated before day 15 in the periods 1981-1990, 1991-2000 and 2001-2011 were 24% (63/263), 63% (170/269) and 96% (176/184), respectively. CONCLUSION: The sensitivity and specificity of the screening procedure has considerably increased since 1995 compared with the period before 1995. In recent years patients with severe CH were treated considerably earlier than in the first years of the screening. Neonatal screening for CH may be considered as an important success for public health care.

[Immune, inflammatory, and nutritional protein profile in children with iron deficiency in Côte d'Ivoire]. / Déficit en fer, profil protéique immunitaire, inflammatoire et nutritionnel chez l'enfant de Côte-d'Ivoire.

Throughout the world and particularly in sub-Saharan Africa, deficiencies in trace elements constitute a real public health problem because of the insufficient nutritional quality of food. These trace elements are necessary for many of the body's biochemical reactions. The role of microelements such as vitamin A and zinc has been established in the functioning of the immune system and secretion of inflammatory reaction proteins, but the role of iron in these functions remains to be elucidated. The sample consists of 186 children (3/4) 80 with an iron deficiency and 106 with normal iron status. They range in age from 5 to 15 years and all attend school in the department of Adzope. The study excluded all children with parasites that might affect blood iron, protein and other hematological indicators, in particular, Plasmodium falciparum, Giardia intestinalis, Trichomonas intestinalis, Ascaris lumbricoides, and Ancylostoma. Inflammatory, immune and nutritional proteins were measured by radial immunodiffusion (Mancini's method). Ferritin was measured by a specific immunoenzymatic assay. Hematological indicators were tested by an automatic blood cell counter. Nutritional status was estimated by the weight/height ratio (W/H). This analysis showed that iron deficiency was associated with reduced IgG levels (p < 0.05), although immunoglobulins A and M remained stable (p > 0.05. Iron deficiency was also associated with reduced levels of thyroxine-binding prealbumin (TBPA) and albumin (p < 0.05). Inflammatory proteins did not differ significantly between the two groups (p > 0.05). Furthermore, the prognostic inflammatory and nutritional index (PINI) did not show any inflammatory, vital or nutritional risk, because it was lower than or equal to 2. Finally, malnutrition was not observed in the iron-deficient children: the difference in the weight/height ratio (W/H = 96.58 +/- 2.4%) between the children with iron deficiency and those with normal iron status (98.7 +/- 4.3%) did not differ significantly. The reduced IgG associated with iron deficiency may be attributed to the role that iron plays in the proliferation and maturation of lymphocytes. Reduced iron levels would thus lead to slowing down the hematopoietic mechanism, resulting in a decrease in B lymphocyte production and thus inevitably a reduction in IgG synthesis. The reduction in albumin and TBPA associated with the iron deficiency but in the absence of any sign of malnutrition (W/H > 96%) or inflammatory risk (PINI < 2) in either study group shows that iron may play a dominant role during protein synthesis. Iron deficiency might limit the energy of cellular tissues, leading to a reduction in RNA activity (transcription and translation), which would in turn decrease ribosome activity in tissues and thus reduce amino acid synthesis in cells, resulting in the reduction observed in protein synthesis. The lack of difference between the study groups in inflammatory proteins, notably CRP and alpha1-GPA, indicates that iron deficiency does not appear to be related to an inflammatory process. This study of children without any apparent clinical signs of iron deficiency shows that such a deficiency may be associated with a disruption in protein production. The proteins concerned include IgG, TBPA and albumin. The public authorities should pay particular attention to improving children's diets, especially their micronutrient levels, including for iron, vitamin A and zinc.

Protocol for the purification of proteins from biological extracts for identification by mass spectrometry.

When a protein signal is selected by mass spectrometry as being a potential biomarker, it is necessary to formally identify it. This process involves separation of the protein in question and its identification by either peptide fingerprinting or tandem mass spectrometry sequencing. In the following pages, a simple and rapid protocol is described. Basically, the protocol consists of an initial rational selection of a few sorbents followed by alignment of these as a series of columns to obtain the purified target protein. This preparation is then submitted to electrophoresis, the band is excised and the trypsin digest is analyzed by either mass spectrometry (mass fingerprinting approach) or by LC-MS/MS (sequencing). The development of the process takes only a few days. Experimental data for the isolation and identification of proteins are discussed and two examples are shown.

Effects of evening vs morning thyroxine ingestion on serum thyroid hormone profiles in hypothyroid patients.

OBJECTIVE: Standard drug information resources recommend that l-thyroxine be taken half an hour before breakfast on an empty stomach, to prevent interference of its intestinal uptake by food or medication. We observed cases in which TSH levels improved markedly after changing the administration time of l-thyroxine to the late evening. We therefore conducted a pilot-study to investigate whether l-thyroxine administration at bedtime improves TSH and thyroid hormones, and whether the circadian rhythm of TSH remains intact. DESIGN Patients were studied on two occasions: on a stable regimen of morning thyroxine administration and two months after switching to night-time thyroxine using the same dose. On each occasion patients were admitted for 24 h and serial blood samples were obtained. PATIENTS: We investigated 12 women treated with l-thyroxine because of primary hypothyroidism, who used no medication known to interfere with l-thyroxine uptake. MEASUREMENTS: Patients were admitted to hospital and blood samples were obtained at hourly intervals for 24 h via an indwelling catheter. Following this first hospital admission, all women were asked to switch the administration time from morning to bedtime or vice versa. After 2 months they were readmitted for a 24-h period of hourly blood sampling. Blood samples were analysed for serum TSH (immunometric assay), FT4 and T3 (competitive immunoassay), T4 and rT3 (radioimmunoassay), serum TBG (immunometric assay) and total protein and albumin (colourimetric methods). RESULTS: A significant difference in TSH and thyroid hormones was found after switching to bedtime administration of l-thyroxine. Twenty-four-hour average serum values amounted to (mean +/- SD, morning vs bedtime ingestion): TSH, 5.1 +/- 0.9 vs 1.2 +/- 0.3 mU/l (P < 0.01); FT4, 16.7 +/- 1.0 vs 19.3 +/- 0.7 pmol/l (P < 0.01); T3, 1.5 +/- 0.05 vs 1.6 +/- 0.1 nmol/l (P < 0.01). There was no significant change in T4, rT3, albumin and TBG serum levels, nor in the T3/rT3 ratio. The relative amplitude and time of the nocturnal TSH surge remained intact. CONCLUSIONS: l-thyroxine taken at bedtime by patients with primary hypothyroidism is associated with higher thyroid hormone concentrations and lower TSH concentrations compared to the same l-thyroxine dose taken in the morning. At the same time, the circadian TSH rhythm stays intact. Our findings are best explained by a better gastrointestinal uptake of l-thyroxine during the night.

Effects of transdermal and oral contraceptives on estrogen-sensitive hepatic proteins.

OBJECTIVE: The purpose of this study was to evaluate the effects of the contraceptive patch and an oral contraceptive (OC) on serum concentrations of estrogen-sensitive hepatic proteins, ethinyl estradiol (EE) and levonorgestrel (LNG). METHODS: Twenty-four women were randomized to receive three cycles of a contraceptive patch that delivers EE 20 microg/day and norelgestromin 150 microg/day or an OC that contains EE 35 microg and norgestimate 250 microg. Blood samples were taken at baseline and at the end of Cycle 3. Serum levels of sex-hormone-binding globulin (SHBG), thyroxine-binding globulin (TBG), corticosteroid-binding globulin (CBG) and C-reactive protein (CRP) were quantified by immunoassay methods. EE and LNG levels in patch users were measured by radioimmunoassay (RIA) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay. The paired t test and Student's t test were used for statistical analysis. RESULTS: Nineteen women completed the study (patch, n=10; OC, n=9). Treatment with both the patch and OC resulted in significant increases from baseline in SHBG, TBG and CBG. The increase in CRP was significant in the patch group and approached significance in the OC group. The increases in SHBG and TBG observed with the patch were significantly greater than those associated with the OC. By way of RIA and LC-MS/MS assay methods, the patch was associated with mean EE levels of 114 and 111 pg/mL, respectively. CONCLUSIONS: The serum concentrations of estrogen-sensitive hepatic proteins and EE associated with the patch suggest that this new contraceptive system may have relatively large net estrogen effects.

Congenital hypothyroidism and the second newborn metabolic screening in Colorado, USA.

OBJECTIVE: To evaluate the effectiveness of a second newborn screening for congenital hypothyroidism (CH). METHODS: All infants born in Colorado, USA, from July 1996 through June 2004 had a total thyroxine measured with secondary thyroid stimulating hormone determination. RESULTS: The number of first and second newborn screens completed was 494,324 and 471,877, respectively. The first screen identified 185 cases of CH (incidence of 1:2,703). The second screen identified an additional 42 cases. Overall, the incidence based on both the first and second screenings was 1:2,174. The false negative rate for the first screen was 15.6%. In the absence of a second screen, one infant with CH out of every 11,111 babies screened would have been missed. The addition of the second screen increased the cost-per-case identified from dollars 6,108 to dollars 9,730. CONCLUSIONS: With only one newborn screen for CH, the number of missed cases is significant and higher than previously reported.

The relationship between iron status and thyroid hormone concentration in iron-deficient adolescent Iranian girls.

Extensive data from animal and human studies indicate that iron deficiency impairs thyroid metabolism. The aim of this study was to determine thyroid hormone status in iron-deficient adolescent girls. By stepwise random sampling from among all public high schools for girls in Lar and its vicinity in southern Iran, 103 out of 431 iron deficient subjects were selected. Urine and serum samples were collected and assayed for urinary iodine and serum ferritin, iron, total iron binding capacity (TIBC), thyroid stimulating hormone (TSH), thyroxine (T4), triiodothyronine (T3), free thyroid hormones (fT4 and fT3), triiodothyronine resin uptake (T3RU), reverse triiodothyronine (rT3), selenium and albumin concentrations. Hematological indices for iron status confirmed that all subjects were iron-deficient. There was a significant correlation between T4 and ferritin (r = 0.52, P < 0.001) and between TSH and ferritin (r = -0.3, P < 0.05). Subjects with low serum ferritin had a higher ratio of T3/T4 (r = -0.42, P < 0.01). Using stepwise regression analysis, only ferritin contributed significantly to the rT3 concentration (r = -0.35, P < 0.01). The results indicate that the degree of iron deficiency may affect thyroid hormone status in iron-deficient adolescent girls.

Impact of varying levels of protein intake on protein status indicators after gastric bypass in patients with multiple complications requiring nutritional support.

BACKGROUND: <6% of patients who undergo Roux-en-Y gastric bypass (RYGBP) for morbid obesity require nutritional support after surgery. Protein and caloric needs have been estimated as 14-21 kcal, 1.2 g protein/kg/current body weight/day in uncomplicated morbidly obese patients. This study assesses the effect of varying protein-calorie intake in complicated patients after RYGBP on two markers of protein status: thyroxine-binding prealbumin (TBPA) and serum albumin. METHODS: This 25-month retrospective study consisted of 22 patients with postoperative complications. Serum albumin, TBPA, medical nutrition care-plans, laboratory data and history were reviewed. These post-RYGBP patients who had BMI>35 and no multi-system organ failure or fistulas, had complications after surgery requiring nutrition support services (NSS). Serum albumin and TBPA were matched to fed levels of protein using random coefficient regression analysis. RESULTS: Mean incremental increases of 2.34 mg/dl (TBPA, P=0.0113) and 0.11 g/dl (serum albumin, P=0.0272) were found with each 0.5 g protein intake increase/kg ideal body weight/day (kg/IBW/day). Patients required NSS for 23+/-21 (mean+/-SD) days, with 15+/-19 days fed at goal rate. Initial serum albumin was 2.3+/-0.5, with a final measure of 2.7+/-0.5 g/dl. Goal protein and calorie intake were 2.1 g and 17 kcal/kg IBW/day versus actual intake of 1.6 g and 13 kcal/kg IBW/day. CONCLUSION: Morbidly obese patients requiring NSS following RYGBP risk iatrogenic protein malnutrition. There was a positive linear relationship between protein status and protein intake that warrants further study of higher protein feeding in complicated post-RYGBP patients.

Clinical effectiveness and cost-effectiveness of the use of the thyroxine/thyroxine-binding globulin ratio to detect congenital hypothyroidism of thyroidal and central origin in a neonatal screening program.

CONTEXT: Since the introduction of screening for congenital hypothyroidism (CH) in 1974, the optimal laboratory strategy has been the subject of debate. OBJECTIVE: To assess the clinical effectiveness and cost-effectiveness of various types of thyroxine (T(4))-based strategies to screen for CH. DESIGN, SETTING, AND PARTICIPANTS: In the Netherlands, since January 1, 1995, a primary T(4) determination with supplemental thyroid-stimulating hormone (TSH) and T(4)-binding globulin (TBG) measurements has been used. Results were calculated from cumulative findings for 1181079 children screened between January 1, 1995, and December 31, 2000. MAIN OUTCOME MEASURES: Rates of detection of patients with CH of thyroidal origin (CH-T) or CH of central origin (CH-C), false-positive rates, laboratory costs, and costs of initial diagnostic evaluations. RESULTS: All known infants (n = 393) with CH-T and 92% (n = 66) of infants with CH-C were detected on the basis of low T(4) levels, TSH elevation, and/or low T(4)/TBG ratios. If the decision to refer had been based solely on TSH elevation, then 94% of patients with CH-T and none of the patients with CH-C would have been detected. If low T(4) levels (

Changes in thyroid hormone state in children receiving chemotherapy.

OBJECTIVE: The concentrations of thyroid function determinants may change during severe illness. Our goal was to quantify their changes in children with cancer during chemotherapy, and to correlate them to clinical condition and type of drugs. DESIGN: During a 3-month period all patients admitted for chemotherapy to the paediatric oncology ward were evaluated for inclusion. Patients with brain tumours, neuroblastoma (cranio)spinal irradiation and use of dexamethasone before the first blood sample were excluded. MEASUREMENTS: Plasma concentrations of T4, T3, rT3, thyroxine-binding globulin (TBG), thyroglobulin (Tg), TSH, IGF-1, cortisol, PRL and physical well-being by means of questionnaires were measured before and during chemotherapy. RESULTS: In 19 children, 46 courses of chemotherapy and 123 plasma samples were analysed. During chemotherapy, mean concentrations of TSH, T3, Tg and cortisol decreased to 53, 67, 69 and 15% of the baseline value, respectively. Mean plasma rT3 increased to 217% of baseline. In 87% of all courses, one or more thyroid parameter(s) was aberrant. Furthermore, in 23 samples (19%) from 10 patients (53%), the concentration of IGF-1 was below the reference value (adjusted for sex and age). Small changes were seen in scores for clinical condition but none was related to a change in thyroid function determinant. Most changes in thyroid hormones could be attributed to using dexamethasone. CONCLUSIONS: These results demonstrate that, in children, thyroid hormone state changes significantly during chemotherapy, apparently not related to physical well-being but to the drugs administered. Future investigations should focus on the impact for patient care and possibilities of (preventive) intervention.

Developmental trends in cord and postpartum serum thyroid hormones in preterm infants.

The purpose of this study was first to clarify postnatal trends in sera T(4), free T(4) (FT(4)), T(4)-binding globulin, TSH, T(3), rT(3), and T(4) sulfate levels in cord and at 7, 14, and 28 d in groups of preterm infants at 23-27 wk (n = 101), 28-30 wk (n = 196), and 31-34 (n = 253) wk gestation, and second to compare these trends to those of term infants and also with cord sera levels of equivalent gestational ages (n = 812; 23-42 wk gestation). In all preterm groups, TSH and rT(3) decrease to below, T(4)-binding globulin increases to within, and T(3) and T(4) sulfate increase to above cord levels of equivalent gestational age. Term infants are hyperthyroxinemic relative to cord and nonpregnant adult levels of T(4). Postnatal T(4) increases are attenuated in 31- to 34-wk infants, absent in 28- to 30-wk infants (although levels are equivalent to gestational age), and crucially reversed in 23- to 27-wk infants. This immature group is hypothyroxinemic relative to other groups and to cord levels of equivalent gestational age. Compared with term infants, postnatal FT(4) increases are lower in 31- to 34-wk infants, attenuated in 28- to 30-wk infants, and absent in 23- to 27-wk infants. The 23- to 27-wk group is distinctive; they are hypothyroxinemic on T(4) levels, yet FT(4) levels are within the cord levels of equivalent gestational age.

Iron deficiency anaemia in childhood and thyroid function.

Studies in animals and adults have indicated iron deficiency anaemia to be associated with altered thyroid hormone metabolism. The aim of the present study was to determine the effect of iron deficiency anaemia on the thyroid function of young children. Concentrations of thyroxine (T4) and triiodothyronine (T3), free thyroid hormones (fT4 and fT3), thyroxine binding globulin (TBG), and thyroid stimulating hormone (TSH) were measured in the basal state and in response to an intravenous bolus of thyrotropin releasing hormone (TRH) in nine children one to three years of age with iron deficiency anaemia (IDA) before and after treatment with oral iron. The results of the anaemic children were also compared to basal and stimulated concentrations of thyroid hormones, TBG, and TSH of eight iron sufficient, age-matched children. Seven of the IDA and 6 of the control children were male. The mean haemoglobin (Hb) and serum ferritin (SF) in the IDA children at baseline were 93g/L (range 81-102) and 6g/L (range 1-12) which increased to 121g/L (range 114-129) and 54g/L (range 19-175), respectively, after a mean of 2.3 months (SD 0.5) of iron therapy. In the control group, mean Hb and SF were 125g/L (range 114-130) and 51 g/L (range 24-144), respectively. The basal values of TBG and thyroid hormones of the IDA children before and after iron treatment were not different from the control children. Similarly, there was no statistical difference in the thyroid hormones in the IDA children before compared to after resolution of the anaemia. Compared to the control children, the TSH response over time to TRH, TSH area under the curve (TSHAUC), and the peak TSH value after stimulation were all lower in the IDA children both before and after resolution of anaemia, but the differences were not significant. Iron therapy and resolution of anaemia had no effect among the IDA children. The time to reach the peak TSH concentration was longer in the IDA children (P=0.08) than the control children before iron therapy. While the time to peak TSH decreased upon resolution of the anaemia, the difference was not significant. There was no effect of Hb concentration, age, or anthropometry with TSH, TSHAUC, or time to peak TSH after TRH stimulation in the IDA children before treatment. Normal thyroid function was preserved in these children with iron deficiency anaemia, however three of nine children had minor abnormalities of hypothalamic-pituitary function. These results indicate that hypothyroidism is unlikely to be a major cause of impaired psychomotor development or growth in young children with iron deficiency anaemia.

Thyroxine-binding globulin cleavage in cord blood.

Thyroxine-binding globulin, a member of the serine protease inhibitor superfamily of proteins (serpins), releases T(4) on cleavage by polymorphonuclear elastase. Such cleavage, previously shown to occur during sepsis and with an exogenous inflammatory stimulus, is now demonstrated in the cord blood of normal babies and appears to be part of a physiological inflammatory response in the newborn. In association with the neonatal TSH surge, thyroxine-binding globulin cleavage is likely to contribute to an increased flux of T(4) to neonatal tissues at a time when T(4)-sensitive morphogenic and biochemical changes are occurring.

Thyroid hormone metabolism in pediatric cardiac patients treated by continuous povidone-iodine irrigation for deep sternal wound infection.

OBJECTIVE: The purpose of this study was to assess the influence of povidone-iodine mediastinal irrigation used for the treatment of deep sternal wound infection (DSWI) on thyroid function. METHODS: Thyroid function was studied in 18 pediatric cardiac patients treated with continuous povidone-iodine irrigation for DSWI. The median age of patients was 8 months (18 days-5.3 years). Serum concentrations of total triiodothyronine (TT3), total thyroxine (TT4), free triiodothyronine (FT3), free thyroxine (FT4), thyroid-stimulating hormone (TSH), reverse triiodothyronine (rT3) and thyroxine-binding globulin (TBG) were measured at three time points: (a) prior to mediastinal reexploration (before povidone-iodine exposure); (b) immediately after discontinuation of povidone-iodine irrigation; (c) 2 weeks after discontinuation of mediastinal irrigation. Urinary iodine excretion was examined on the last day of povidone-iodine exposure. RESULTS: Prior to the mediastinal reexploration, the median TT3 and TT4 levels were below the normal range, then increased significantly to concentrations within the normal range. The median serum FT3 levels were within the normal range throughout the observation period, though a significant increase of FT3 levels was observed after discontinuation of irrigation. The median serum FT4 concentrations were within the normal range prior to irrigation and did not change significantly. The median rT3 levels were within the normal range, close to upper normal limit. The median TBG levels were within the normal range throughout the observation period, though a significant increase of TBG levels was observed during the period of mediastinal irrigation. The median TSH level was within the normal range prior to mediastinal irrigation and did not change significantly. Urinary iodine concentrations in infants with povidone-iodine irrigation were significantly higher 6700 microg/l (range, 1600-15000 microg/l) than in the group of 53 healthy infants 200 microg/l (range, 20-780 microg/l, P<0,001). CONCLUSIONS: Our data showed that the use of povidone-iodine irrigation in the patients with DSWI has not lead to any significant alteration in thyroid function within the study period.

The quantity of thyroid hormone in human milk is too low to influence plasma thyroid hormone levels in the very preterm infant.

BACKGROUND: Thyroid hormone is crucial for brain development during foetal and neonatal life. In very preterm infants, transient low levels of plasma T4 and T3 are commonly found, a phenomenon referred to as transient hypothyroxinaemia of prematurity. We investigated whether breast milk is a substantial resource of thyroid hormone for very preterm neonates and can alleviate transient hypothyroxinaemia. Both the influence of breast feeding on plasma thyroid hormone levels and the thyroid hormone concentration in preterm human milk were studied. METHODS: Two groups were formed from the placebo group of a randomized thyroxine supplementation trial in infants born at < 30 weeks' gestational age on the basis of the mean breast milk intake during the third, fourth and fifth weeks of life. One group received more than 50% breast milk (mean breast milk intake 84%, n = 32) and the other group less than 25% breast milk (mean breast milk intake 3.3%, n = 25). Plasma thyroid hormone concentrations were compared between the two groups. Breast milk was collected from mothers of infants participating in the same trial and the thyroxine concentration in breast milk was measured with RIA after extraction. RESULTS: No significant differences were found between both groups in plasma concentrations of T4, free T4, T3, TSH, rT3 and thyroxine-binding globulin (TBG), which were measured once a week. Thyroxine concentration in breast milk ranged between 0.17 microg/l and 1.83 microg/l (mean 0.83, SD 0.3 microg/l) resulting in a maximum T4 supply of 0.3 microg/kg via ingested breast milk. In formula milk, the T4 concentration was equally low. Protease treatment did not influence the measured T4 concentrations. CONCLUSIONS: No differences in plasma thyroid hormone between breast milk-fed and formula-fed infants were found. The amount of T4 present in human milk and formula milk is too low to alter the hypothyroxinaemic state of preterm infants.

Characterization and primary structures of bovine and porcine thyroxine-binding globulin.

Thyroxine-binding globulin (TBG) is the major serum transport protein for iodothyronines in most of the large, omni- or herbivorous mammals. Characterization of human TBG (hTBG), including its 20 known natural variants, allowed the identification of the ligand-binding site and a correlation of diminished synthesis or loss of function with mutations in the TBG gene. Further refinement of the structure-function correlation, especially the high binding affinity and heat stability, requires characterization of other mammalian TBGs, of which only rat and sheep TBG were available. We now present some of the chemical and physical properties of bovine TBG (bTBG) and porcine TBG (pTBG) and their primary structures deduced from their cDNA sequences. The serum concentrations of bTBG and pTBG estimated by Scatchard analysis of T(4)-binding were similar to hTBG. The T(4)-binding affinity of human, bovine and porcine TBGs were all similar, at 1.2x10(10) M(-1). However, heat stability of the animal TBGs was reduced, with a half life of denaturation of 7 min (bTBG) and 5 min (pTBG) at 55 degreeC, compared with 21 min for hTBG. Nucleotide alignment revealed identity with hTBG of 85.5% (bTBG) and 83.7% (pTBG) and amino acid identity of 82.8% (bTBG) and 82.6% (pTBG). As expected, the relevant parts of the ligand-binding domain (amino acids 215-291, and 363-395) were highly conserved at more than 95% similarity. Comparison of the five known mammalian TBGs allows focusing of future mutagenesis experiments to further characterize the properties of the molecule.