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Screening for congenital hypothyroidism for all newborns of the former western parts of the city of Berlin was instituted in 1978 by determination of the TSH levels in dried filter paper blood spots of the 3rd to 5th day of life. Since 1991 the newborns of the former eastern parts have been also included in the same screening program. From 1978 to 1995 a total number of 104 newborns with permanent congenital hypothyroidism have been detected resulting in a prevalence of congenital hypothyroidism of 1:3800. The etiological diagnosis of hypothyroidism was made by imaging studies (ultrasonography or 99mTc scintigraphy) and assessment of serum thyroglobulin and thyroid hormone levels. Using this approach in 37 children (30 female, 7 male) the diagnosis of athyrosis, in 20 children (15 female, 5 male) the diagnosis of ectopy and in 21 children (18 female, 3 male) the diagnosis of thyroid hypoplasia was made, 16 children (8 female, 8 male) had a normally sized gland and 4 (1 female, 3 male) had congenital goiter. In 86% of all patients the age at onset of thyroxine (L-T4) replacement therapy was 8 or 9 days of life (3-42 days) and the median initial L-T4 dose was 14 micrograms/kg/day (10-16 micrograms/kg/day). The intellectual outcome of 77 children (2-16 years) was studied and normal scores for the intelligence (IQ) and developmental (DQ) quotients were found in 71 (92%). Outcome was not correlated to the age at onset of treatment, the initial dose and the severity of hypothyroidism, but there was a positive correlation of the socioeconomic status of the family and the IQ of the patients. The results of the screening program in Berlin document that an early and efficient thyroxine replacement can normalize the intellectual outcome of patients with congenital hypothyroidism independent of the severity of the disease as assessed by the residual thyroid function detectable at diagnosis.

Screening newborns for inherited disorders provides an opportunity for pre-symptomatic identification and early intervention to prevent or mitigate morbidity and mortality associated with these conditions. Since the introduction of newborn screening in 1962 to screen for phenylketonuria, technological advances have enabled the screening panel to expand substantially so that it now includes more than 50 disorders. Newborn screening will continue to evolve,, and deployment of improved methodologies and incorporation of additional disorders are expected. This article provides an overview of the current state of newborn screening, and describes the disorders detectable, the methodologies employed, and the challenges involved in analyses of specimens obtained from newborns.

Hypothyroidism and autoimmune thyroiditis are more prevalent than previously considered in women during pregnancy and the postpartum, and are associated with adverse effects on the mother and her fetus. We determined the efficacy and accuracy of screening women for primary hypothyroidism and autoimmune thyroiditis by testing TSH and two thyroid antibodies (TAb): thyroperoxidase antibodies (TPOAb) and thyroglobulin antibodies (TgAb), in eluates of filter paper specimens collected during early pregnancy and the postpartum.We enrolled 494 first-trimester pregnant women with no exclusion criteria into a prospective study to detect primary hypothyroidism and autoimmune thyroiditis. Finger stick blood was applied to filter paper, dried in room air, eluted, and promptly tested for TSH and TAb. A total of 178 of the pregnant women (36%) were tested in the early postpartum. Women with abnormal results had confirmatory serum tests.It was found that 91 pregnant women (18.4%) and 43 postpartum women (24.2%) had abnormal TSH values (4.0 mU/L) and/or positive TAb; 140 pregnant women (28.3%) had TSH values2.5 mU/L. All subjects with TSH values4.0 mU/L tested positive for TAb. Eighteen women (3.6%) who tested normal during pregnancy tested abnormal in the postpartum.This study confirms that TSH and TPOAb measured in eluates of blood-spotted filter paper specimens are excellent screening tests to detect primary hypothyroidism and autoimmune thyroiditis in pregnant and postpartum women. Results are very comparable to serum data in this population published in the literature.

Background Radioiodine is the treatment of choice for relapsed hyperthyroidism although the optimum protocol is uncertain. Fixed dose radioiodine is increasingly popular but responses may vary. Aim To assess the outcome of 131I therapy in hyperthyroidism using a standard dose regimen in a regional referral centre and to explore factors influencing outcome. Methods We studied 449 patients (M:F 82:367; age range 13-89y, median 42y) with hyperthyroidism treated between 2003 and 2007 with a standard dose of 550MBq 131I. Patients were classified as either Graves’ disease, toxic multinodular goitre or indeterminate aetiology. Antithyroid drugs were routinely stopped at least 1 week before radioiodine. Results One year after radioiodine 334 (74%) were hypothyroid, 85 (19%) were euthyroid and 30 (7%) had required a further dose of 131I. Patients with Graves’ disease were more likely to become hypothyroid than those with toxic multinodular goitre (78% v 37%, p80pmol/L (normal range 9.0 – 19.0 pmol/L) at presentation was associated with an increased failure rate (17% compared with 5% and 3% for 40-79pmol/L and

Introduction: The preparation of bloodspot calibrators and quality control samples for neonatal thyrotropin (TSH), as part of the national quality assessment scheme in The Netherlands, is sometimes problematic. Often, the recovery of the added TSH is well above 100% when measured in plasma. Results from other External Quality Assessment Schemes for neonatal TSH show similar problems. Methods and Results: A questionnaire was sent to 17 kit manufacturers and organizers of EQAS concerning the preparation of bloodspot calibrators for TSH; 13 completed questionnaires were returned. There are large differences with respect to type of filterpaper, matrix, assessment of TSH-concentrations to the calibrators, size of the bloodspots, shelf life, etc. Discussion: Attention is focused on several items to be considered when attempting standardization of the preparation of TSH bloodspot calibrators.

Thyroid-antibody screening is recommended for several adult populations, including the elderly and pregnant and postpartum women, and analysis of filter paper-dried blood spots (DBSs) has been a cost-effective means of screening newborns for hypothyroidism for more than 30 years. The aim of this study was to show that DBS specimens can be used to test adults for thyroid-stimulating hormone (TSH) antibodies and thyroid antibodies during screening for thyroid disease in a community setting. Thyroid peroxidase antibody and thyroglobilin antibody enzyme-linked immunosorbent assay kits for serum have been adapted for use with DBSs. A TSH assay for newborn DBSs was adapted for the screening of adults. Parallel specimens, serum and DBSs, were collected during routine care of patients attending a thyroid clinic. In addition, 962 DBS specimens were collected from volunteers at community centers and nursing homes. Twelve months later, a second specimen was collected from each of 411 original volunteers. Antibody results are reported as normal and positive. Ninety-seven percent of the serum/DBS results correlated exactly. Of 962 volunteers from nursing homes and community centers, testing for antibodies was positive or indeterminate (borderline) in 266 (27%). Eighty percent of the patients with an initially normal TSH reading who were positive on the 12-month collection were positive for at least one antibody in the first collection. The use of DBSs is a convenient and efficient way to screen for thyroid antibody as a means of detecting occult thyroid disease in adults.

The lanthanide fluorescence immunoassay was elaborated for quantitative determination of the total thyroxin T4 in bloodstain dried in filter paper; the fields of its clinical application were defined. The method is based on the hardphase concurrent immunoassay with specific monoclonal antibodies to T4 marked by chelates of europium ions and with conjugate of the T3 heterologous hapten sorbed in plate holes with bovine serum albumin. Measurements of the fluorescence intensity were made by a fluorometer in the time resolution mode. The method ensures the T4 determination in a dry bloodstain with a diameter of 3.4 mm within the concentration range of 0 to 400 nmol/l and with the variation coefficient of below or equal to 15%; the results correlated with the findings of the T4 analysis by the DELFIA Neo T4 set, "Wallac Oy", Finland. The method efficiency was demonstrated for screening and verifying the congenital thyroid deficiency in newborns; it was also confirmed that the method can be used for monitoring the functional thyroid condition in adult patients.

Objective: Symptoms of hypothyroidism in adults can be mistaken for medical and psychiatric diseases, as well as for general signs of ageing such as weakness, lethargy and fatigue. The incidence of hypothyroidism is many-fold higher in adults than in newborn children. The latter have been routinely screened for the condition using filter paper dried blood spots (DBS) for nearly three decades but this cost-effective screening technique has only recently been applied to adults. This study was undertaken to show that DBS testing in adults and older children is an accurate way to screen for hypothyroidism. Methods: Serum and DBS specimens were collected from adults and children. Assays were run on both specimens and the results correlated. In addition 972 specimens were collected from adults at community centres and nursing homes. Follow-up studies were performed on patients with positive results. Results: The correlation coefficient for 118 matched serum and DBS specimens was 0.99. Thyroid-stimulating hormone (TSH) values were elevated in 50 of the 972 adults from nursing homes and community centres. Thirteen of these individuals were on thyroid medication and 28 had either high serum TSH or high thyroglobulin (TgAb) or thyroid peroxidase (TPOAb) antibody levels. Conclusions: Individuals can be screened for hypothyroidism by collecting finger stick DBS specimens at community centres, nursing homes and other locations which can be mailed by regular postal service to a central laboratory for accurate and inexpensive testing.

A simple and reliable method of paired TSH assay was developed and used in screening for neonatal primary hypothyroidism. In this method, a paired assay is first done. Equal parts of the extracts of dried blood spots on filter paper (9 mm diameter) from two infants 4-7 days old are combined and assayed for TSH by double antibody RIA. If the value obtained is over the cut-off point, the extracts are assayed separately for TSH in a second assay to identify the abnormal sample. Two systems, A and B, with different cut-off points were tested. On the basis of reference blood samples (serum levels of TSH, 80 microU/ml in system A and 40 microU/ml in system B), the cut-off point was selected as follows: upper 5 (A) or 4 (B) percentile in the paired assay and values of reference blood samples in the second individual assay. Four cases (2 in A and 2 in B) of neonatal primary hypothyroidism were found among 25 infants (23 in A and 2 in B) who were recalled from a general population 41,400 infants (24,200 in A and 17,200 in B) by 22,700 assays. This paired TSH neonatal hypothyroidism.

The radioimmunoassay described measures TSH in dried whole blood spots collected from neonates onto filter paper Guthrie cards. Microgranular cellulose is added to the precipitating reagent at the critical separation stage of the assay to overcome imprecision caused by the presence of the filter paper sample disc in the tube. The method was developed for a regional neonatal screening unit and has been found to be very reliable during ten months' routine use. It was required to be as precise, sensitive, accurate, rapid, simple, and inexpensive as possible and suitable for use with automatic diluting equipment in order to process large numbers of samples. Other methods were examined for their suitability and found not to fulfil one or more of the above criteria.

A multicentre study for the detection of congenital hypothyroidism by enzyme-immunoassay of thyrotropin in filter paper blood spots is described. The method is accurate, with good precision and adequate sensitivity. No interference due to common blood constituents has been observed. Moreover, the TSH determination can easily be performed by photometric measurement of enzyme activity. Large series of samples can be processed by automation using common laboratory analysers. Values show good agreement with those obtained by RIA methods. We present a method for screening for congenital hypothyroidism which can be performed in all clinical laboratories.

An immunoradiometric two-site assay for thyrotrophin (TSH) in dried blood filter paper spots is described. The assay is automated by means of the Kemtek 3000 automated immunoassay system. The technique uses a 6.0 mm disc punched from the dried blood samples collected as part of the screening programme for phenylketonuria. The method is sensitive and precise, and results correlate well with those obtained in TSH assays of serum samples. The procedure is rapid, results being available within 24 h of receipt of samples. Of 25204 specimens so far screened by this assay, 99.9% have TSH levels less than 15 mU/1. One false positive result has been obtained and six confirmed cases of neonatal hypothyroidism detected, giving a prevalence of 1 in 4200.

We have simultaneously measured T4 and TSH concentrations from 93,000 consecutive filter paper blood samples received by the Quebec network for Genetic Medicine. T4 was measured using the Micromedic Automated System and TSH was measured by RIA kits from Pharmacia and Becton Dickinson, each for a 6-month period. Statistical analyses to assess quality control parameters for the various methods revealed that the T4 assay had greater precision and reproducibility than either of the 2 commercially available TSH kits. The number of false negative results was similar for both methods. Eight infants with ectopic thyroid glands were correctly detected by the T4 method. One infant with secondary hypothyroidism was detected by the T4 approach and missed by the TSH methodology. Three infants overall would have been missed using either the T4 or TSH approach. These data indicate that primary T4 screening has similar sensitivity compared to TSH kits for mass screening programs for congenital hypothyroidism.

Screening for hypothyroidism has been achieved by seeking raised concentrations of thyroid stimulating hormone (h-TSH) in dried blood samples collected on absorbent paper from some 1700 newborn babies each week using double antibody radioimmunoassay and 125I-label counting on a NE1600 gamma counter linked to a HP9815S desk top calculator. Specificity, accuracy, and quality control of the assay have been investigated. The calculator programme stores all count, time, and calculated data and uses log-log function formation and third-power homoscedastic polynomial fit to the calibration data followed, if necessary, by elimination of outliers set at p= 0·5 %. Branching in the calculation procedure provides the ability to process sample data in subgroups, completely to reprocess data (including presentation in a form suitable for making a histogram plot), and to reset without loss of data in case of instrument or main power failure. Variance analysis was used to select the minimum duration of count (50 s), and it was demonstrated that homoscedasticity pertains in the regression calculation.

We have recently developed an immunoassay that can measure thyroxine rapidly and accurately in the eluate of 40 mul of dried blood spotted on filter paper at the fifth day of life. The method is completely automated and by using the samples received by the Central Laboratory of the Quebec Network for Genetic Medicine and their follow-up facilities, we are now screening every newborn in the province of Quebec for neonatal hypothyroidism. To date, from 47,000 measurements, three newborn infants with abnormally low TBG and seven hypothyroid infants have been detected. From these data we conclude that the frequency of congenital hypothyroidism is about one in 7,000 births and that our method is effective in detecting thyroid hormone abnormalities with an acceptable percentage of false positive measurements; no false negative results have occurred to our knowledge.

We describe a sensitive, simple, and rapid two-site immunoenzymometric assay for thyrotropin in dried blood samples on filter paper, for use in screening for neonatal primary hypothyroidism. In this method, two dried-blood spots of 3 mm diameter (equivalent to about 5.4 microliter of blood) are incubated overnight with anti-thyrotropin-beta-D-galactosidase complex in an anti-thyrotropin coated tube. Then the enzyme activity in the washed tube is determined fluorophotometrically. The range of thyrotropin measurable is 10 to 160 milli-int. units/L blood. Values for thyrotropin in dried blood samples determined by this method and those determined by radioimmunoassay correlated highly (r = 0.96).

A method for measuring thyrotropin (TSH) in eluates of dried blood samples on filter paper was evaluated and improved as a screening test for neonatal hypothyroidism. A linear relationship between the volume of eluate and the TSH value and good recoveries of endogenous TSH (104%) and added TSH (89%) were obtained, indicating that TSH in dried blood was extracted well by overnight elution and determined accurately by radioimmunoassay. The TSH content in the central portion of a spot was less (71%) than that in the peripheral portion. The TSH in dried blood samples on filter paper was stable at 4 degrees C, room temperature (about 25 degrees C) or 37 degrees C for 1 month. The TSH values of eluates were correlated with those of whole blood (r = 0.90) and serum (r = 0.81). Cases of primary hypothyroidism cound be readily differentiated from normal subjects by this method, even when eluates of their blood were combined with those of normal blood for assay of TSH.

We have evaluated a time-resolved immunofluorometric assay (IFMA) for determining thyrotropin. This "sandwich"-type system involves two monoclonal antibodies directed against different epitopes. A linear relationship between signal and thyrotropin concentration was observed up to 6000 milli-int. units/L. This procedure takes one day, vs six days with our present RIA technique, and requires only a tenth as much sample. Furthermore, intra- and interassay CVs are lower than with RIA. Assay of 19 paper-disc blood specimens from newborns identified as having congenital hypothyroidism, both by RIA and by clinical evidence, also gave positive results with IFMA. In prospective assay of 3944 specimens by both methods we identified one case of congenital hypothyroidism, which was detected by both techniques. Technical false-positive reactions, identified as such by repeated analyses, were fewer with the IFMA method than with RIA.

We report the use of Staphylococcus aureus rich in protein A as an immunoadsorbent in the radioimmunoassay of thyrotropin and thyroxine in blood collected from newborn infants and dried on filter paper. The procedures for thyrotropin and thyroxine, which differ mainly with respect to concentration and volume of the Staphylococcus aureus suspension, are reliable, rapid, and technically simple. The ease of this method, coupled with its dependability, offers certain advantages over some of the commonly used techniques of separation, and therefore it provides a unique approach to measurement of these analytes in laboratory screening programs for neonatal hypothyroidism.

Filter paper discs saturated with dried blood can be used in the Immunotube solid-phase thyroxine radioimmunoassay. This assay utilizes polypropylene tubes to which antibody to thyroxine is covalently bound. The filter paper standards and samples are placed in the tubes, followed by an assay buffer that contains I125-labeled thyroxine and compounds to displace thyroxine from its binding proteins. After incubation, bound and free thyroxine are separated by aspirating or decanting the disc and buffer from the tube. The test can be used with 0.32 cm (1/8 inch) or 0.64 cm(1/4 inch) discs, and gives quantitative results that correlate well with those for serum samples. The intra-assay coefficient of variation is less than 10%. The assay may readily be mechanized with existing disc-punching equipment, and results of its use in mass screening programs are described.

A double antibody heterologous enzyme immunoassay was developed for measuring thyroxine (T4) in serum and in dried blood samples on filter paper for use in screening for neonatal hypothyroidism. In this method, ethanol extracts of 100 microliter of serum or dried blood spots of 9 mm in diameter were incubated with T4-alkaline phosphatase conjugate prepared with glutaraldehyde, and antiserum to T4-bovine serum albumin conjugate prepared with carbodiimide. The enzyme activity in the precipitates formed with second antibody was then determined photometrically at 500 nm. The measurable range of T4 was 1-32 micrograms/dl. A linear relation was obtained between the sample volume and the T4 value, and a high correlation was found between values for T4 determined by this method and those for T4 in serum determined by radioimmunoassay (coefficient of correlation: r=0.97 for serum and r=0.88 for dried blood samples).

We adapted a commercial immunoradiometric assay (IRMA) to measure thyrotropin in filter-paper blood spots. Two 3-mm blood spots are used for each standard and sample. These are incubated for 2 h with radiolabeled antibody and for 30 min with magnetic antibody, followed by a 10-min separation procedure. Assay sensitivity is 6 milli-int. units/L. Coefficients of variation (precision profile of the standard curve) ranged from 4.3 to 9.6%. The coefficient of correlation (r) between thyrotropin concentrations in the blood spots and in serum was 0.93. Pre-elution of the blood spots is necessary for short incubation time. Short incubation time, little need for specialized equipment, the high precision and sensitivity characteristic of IRMA, and ease of collection, transport, and storage of the blood-spot samples make this assay suitable for neonatal hypothyroid screening.

We describe a simple method for evaluating thyroxine-binding globulin capacity and concentration from a single 1-cm blood spot on filter-paper used in a screening program for neonatal hypothyroidism. This method permits prompt diagnosis of about 90% of the infants with thyroxine-binding globulin deficiency in our abnormal low-thyroxine, low-thyrotropin population. There was excellent equivalence between results obtained by our method and by the method of Chopra et al. (J. Clin. Endocrinol. Metab. 35:565, 1972), and minimal overlap between the population with low thyroxine-binding globulin and the low-thyroxine, normal thyrotropin population. We recommend this method to all programs in which a primary thyroxine measurement is used in screening for congenital hypothyroidism.

To reduce the number of false-positives in our screening program for neonatal hypothyroidism, we investigated the effect of age, body weight, and interval between sampling and analysis on results for filter paper spot thyroxine determination. The statistical analysis included an analysis of covariance. The age of the infant did not influence the results, the interval influenced them minimally, but weight did require a correction factor of about 0.16 ng/kg below the mean weight. This correction factor should reduce our false-positives from 0.75% to 0.60%. Furthermore, because all hypothyroid infants had results that fell more than 2.8 SD below the geometric mean, we intend to measure thyrotropin concentrations in the blood eluate of all the spots with a thyroxine concentration below that cut-off point.