0021-972X/78/4705-1028502.00/0 Journal of Clinical Endocrinology and Metabolism Copyright © 1978 by The Endocrine Society
Vol. 47, No. 5 Printed in U.S.A.
A New Method of Paired Thyrotropin Assay as a Screening Test for Neonatal Hypothyroidism* KIYOSHI MIYAI, TOSHIAKI OURA, MINORU KAWASHIMA, TSUNEO TSURUHARA, YUTAKA HASE, KIYOSHI ICHIHARA, NOBUYUKI AMINO, KEIKO NISHI, TOMIKO FUJIE, KIYOMI NAKATANI, MIZUO AZUKIZAWA, AND OSAMU NOSE Central Laboratory for Clinical Investigation (KM., K.I., N.A., K.N., T.F., K.N.), the Department of Medicine and Geriatrics (M.A.), and the Department of Pediatrics (O.N.), Osaka University Medical School; the Children's Medical Center of Osaka City (T.O., T.T., Y.H.); and the Osaka Kessei Laboratories (M.K.), Osaka Japan ABSTRACT. A simple and reliable method of paired TSH assay was developed and used in screening for neonatal primary hypothyroidism. In this method, a
and 40 juU/ml in system B), the cut-off point was selected as follows: upper 5 (A) or 4 (B) percentile in the
paired assay is first done. Equal parts of the extracts of
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 of 41,400 infants (24,200 in A and 17,200 in B) by 22,700 assays. This paired TSH assay system saves labor and expense for screening neonatal hypothyroidism. (J Clin Endocrinol Metab 47: 1028, 1978)
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 cutoff points were tested. On the basis of reference blood samples (serum levels of TSH, 80 /iU/ml in system A
B
ECAUSE the irreversible mental retardation of congenital hypothyroidism can be prevented by early treatment, the importance of early diagnosis of this disease has been emphasized. The difficulty in early diagnosis by clinical features alone has prompted the organization of various mass screening programs during the past 3 yr (1). Several methods for mass screening have been developed; namely, measurements of T4 (2-7) and/or TSH (3, 5-12) in cord blood (5-9) or in dried blood spots on filter paper (2-7, 9-12) collected for screening for other metabolic disorders. Recently, we developed a new "paired TSH assay method," in which samples from two infants are combined for measurement of TSH by sensitive RIA. This method of mass screening saves considerable labor and ex-
Received December 27, 1977. Address requests for reprints to: K. Miyai, Central Laboratory for Clinical Investigation, Osaka University Hospital, 1-1-50, Fukushima, Fukushima-ku, Osaka 553, Japan. * This work was supported in part by grants from the Ministry of Health and Welfare and the Ministry of Education of Japan.
paired assay and values of reference blood samples in
pense. Since 1975, we have screened babies by this method and have detected infants with congenital primary hypothyroidism in the general population. The present paper describes the method and an evaluation of our program. Materials and Methods Subjects A total of 41,400 dried blood samples from babies in the general population and 16 cases of primary hypothyroidism were tested. To screen babies in the general population, blood from 4- to 7-day-old infants, taken by heel puncture, was spotted on filter paper. The dried spots were assayed for TSH within 10 days. Immunoreactive TSH did not change during this time (12). Reference blood spots were prepared by spotting 30 \i\ blood from established cases of primary hypothyroidism and were stored frozen. Extraction procedure Two systems were used: system A was used in earlier experiments and system B in later ones.
1028
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TSH FOR HYPOTHYROID SCREENING System A. As shown in Fig. 1, dried blood spots of 9 mm diameter (equivalent to about 25 /xl blood) were extracted overnight with 300 /xl 0.01 M phosphate-buffered saline (PBS; pH 7.8). For the first paired assay, 200-/xl samples of the extracts (equivalent to about 16 JU.1 blood) from two subjects were mixed and the TSH content of the mixture was determined by a one-step method of RIA. For the second individual assay, the remainder of the extract (100 /il of the 300-ju,l extract, equivalent to about 8 fi\ blood) was removed and the spot was rinsed with 300 /u.1 PBS. The TSH content of the total extract (the remaining extract plus the rinsing fluid) from a single subject was then determined by the same RIA method. System B. Dried blood spots were extracted with 400 /xl PBS and 200-/xl samples of the extracts (equivalent to about 12 /xl blood) from two subjects were mixed and used for the first paired assays. In the second individual assays, the remainder of extract (200 /il of the 400-/il extract, equivalent to about 12 /xl blood) was removed and rinsed with 200 /xl PBS. In both assays, a two-step method of TSH RIA was used. The concentration of TSH in the eluate was converted to microunits per ml blood assuming that 25 /xl blood in one spot (9 mm diameter) are completely extracted. RIA of TSH, TA> and T 3 TSH was measured by RIA (13, 14) with modified double antibody methods as follows. Purified human TSH (5.5 U/mg) for use as a standard and for labeling with 125I, potent antihumanTSH rabbit serum (first antibody), and antirabbit y-globulin goat serum (second antibody) were obtained from Daiichi Radioisotope Laboratories and Eiken Immunochemical Laboratories. All reagents were prepared in 0.5% bovine serum albumin in PBS (diluent). subject - 1
1029
One-step method for system A. A mixture of 100 /il [125I]TSH (0.01-0.02 /xCi; SA, 100-150 /iCi//xg), 100 id anti-TSH, and 400 /xl eluate or 100 ill standard TSH (0.6-320 juU/ml) with 300 /il diluent was incubated at room temperature (about 25 C) overnight. Then 100 /xl second antibody were added and incubation was continued at 4 C overnight. The mixture was centrifuged at 2000 X g for 60 min, the supernatant was discarded, and the radioactivity of the precipitate was measured in a well-type scintillation counter. Two-step method for system B. In this method, the amount of anti-TSH was half that in system A and addition of [125I]TSH was delayed. A mixture of 50 /il anti-TSH and 400 /xl eluate or 100 /xl standard TSH with 300 /xl diluent was incubated at room temperature (about 25 C) overnight. Then 50-100 /xl [I25I]TSH were added and the mixture was incubated again overnight. Finally, 50-100 /xl second antibody were added and incubation was continued at 4 C overnight. The subsequent procedure was the same as in the one-step method. Mean percentages of [125I]TSH bound were about 32% (A) and 27% (B). The subjects recalled were examined by determining their serum TSH, T4, and T 3 concentrations. Serum T4 (15) and T 3 (16) were measured by modified double antibody RIA using commercial kits obtained from Eiken Immunochemical Laborato-
Results Comparison of TSH RIA methods The two-step method (B) was more sensitive than the one-step method (A). The 50% intercept in method B was 0.5 juU/tube, whereas that in method A was 2.0 ixU/tube; moreover, the minimum detectable TSH level
subject s 2
FIG. 1. Screening program for detecting congenital hypothyroidism by the paired TSH assay method.
reference blood spot (80(A) or 40(B)N V/'U mi serum /
reference blood spot /80(A) or 40(B)\ \fU mi serum '
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
JCK & M . 1978 No 5 Vol47
MIYAI ET AL.
1030
was about 0.06 /xU/tube with method B and 0.2 /xU/tube with method A. There was little difference between the standard curves obtained with kits from Daiichi Radioisotope Laboratories and Eiken Immunochemical Laboratories. Model experiment Figure 2 shows the results of a model experiment using the paired TSH assay method. Abnormal pairs (normal plus hypothyroid subjects) were clearly separated from normal pairs (normal plus normal subjects). The separation was clearer with system B than with system A. Cut-off point The over-all frequency distributions of TSH concentrations in the extracts are shown in Fig. 3. Reference blood spots were assayed for TSH in every assay as positive controls; the mean TSH values of these samples are indicated by arrows in Fig. 3. In the first paired assay of system A (Fig. 3a), the TSH values of the extract from the reference blood spot (serum TSH, 80 /xU/ml) were located at the 2.4 percentile of the general population. To avoid between-assay variability and false negatives, we selected the upper 5 percentile of samples (about twice the 2.4 percentile) in every assay. In system B (Fig. 3b), the reference blood spot (serum
TSH, 40 juU/ml) was located at the 2 percentjle level. Thus, samples with values in the upper 4 percentile were selected for the second individual assays. In the second individual assay in both systems A and B, subjects above values of the reference blood spots were selected for recall. These cut-off points are shown in Fig. 1. Results of screening Table 1 summarizes the results of screening for congenital hypothyroidism in the Osaka area from November 1975 to June 1977. With system A, of 24,200 infants tested in a general population, 44 were selected for recall; in fact, only 23 of these could be recalled. The serum TSH, T4, and T 3 values for these infants are shown in Table 2. Two infants (cases K. S. and K. I.) were diagnosed as cases of neonatal primary hypothyroidism. With system B, a total of 17,200 babies from a general population were screened and two subjects were selected for recall. These (Y. Y. and K. N.) were examined, tested, and diagnosed as cases of primary hypothyroidism. Discussion The reliability of TSH measurements by RIA in extracts of dried blood samples on filter paper in our laboratory has been reported previously (12). In these experiments, we tested the stability of TSH in dried blood
System A
System B
Serum T S H
FIG. 2. Model experiment of the paired assay method for TSH. Individual, Extracts of blood spots from primary hypothyroid patients (A, *, • , • , • , • ) and normal subjects (O) were assayed separately for TSH. Paired, Extracts from two subjects were combined and assayed for TSH as a normal pair (normal plus normal subject; ©) or an abnormal pair (normal subject plus hypothyroid patient; ®, ®, ®, ®, ®, ®) , Upper or lower limits.
{HU nt)
100
0 A • • • • *
80
""" nal 320 100 120 75 53 50
60
40
60
20 20
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
TSH FOR HYPOTHYROID SCREENING
1031
[ a ) First paired assay of system A 3000
2000 2.4
C, (40)
Percentile
K.S. (528A>U ml serum)
C. (BO)
1000
20
40
80
60
100
TSH in eluate (//U
ml blood )
200
300
over
FIG. 3. Distributions of TSH values in extracts of dried blood from infants in the first paired assay of system A (a) and system B (b).
( b ) First paired assay of system B
1000
500
(40)
10
20
C. K.N. (80) (316)
30
40
TSH in eluate
60 mfl blood )
spots, the reproducibility of extraction, the recovery of added TSH, the linearity of values on dilution, and the correlation of blood spot TSH values with serum TSH results. The sensitivity of the improved method (system B) is about 0.06 juU/tube, approximately 4 times that of system A. This higher sensitivity is comparable to that reported by Larsen et al. (0.066 juU/3 jul/tube) (11) and Pekary et al. (0.02 jiiU/tube) (14). Our method can recognize TSH concentrations as low as 5 /iU/ml blood, even using half the extract (200 /xl of 400 /d) from one blood spot of 9 mm diameter, equivalent to about 25 /il blood.
The principle of the paired TSH assay method for screening is as follows. Assuming that the blood concentration of TSH is under 20 /xU/ml in normal infants and over 50 juU/ml in hypothyroid infants, the TSH concentration in combined samples from two individuals should be less than 20 juU/ml for pairs of normal infants (normal plus normal), but more than 25 |iiU/ml in abnormal pairs (hypothyroid plus normal). Therefore, it should be possible to distinguish these pairs; this was in fact proved by the results of the model experiment shown in Fig. 2. When an abnormal pair is found in the first paired assay, the extracts in
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
1032
MIYAI ET
JCR&M • 1978 Vol47 • No 5
AL.
TABLE 1. Summary of results of screening tests for hypothyroidism on newborn infants No. of subjects System A
Total (%)
System B
24,200; 12,100 assays (100) 1st Paired assay 1,340; 1,340 assays 2nd Individual assay 44 (0.18) Recall requested 23 (0.1) Recalled and examined 2 (0.008) Primary hypothyroidism Percentage is in parentheses.
17,200;; 8,600 assays (100) 660; 660 assays 2 (0.01) 2 (0.01) 2 (0.01)
41,400; 20,700 assays (100) 2,000; 2,000 assays 46 (0.1) 25 (0.06) 4 (0.01)
TABLE 2. Summary of infants with primary hypothyroidism in screening tests Screening TSH on eluate (juU/ml blood) Case
Sex
System „ . , Paired
Examinations on recall Age
Individ- (weeks) , ual
Diagnosis
Serum TSH
Serum T4
Serum T., 105
F
A
236
172
12
(/lU/ml) 528
(Mg/dl) 2.2
(ng/dl)
K. S. K.I.
F
A
115
85
6
188
6.0
166
Y. Y.
F
B
114
136
6
680
4.9
202
K. N.
F
B
37
78
8
316
4.6
135
7.014.6*
192260''
2.84
Vol. 47, No. 5 Printed in U.S.A.
A New Method of Paired Thyrotropin Assay as a Screening Test for Neonatal Hypothyroidism* KIYOSHI MIYAI, TOSHIAKI OURA, MINORU KAWASHIMA, TSUNEO TSURUHARA, YUTAKA HASE, KIYOSHI ICHIHARA, NOBUYUKI AMINO, KEIKO NISHI, TOMIKO FUJIE, KIYOMI NAKATANI, MIZUO AZUKIZAWA, AND OSAMU NOSE Central Laboratory for Clinical Investigation (KM., K.I., N.A., K.N., T.F., K.N.), the Department of Medicine and Geriatrics (M.A.), and the Department of Pediatrics (O.N.), Osaka University Medical School; the Children's Medical Center of Osaka City (T.O., T.T., Y.H.); and the Osaka Kessei Laboratories (M.K.), Osaka Japan ABSTRACT. A simple and reliable method of paired TSH assay was developed and used in screening for neonatal primary hypothyroidism. In this method, a
and 40 juU/ml in system B), the cut-off point was selected as follows: upper 5 (A) or 4 (B) percentile in the
paired assay is first done. Equal parts of the extracts of
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 of 41,400 infants (24,200 in A and 17,200 in B) by 22,700 assays. This paired TSH assay system saves labor and expense for screening neonatal hypothyroidism. (J Clin Endocrinol Metab 47: 1028, 1978)
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 cutoff points were tested. On the basis of reference blood samples (serum levels of TSH, 80 /iU/ml in system A
B
ECAUSE the irreversible mental retardation of congenital hypothyroidism can be prevented by early treatment, the importance of early diagnosis of this disease has been emphasized. The difficulty in early diagnosis by clinical features alone has prompted the organization of various mass screening programs during the past 3 yr (1). Several methods for mass screening have been developed; namely, measurements of T4 (2-7) and/or TSH (3, 5-12) in cord blood (5-9) or in dried blood spots on filter paper (2-7, 9-12) collected for screening for other metabolic disorders. Recently, we developed a new "paired TSH assay method," in which samples from two infants are combined for measurement of TSH by sensitive RIA. This method of mass screening saves considerable labor and ex-
Received December 27, 1977. Address requests for reprints to: K. Miyai, Central Laboratory for Clinical Investigation, Osaka University Hospital, 1-1-50, Fukushima, Fukushima-ku, Osaka 553, Japan. * This work was supported in part by grants from the Ministry of Health and Welfare and the Ministry of Education of Japan.
paired assay and values of reference blood samples in
pense. Since 1975, we have screened babies by this method and have detected infants with congenital primary hypothyroidism in the general population. The present paper describes the method and an evaluation of our program. Materials and Methods Subjects A total of 41,400 dried blood samples from babies in the general population and 16 cases of primary hypothyroidism were tested. To screen babies in the general population, blood from 4- to 7-day-old infants, taken by heel puncture, was spotted on filter paper. The dried spots were assayed for TSH within 10 days. Immunoreactive TSH did not change during this time (12). Reference blood spots were prepared by spotting 30 \i\ blood from established cases of primary hypothyroidism and were stored frozen. Extraction procedure Two systems were used: system A was used in earlier experiments and system B in later ones.
1028
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TSH FOR HYPOTHYROID SCREENING System A. As shown in Fig. 1, dried blood spots of 9 mm diameter (equivalent to about 25 /xl blood) were extracted overnight with 300 /xl 0.01 M phosphate-buffered saline (PBS; pH 7.8). For the first paired assay, 200-/xl samples of the extracts (equivalent to about 16 JU.1 blood) from two subjects were mixed and the TSH content of the mixture was determined by a one-step method of RIA. For the second individual assay, the remainder of the extract (100 /il of the 300-ju,l extract, equivalent to about 8 fi\ blood) was removed and the spot was rinsed with 300 /u.1 PBS. The TSH content of the total extract (the remaining extract plus the rinsing fluid) from a single subject was then determined by the same RIA method. System B. Dried blood spots were extracted with 400 /xl PBS and 200-/xl samples of the extracts (equivalent to about 12 /xl blood) from two subjects were mixed and used for the first paired assays. In the second individual assays, the remainder of extract (200 /il of the 400-/il extract, equivalent to about 12 /xl blood) was removed and rinsed with 200 /xl PBS. In both assays, a two-step method of TSH RIA was used. The concentration of TSH in the eluate was converted to microunits per ml blood assuming that 25 /xl blood in one spot (9 mm diameter) are completely extracted. RIA of TSH, TA> and T 3 TSH was measured by RIA (13, 14) with modified double antibody methods as follows. Purified human TSH (5.5 U/mg) for use as a standard and for labeling with 125I, potent antihumanTSH rabbit serum (first antibody), and antirabbit y-globulin goat serum (second antibody) were obtained from Daiichi Radioisotope Laboratories and Eiken Immunochemical Laboratories. All reagents were prepared in 0.5% bovine serum albumin in PBS (diluent). subject - 1
1029
One-step method for system A. A mixture of 100 /il [125I]TSH (0.01-0.02 /xCi; SA, 100-150 /iCi//xg), 100 id anti-TSH, and 400 /xl eluate or 100 ill standard TSH (0.6-320 juU/ml) with 300 /il diluent was incubated at room temperature (about 25 C) overnight. Then 100 /xl second antibody were added and incubation was continued at 4 C overnight. The mixture was centrifuged at 2000 X g for 60 min, the supernatant was discarded, and the radioactivity of the precipitate was measured in a well-type scintillation counter. Two-step method for system B. In this method, the amount of anti-TSH was half that in system A and addition of [125I]TSH was delayed. A mixture of 50 /il anti-TSH and 400 /xl eluate or 100 /xl standard TSH with 300 /xl diluent was incubated at room temperature (about 25 C) overnight. Then 50-100 /xl [I25I]TSH were added and the mixture was incubated again overnight. Finally, 50-100 /xl second antibody were added and incubation was continued at 4 C overnight. The subsequent procedure was the same as in the one-step method. Mean percentages of [125I]TSH bound were about 32% (A) and 27% (B). The subjects recalled were examined by determining their serum TSH, T4, and T 3 concentrations. Serum T4 (15) and T 3 (16) were measured by modified double antibody RIA using commercial kits obtained from Eiken Immunochemical Laborato-
Results Comparison of TSH RIA methods The two-step method (B) was more sensitive than the one-step method (A). The 50% intercept in method B was 0.5 juU/tube, whereas that in method A was 2.0 ixU/tube; moreover, the minimum detectable TSH level
subject s 2
FIG. 1. Screening program for detecting congenital hypothyroidism by the paired TSH assay method.
reference blood spot (80(A) or 40(B)N V/'U mi serum /
reference blood spot /80(A) or 40(B)\ \fU mi serum '
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
JCK & M . 1978 No 5 Vol47
MIYAI ET AL.
1030
was about 0.06 /xU/tube with method B and 0.2 /xU/tube with method A. There was little difference between the standard curves obtained with kits from Daiichi Radioisotope Laboratories and Eiken Immunochemical Laboratories. Model experiment Figure 2 shows the results of a model experiment using the paired TSH assay method. Abnormal pairs (normal plus hypothyroid subjects) were clearly separated from normal pairs (normal plus normal subjects). The separation was clearer with system B than with system A. Cut-off point The over-all frequency distributions of TSH concentrations in the extracts are shown in Fig. 3. Reference blood spots were assayed for TSH in every assay as positive controls; the mean TSH values of these samples are indicated by arrows in Fig. 3. In the first paired assay of system A (Fig. 3a), the TSH values of the extract from the reference blood spot (serum TSH, 80 /xU/ml) were located at the 2.4 percentile of the general population. To avoid between-assay variability and false negatives, we selected the upper 5 percentile of samples (about twice the 2.4 percentile) in every assay. In system B (Fig. 3b), the reference blood spot (serum
TSH, 40 juU/ml) was located at the 2 percentjle level. Thus, samples with values in the upper 4 percentile were selected for the second individual assays. In the second individual assay in both systems A and B, subjects above values of the reference blood spots were selected for recall. These cut-off points are shown in Fig. 1. Results of screening Table 1 summarizes the results of screening for congenital hypothyroidism in the Osaka area from November 1975 to June 1977. With system A, of 24,200 infants tested in a general population, 44 were selected for recall; in fact, only 23 of these could be recalled. The serum TSH, T4, and T 3 values for these infants are shown in Table 2. Two infants (cases K. S. and K. I.) were diagnosed as cases of neonatal primary hypothyroidism. With system B, a total of 17,200 babies from a general population were screened and two subjects were selected for recall. These (Y. Y. and K. N.) were examined, tested, and diagnosed as cases of primary hypothyroidism. Discussion The reliability of TSH measurements by RIA in extracts of dried blood samples on filter paper in our laboratory has been reported previously (12). In these experiments, we tested the stability of TSH in dried blood
System A
System B
Serum T S H
FIG. 2. Model experiment of the paired assay method for TSH. Individual, Extracts of blood spots from primary hypothyroid patients (A, *, • , • , • , • ) and normal subjects (O) were assayed separately for TSH. Paired, Extracts from two subjects were combined and assayed for TSH as a normal pair (normal plus normal subject; ©) or an abnormal pair (normal subject plus hypothyroid patient; ®, ®, ®, ®, ®, ®) , Upper or lower limits.
{HU nt)
100
0 A • • • • *
80
""" nal 320 100 120 75 53 50
60
40
60
20 20
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
TSH FOR HYPOTHYROID SCREENING
1031
[ a ) First paired assay of system A 3000
2000 2.4
C, (40)
Percentile
K.S. (528A>U ml serum)
C. (BO)
1000
20
40
80
60
100
TSH in eluate (//U
ml blood )
200
300
over
FIG. 3. Distributions of TSH values in extracts of dried blood from infants in the first paired assay of system A (a) and system B (b).
( b ) First paired assay of system B
1000
500
(40)
10
20
C. K.N. (80) (316)
30
40
TSH in eluate
60 mfl blood )
spots, the reproducibility of extraction, the recovery of added TSH, the linearity of values on dilution, and the correlation of blood spot TSH values with serum TSH results. The sensitivity of the improved method (system B) is about 0.06 juU/tube, approximately 4 times that of system A. This higher sensitivity is comparable to that reported by Larsen et al. (0.066 juU/3 jul/tube) (11) and Pekary et al. (0.02 jiiU/tube) (14). Our method can recognize TSH concentrations as low as 5 /iU/ml blood, even using half the extract (200 /xl of 400 /d) from one blood spot of 9 mm diameter, equivalent to about 25 /il blood.
The principle of the paired TSH assay method for screening is as follows. Assuming that the blood concentration of TSH is under 20 /xU/ml in normal infants and over 50 juU/ml in hypothyroid infants, the TSH concentration in combined samples from two individuals should be less than 20 juU/ml for pairs of normal infants (normal plus normal), but more than 25 |iiU/ml in abnormal pairs (hypothyroid plus normal). Therefore, it should be possible to distinguish these pairs; this was in fact proved by the results of the model experiment shown in Fig. 2. When an abnormal pair is found in the first paired assay, the extracts in
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 12 November 2015. at 14:41 For personal use only. No other uses without permission. . All rights reserved.
1032
MIYAI ET
JCR&M • 1978 Vol47 • No 5
AL.
TABLE 1. Summary of results of screening tests for hypothyroidism on newborn infants No. of subjects System A
Total (%)
System B
24,200; 12,100 assays (100) 1st Paired assay 1,340; 1,340 assays 2nd Individual assay 44 (0.18) Recall requested 23 (0.1) Recalled and examined 2 (0.008) Primary hypothyroidism Percentage is in parentheses.
17,200;; 8,600 assays (100) 660; 660 assays 2 (0.01) 2 (0.01) 2 (0.01)
41,400; 20,700 assays (100) 2,000; 2,000 assays 46 (0.1) 25 (0.06) 4 (0.01)
TABLE 2. Summary of infants with primary hypothyroidism in screening tests Screening TSH on eluate (juU/ml blood) Case
Sex
System „ . , Paired
Examinations on recall Age
Individ- (weeks) , ual
Diagnosis
Serum TSH
Serum T4
Serum T., 105
F
A
236
172
12
(/lU/ml) 528
(Mg/dl) 2.2
(ng/dl)
K. S. K.I.
F
A
115
85
6
188
6.0
166
Y. Y.
F
B
114
136
6
680
4.9
202
K. N.
F
B
37
78
8
316
4.6
135
7.014.6*
192260''
2.84
