Somatomedin levels in the diagnosis and therapy of growth hormone deficiency

Serum somatomedin-C (SM-C) and somatomedin (SM) concentrations were measured by, respectively, radioimmuno (SM-C RIA) and radioreceptor assays (SM RRA) in 3 groups of children with short stature. The patient population was different from previously reported series in that it was urban Brazilian, low income, and significantly older. Group A consisted of 6 male and 3 female children, aged 7.7-16.0 years, whose average peak plasma immunoreactive growth hormone (GH) was above 10 ng/ml. Group B contained 8 male and 5 female untreated GH-deficient patients, ranging in age from 9.5 to 21.0 years. In Group C there were 4 male and 1 female GH-deficient subjects treated with I.M. injections of GH (0.1 U/kg) from 1 month to 7 years. The mean +/- SE basal RIA SM-C (ng/ml) concentrations were significantly lower in groups B (34.2 +/- 8.8) and C (43.8 +/- 13.7) than A (214.3 +/- 42.7): A X B, P less than 0.001 and A X C, P less than 0.02. Likewise the mean +/- SE basal RRA SM (ng/ml) concentrations were significantly lower in groups B (78.9 +/- 17.6) and C (90.8 +/- 19.3) than group A (316.3 +/- 43.0): A X B, P less than 0.001 and A X C, P less than 0.002. A significant linear correlation was observed between RIA and RRA in group B (r = 0.84; P less than 0.001) and C (r = 0.96; P less than 0.01), but not for A (r = 0.61; P greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Radioreceptor assay for atrial natriuretic factor

Interest in accurate measurement of atrial natriuretic factor (ANF) in biological fluids and various tissues has been stimulated by recent data indicating the possible role of ANF in the homeostasis of salt and water. The presence of high-affinity binding sites for ANF in rat glomeruli has allowed us to develop a rapid, sensitive, and simple radioreceptor assay (RRA). A saturable high-affinity binding site on the membranes of rat glomeruli has been characterized by a dissociation constant of 33 pM and binding capacity of 396 fmol/mg protein. Rat plasma extracts or atrial homogenates or standards were incubated with radioiodinated ANF and a preparation of rat glomerular membranes. The receptor-bound and free radioactivity were separated by filtration on Whatman GF/C paper after 1 h incubation at room temperature. The sensitivity of the RRA was 2.08 fmol. The effective concentration of standard ANF that displaced 50% of labeled receptor-bound ANF (EC50) was 43.3 +/- 2.6 fmol/ml (n = 7). Both intra- and interassay coefficients of variation were smaller than 11%. This RRA assay has been compared with radioimmunoassay (RIA). High correlations for 19 plasma extracts and 34 atrial homogenates (r = 0.973 and r = 0.954, respectively) tested by RRA and RIA were obtained. This good correlation between the two methods suggests that the immunoreactive material found in rat plasma and atrial homogenates also displays biological activity.     

A new spectrofluorimetric method for renal renin activity: a comparative study with other methods

A new spectrofluorimetric method for protein estimation has been developed and applied to study the renin-angiotensin system. The fluorometric reagent, 1-4-diamino-2-3-dichloro-anthraquinone, is introduced in this field and used for the first time. Renal semipurified rat renin is incubated with a synthetic substrate (N-acetyl-tetra-decapeptide) at 0 degrees C and 37 degrees C, at optimal pH, during 3 hours. The incubated mixture is studied by bioassay (BA), radioimmunoassay (RIA) and spectrofluorimetric analysis (SFA), and values obtained with these methods are compared. Blank samples for spectrofluorimetric analysis were prepared by substituting the incubated mixture with the unincubated components of the reaction. Its fluorescence values were subtracted from those of the incubated mixture. Precision and sensitivity for RIA and SFA were similar in both cases, but different in the case of BA. Renal renin activity (RRA) values for RIA (7.15 x 10-(3) nM/ml angio. I/3 h) and SFA (7.08 x 10-(3) nM/ml angio. I/h) were statistically equal (t = 1.05; p less than 0.05), while RRA values for BA (6.23 x 10-(2) nM/ml angio. I/3 h), were higher and significantly different from the statistical point of view. The graphical representation of RRA values for RIA versus SFA values gives an objective expression where RRA(RIA) = 0.78 RRA(SFA) + 0.02.     

Evidence that most of the radioimmunoassayable inhibin secreted by the corpus luteum of the common marmoset monkey is of a non-dimeric form.

Although recent data for several species of primate, including human and marmoset, indicate that the corpus luteum secretes high levels of radioimmunoassayable inhibin, the nature of the immunoreactive (ir) inhibin detected has not been established. In this study, plasma ir-inhibin levels during the ovarian cycle of the marmoset (n = 12 animals) were measured by alpha-subunit-directed inhibin RIA, and values were compared with those estimated by a recently developed two-site immunoradiometric assay (IRMA) specific for inhibin alpha-beta dimer. Consistent with earlier data, plasma levels of ir-inhibin measured by RIA (overall mean value 133 +/- 7 ng/ml; n = 171) reached values 4-fold higher (p less than 0.001) during the luteal phase (222 +/- 20 ng/ml) than during the follicular phase (58 +/- 8 ng/ml), being directly correlated with plasma progesterone levels (r = 0.65; p less than 0.001). In contrast, plasma ir-inhibin levels estimated by IRMA were substantially lower than those measured by RIA (overall mean value 9.62 +/- 1.08 ng/ml; n = 171) and did not vary significantly during the cycle. Administration of a luteolytic dose of cloprostenol during the late luteal phase/early pregnancy led to an abrupt fall in plasma concentrations of progesterone (95%) and alpha-inhibin measured by RIA (82%), whereas dimeric inhibin levels remained unchanged. Analysis of marmoset luteal extracts (n = 5) by RIA, IRMA, and inhibin bioassay yielded inhibin estimates of 102.6 +/- 21.0, 0.632 +/- 0.103, and less than 2.0 ng/mg, respectively, thus confirming that only a very small proportion of the inhibin produced was dimeric (i.e., bioactive) in nature.(ABSTRACT TRUNCATED AT 250 WORDS)     

Peripheral inhibin levels during estrous cycle in buffalo (Bubalus bubalis)

A sensitive, specific RIA was validated and used for measurement of peripheral plasma immunoreactive inhibin (irinhibin) levels during the estrous cycle in Murrah buffalo. The RIA employed an 125-I iodinated inhibin as tracer and an antiserum against dimeric inhibin. The procedure had a sensitivity of 16 pg/tube, and the nonspecific effects of buffalo plasma were compensated for by including 200 ul bullock plasma in the standards. Separation of free and bound inhibin was affected by the use of a second antibody and precipitation with polyethylene glycol. Blood samples were collected once daily for 30 d from Murrah buffalo (n = 6) during the hot month of July. Cyclic activity and estrus were confirmed by plasma progesterone determination. Peripheral plasma concentrations of ir-inhibin fluctuated between 0.40 +/- 0.07 and 0.67 +/- 0.13 ng/ml during the estrous cycle in buffalo. During the same period, plasma progesterone levels increased from 0.21 +/- 0.01 ng/ml at Day 0 to a peak of 3.30 +/- 0.72 ng/ml on Day 13, declining sharply by Day -5. Ir-inhibin levels exhibited an increase during the follicular phase, with the maximum concentration of 0.65 +/- 0.01 ng/ml occuring on the day of estrus, a decline thereafter, and no pattern during the luteal phase. The differences, however, were not statistically significant throughout the estrous cycle.     

Radioimmunoassay of a bovine pregnancy-associated glycoprotein in serum: its application for pregnancy diagnosis.

A sensitive and specific double-antibody RIA for a bovine pregnancy-associated glycoprotein (bPAG) is described. The limit of detection was 0.2 ng/ml. The assay was specific for bPAG in that pituitary and placental gonadotropic hormones and other placental or serum proteins assayed in serial dilutions did not cross-react. The RIA allowed measurement of bPAG in placental extracts, fetal serum, fetal fluids, and serum or plasma of pregnant cows. About 20% of unbred heifers and nonpregnant cows had detectable levels ranging from 0.30 +/- 0.09 to 0.50 +/- 0.17 ng/ml (mean +/- SD), and 15% of bull sera showed higher concentrations (3.01 +/- 1.73 ng/ml) of bPAG or bPAG-like protein. Variations among animals was observed in fetal serum bPAG concentrations. Bovine PAG was detected in maternal peripheral blood at Day 22 of pregnancy (mean +/- SD, 0.38 +/- 0.13 ng/ml) in some animals and at Day 30 in all pregnant cows. Peripheral serum bPAG levels increased progressively to 3.60 +/- 1.73 ng/ml (mean +/- SD) at Day 30 of pregnancy, to 24.53 +/- 8.81 ng/ml at Day 120, and to 1551.91 +/- 589.68 ng/ml at Day 270. Peak concentration of bPAG was 2462.42 +/- 1017.88 ng/ml and it occurred 1-5 days prior to parturition. After delivery, bPAG concentrations decreased steadily to 499.63 +/- 267.20 ng/ml at Day 14 postpartum (pp), 10.12 +/- 7.84 ng/ml at Day 60 pp, and 1.44 +/- 1.08 ng/ml at Day 90 pp. The undetectable concentration (less than 0.20 ng/ml) was reached by Day 100 +/- 20 pp. An investigation undertaken in Holstein heifers, Holstein cows, and Hereford cows used as recipients for purebred Holstein embryos supplied evidence of the influence of breed of recipient and sex of fetuses on peripheral concentrations of bPAG. A herd of 430 Holstein-Friesian heifers that had received transferred embryos were bled at Day 35 postestrus (pe) for measurement of bPAG. The bPAG was detected in 287 of 430 serum samples analyzed. By rectal palpation performed at Day 45 pe, 267 heifers with detectable levels of bPAG at Day 35 pe were confirmed to be pregnant as were 3 of 143 heifers previously diagnosed as not pregnant by RIA. These results suggest that detection of this placental-specific antigen in the serum could be used as a specific serological method for early pregnancy diagnosis in cattle from 28 days after breeding.     

Radioimmunoassay for insulin-like growth factor I (IGF-I) using biosynthetic IGF-I

We produced antiserum to insulin-like growth factor I (IGF-I), and developed a specific and sensitive radioimmunoassay (RIA) for IGF-I using the biosynthetic IGF-I. This antiserum to IGF-I was specific for IGF-I; no cross-reactivities with multiplication stimulating activity, porcine insulin or human growth hormone (hGH) were detected. The sensitivity was 10-25 pg/tube with 50% displacement at 125 pg/tube. The intra- and inter-assay coefficients of variation for IGF-I were 5.4 and 9.7%, respectively. The plasma IGF-I levels as determined by RIA in normal adults (N = 46), patients with active acromegaly (N = 31), and pituitary dwarfs (N = 31) were 21.6 +/- 1.0, 157.3 +/- 17.0, and 2.5 +/- 0.3 ng/ml (Mean +/- SEM), respectively, indicating the levels were GH-dependent. The plasma IGF-I levels were significantly increased from 2.2 +/- 0.2 to 26.5 +/- 3.2 ng/ml after hGH administrations for three consecutive days in five pituitary dwarfs. The IGF-I levels were low in patients with hypothyroidism and liver cirrhosis, but were normal in patients with chronic renal failure. These data confirm previous reports and this radioimmunoassay proves useful in evaluating plasma IGF-I levels.     

Use of highly specific antibodies against 17alpha-OH-progesterone in a simplified nonchromatographic RIA and in the simultaneous determination of four sex hormones in human plasma.

A highly specific antiserum raised against the 3(O-carboxymethyl)oxime of 17alpha-hydroxyprogesterone (17-OHP) was produced in rabbit and used in the development of a specific radioimmunoassay (RIA) for 17-OHP which included a celite chromatography. Methods allowing the measurement of plasma 17-OHP levels either separately or in combination with that of several plasma androgens (testosterone, delta4-androstenedione and dehydroepiandrosterone) are described. Moreover, RIA meabsurement of plasma 17-OHP levels with or without chromatographic (celite column) purification gave comparable results (mean +/- SD) in 29 normal adult males (118 +/- 34 ng/100 ml) and 35 normal adult females (follicular phase: 46+/- 16 ng/100 ml); luteal phase: 241 +/- 71 ng/100 ml).     

Determination of insulin-like growth factor-I in normal subjects and in patients with growth hormone disorders by radioimmunoassay using biosynthetic homologous peptide

A highly sensitive and specific RIA for IGF-I has been developed using recombinant DNA-derived IGF-I of very high purity and specific antiserum to it. This assay system could detect IGF-I at as low concentrations as 20-30 ng/ml. The intra-assay and interassay coefficients of variation at various concentrations of IGF-I were 4.9 to 6.5% and 5.4 to 8.0%, respectively. The recovery rate of pure IGF-I added to plasma was 77.0 +/- 3.7%. The antiserum did not cross-react with porcine insulin, biosynthetic human insulin, hGH, hEGF, the synthetic C-domain of IGF-I or that of IGF-II, but reacted equally with an analog, Thr59-IGF-I. Plasma IGF-I was extracted by the acid-ethanol method before assay to separate IGF-I from its binding protein. When plasma IGF-I was assayed without extraction, the inhibition curves of serial dilution of plasma samples from several individuals were not parallel to the standard curve of IGF-I. The plasma concentration of IGF-I was 147 +/- 49 ng/ml (mean +/- SD) in 156 normal adults aged from 20-59 years. As reported by others, the IGF-I levels were low in cord plasma (41.8 +/- 23.5 ng/ml) and plasma of patients with GH deficiency (64.6 +/- 42.0 ng/ml), while its levels were high in normal children of pubertal ages (12-13 yr, 365 +/- 126 ng/ml) and in patients with active acromegaly (562 +/- 115 ng/ml). This RIA system is a simple and useful method for determining plasma IGF-I in normal and diseased states.     

Plasma relaxin immunoactivity in the pig at parturition and during nuzzling and suckling.

One sow bled at 30--60-min intervals for 48 h at 5 and 4 days before parturition had mean +/- s.e.m. relaxin levels of 5.0 +/- 0.48 ng/ml and 5.5 +/- 0.44 ng/ml for each 24-h period respectively. This sow and another were bled at frequent intervals during parturition; both showed considerable fluctuations in their relaxin levels but no consistent peaks in relation to each birth. Mean levels during parturition were 10.7 +/- 0.46 ng/ml and 13.4 +/- 0.81 ng/ml respectively, both significantly higher than the levels at 4 and 5 days before birth. Relaxin levels in two lactating sows rose acutely during nursing, showing a 3-fold rise in one animal and an 8-fold rise in the other. Results from a third sow during an extended period of nuzzling and sucking by the piglets showed multiple peaks of relaxin immunoactivity associated with each nuzzling/sucking stimulus.     

High concentration of soluble HLA-DR in the synovial fluid: generation and significance in "rheumatoid-like" inflammatory joint diseases

In the search for its role in inflammatory joint diseases, soluble HLA-DR (sHLA-DR) was quantitated in 72 synovial fluids (SF) by a newly established immunoenzyme assay. Unlike other soluble receptors which accumulated only moderately (sCD25, sCD4) or negligibly (sHLA class I, sCD8) in the SF, SF sHLA-DR levels exceeded serum levels by up to 3 orders of magnitude and varied disease dependently from "control" values (traumatic synovitis and osteoarthritis: 9.9 +/- 6.1 ng/ml). Clear-cut different SF sHLA-DR values in HLA-DR-associated "rheumatoid-like" (136.5 +/- 130.0 ng/ml) vs HLA-B27-associated "spondylarthropathy-like" arthritic forms (28.4 +/- 29.1 ng/ml) were most significant comparing oligoarticular juvenile chronic arthritis type I (147.6 +/- 112.6 ng/ml) and type II (3.3 +/- 1.1 ng/ml), thus offering a new classification marker. Also ex vivo, large amounts of sHLA-DR were released spontaneously by SF mononuclear cells and found to be related to the T-cell activation state. SF sHLA-DR may be shed in large complexes or micelles, as it eluted mainly at >450 kDa on gel filtration. Western blotting revealed that the majority of SF sHLA-DR consisted of full-length alpha- and beta-chains. Minor fractions of smaller sized antigens seemed to be generated by proteolytic cleavage rather than by alternative splicing, since only minute amounts of HLA-DRB mRNA lacking the transmembrane exon could be amplified by RT-PCR. Distinct forms of high-dose sHLA-DR, able to provoke rather than to suppress T-cell responses, are discussed as contributing to some HLA-DR disease association.     

Insulinhypoglycemia but not arginine infusion stimulates circulating plasma growth hormone-releasing hormone (GHRH) concentrations in children

The effect of insulinhypoglycemia and arginine infusion on circulating concentrations of plasma growth hormone-releasing hormone (GHRH) and growth hormone (GH) has been studied in 24 children (4.4 to 14.3 years). Plasma GH and GHRH concentrations were determined by RIA. Basal plasma GHRH levels were detectable in the plasma of all patients ranging from 6.8 to 27.1 pg/ml. Injection of 0.1 U/kg body wt. insulin i.v. resulted in an increase of plasma GHRH levels (11.1 +/- 1.4 pg/ml vs. 18.8 +/- 2.6 pg/ml; P less than 0.01) preceding that of plasma GH (1.5 +/- 0.4 ng/ml vs. 13.6 +/- 1.3 ng/ml; P less than 0.01). Infusion of 0.5 gm/kg body wt. arginine hydrochloride did increase GH concentrations (2.0 +/- 0.6 ng/ml vs. 13.9 +/- 2.3 ng/ml; P less than 0.01) but did not change circulating plasma GHRH levels. Since the source of peripheral GHRH concentrations is not known the importance of these findings remains to be determined.     

Serum androstenedione and testosterone concentrations during pregnancy and nonpregnant cycles in dogs

Concentrations of testosterone and of androstenedione were determined by radioimmunoassay in serum samples collected every 2-5 days throughout the periovulatory and luteal phases of the ovarian cycles of pregnant and nonpregnant beagle bitches. Testosterone levels were consistently lower than those of androstenedione, reached peaks of 29 +/- 4 ng/dl near the time of the preovulatory luteinizing hormone peak, and were reduced to near the limits of detection (less than or equal to 5-10 ng/dl) throughout the luteal phase. Androstenedione levels reached preovulatory peaks of 73 +/- 13 ng/dl, were 54 +/- 7 ng/ml during early estrus, increased (P less than 0.05) to early luteal phase peaks of 76 +/- 8 ng/dl between Days 6 and 18, and then declined to 41 +/- 5 ng/dl by Day 35-40 in both pregnant (n = 8) and nonpregnant (n = 4) bitches. Subsequent protracted increases in androstenedione occurred in 4 of 8 pregnancies but in none of the nonpregnant bitches. From Days 42 to 64 the differences in mean levels between pregnant (45 +/- 2 ng/ml) and nonpregnant (32 +/- 3 ng/ml) bitches was not significant (P greater than 0.05). At parturition androstenedione levels fell (P less than 0.05) abruptly from 39 +/- 7 to 13 +/- 3 ng/dl. These results suggest that, in the bitch, androstenedione is the major circulating androgen during the follicular and luteal phases and that patterns of androstenedione levels during the luteal phase parallel those reported for progesterone in pregnant and nonpregnant bitches, including maintenance of elevated levels throughout gestation and an abrupt decline at parturition.     

A radioimmunoassay for rat ghrelin: evaluation of method and effects of nonylphenol on ghrelin secretion in force-fed young rats

Antiserum YJC 13-31 against the rat ghrelin conjugated to bovine serum albumin (BSA) was produced in the rabbit and a double antibody radioimmunoassay (RIA) for ghrelin has been developed. Characterization results of this antiserum revealed no cross-reaction with human growth hormone and somatostatin. Weak cross-reactions with insulin (0.1%), rat growth hormone (0.1%) and glucagon (0.3%) were observed, which scarcely interfered the assay system. The sensitivity of this RIA was 5 pg per assay tube. With the rat serum samples, the within-assay precision was 7.1% and the between-assay precision was 12.3%. The RIA was also available to detect the ghrelin in rat tissue extracts with good parallelism to the rat ghrelin standard. In application, the serum ghrelin and corticosterone levels in weaned rats were measured by RIA. Gavage of saline was sufficient to raise serum ghrelin from 2.6 +/- 0.18 to 6.7 +/- 0.7 ng/ml (P < 0.01). Gavage with nonylphenol (NP) suppressed the elevation of serum ghrelin levels in a dose-dependent manner. Besides, gavages of saline elevated the serum levels of corticosterone from 108.8 +/- 13.5 to 188.7 +/- 23.5 ng/ml (P < 0.01) but the elevation effects of corticosterone from gavages were overcome by NP in the low dose of 50 mg/kg. It can be speculated that ingestion of NP is harmful to young animals during growth and environmental adaptation.     

Demonstration of potent, gonadotropin-like biological activity in the serum of rats during midpregnancy

The gonadotropin-like activity (GnLa) of serum from pregnant rats was measured using the rat interstitial cell testosterone (RICT) bioassay. Serum GnLA was elevated on day 9 of pregnancy, peaked at 7.2 μg rat LH-RPl equivalents/ml on day 11 and declined to undetectable levels by day 15. Serum LH, measured by homologous RIA, was consistently low (<20 ng/ml) during pregnancy, except near term.Rat placental lactogen (rPL), which was measured in the same serum samples by rat radioreceptor assay (RRA), reached maximal concentrations on days 12 and 13 of pregnancy.These data suggest the presence in pregnancy serum of a potent-gonadotropin-like hormone, different from pituitary LH, whose origin is unknown. Furthermore, there are discrepancies between the times of appearance of this GnLA and rPL.     

Biosynthesis, characterization and inhibition of leukotriene B4 in human whole blood

We have evaluated the biosynthesis, characterization and inhibition of Leukotriene (LT) B4 in unstimulated and in A23187-stimulated human whole blood. LTB4 was assayed by radioimmunoassay (RIA) both in unextracted serum and after extraction and thin-layer chromatography (TLC). Unstimulated human whole blood allowed to clot at 37 degrees C for 60 min produced only trace amounts of LTB4 (0.16 +/- 0.05 ng/ml, mean +/- SD, n = 3). LTB4-like immunoreactivity (ir-LTB4) detectable in unstimulated serum samples was largely overestimated by direct RIA, most likely because of interfering substance(s) unrelated to cyclooxygenase or lipoxygenase activity. Incubation of human whole blood with A23187 (2-10 microM) resulted in a concentration-dependent stimulation of LTB4 production. At 10 microM A23187, ir-LTB4 was 18 +/- 2.4 ng/ml (mean +/- SEM, n = 28). In A23187-stimulated serum samples, LTB4 concentrations measured by direct RIA correlated in a statistically significant fashion with those measured after extraction and TLC. Nafazatrom added in vitro caused a dose-dependent inhibition of A23187-stimulated ir-LTB4 production with an IC50 of 17 microM.     

Radioimmunoassay of cortisol in peripheral blood plasma of buffaloes peripartum

Changes in the concentration of cortisol were observed in the jugular venous plasma of pregnant buffaloes on days 30, 15, 5, 2 and 1 prepartum, at partum, at regular 6-hr intervals up to 72 hr postpartum and on days 4, 6, 10, 18, 34 and 50 postpartum. A radioimmunoassay (RIA) procedure for cortisol standardized in the laboratory was used. Mean plasma cortisol levels showed little fluctuation (P<0.05) between days 30 and 2 prepartum with the values ranging from 1.28 +/- 0.23 to 1.46 +/- 0.13ng/ml. A small (but nonsignificant) rise in the hormone level was observed one day prepartum followed by a sharp increase to a high mean value of 3.78+/-0.36 ng/ml (P<0.05) at parturition. A sharp decline (P<0.05) to a low mean value was recorded within 6 hr postpartum followed by marked fluctuations in the hormone level up to 72 hr postpartum. The hormone levels subsequently varied narrowly between 1.74+/-0.39 and 2.01+/-0.27 ng/ml up to 50 days postpartum.     

Characterization of equine luteinizing hormone by chromatofocusing

Three equine luteinizing hormone (LH) preparations (eLH-A, -B, and -C) recently have been isolated in our laboratory and were shown to differ in average basicity (eLH-A greater than -B greater than -C). The present study further characterizes these preparations by chromatofocusing. Each of these preparations are comprised of a family of isohormones, with 5 major immunoreactive peaks in the pH range of 7 to 4 (approx. pIs = 6.6, 6.1, 5.7, 5.2, and 4.8), with varying amounts of material eluting to either side of the pH gradient. Although similar isoforms are seen in all three LH preparations, the relative proportions of different isoforms vary in a manner reflecting the average charge properties of eLH-A, -B, and -C. While eLH-A contains predominantly basic forms, eLH-C consists largely of acidic material, and eLH-B is composed mostly of isohormones with pIs intermediate to eLH-A and -C. Chromatofocusing of a crude extract from a single horse pituitary gland revealed isohormone peaks corresponding to those found in the highly purified LH preparations. Peak fractions of the various isoforms were used to generate a variety of activity ratios (LH bioactivity:LH radioimmunoassay (RIA), LH radioreceptorassay (RRA):LH RIA, LH bioactivity:LH RRA, follicle-stimulating hormone (FSH) RRA:LH RIA, and FSH RRA:LH RRA activity ratios). The LH bioactivity:LH receptor binding potency ratio showed a linear increase with increasing isohormone acidity (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of residual C-19 steroids in plasma and prostatic tissue of human, rat and guinea pig after castration: unique importance of extratesticular androgens in men

The concentrations of dehydroepiandrosterone (DHEA), its sulfate (DHEAS), androstenedione (A-dione), testosterone (T) and dihydrotestosterone (DHT) have been measured before and after castration in men and two animal models, namely the rat and the guinea pig. In adult men, the pre-castration levels of plasma DHEAS and DHEA were measured at 1839 +/- 320 and 2.4 +/- 0.5 ng/ml, respectively, while in both animal models, the concentrations of these two steroids were below 0.3 ng/ml. Orchiectomy in men reduced plasma T and DHT levels from 2.9 +/- 0.1 and 0.60 +/- 0.10 to 0.42 +/- 0.21 and 0.05 +/- 0.01 ng/ml (P less than 0.01), respectively, while there was no significant effect observed on DHEAS, DHEA and A-dione levels. By contrast, castration in the rat reduced the low levels of circulating DHEA and A-dione below the detection of the radioimmunoassay (RIA) used. In castrated guinea pig, a small quantity of plasma A-dione (0.07 +/- 0.02 ng/ml) was measured while DHEA was undetectable. Moreover, in the rat and guinea pig, plasma T and DHT levels became undetectable. Following administration of the antiandrogen Flutamide for two weeks in the castrated rat and guinea pig, prostate weight was not further reduced, thus indicating that there is no significant androgenic activity left following castration of these two species. In fact, castration in the rat and guinea pig caused a decrease in prostatic levels of DHT from 4.24 +/- 0.351 and 9.42 +/- 1.43 ng/g, respectively, to undetectable levels. In men, on the other hand, the prostatic DHT levels were only inhibited from 5.24 +/- 0.59 to 2.70 +/- 1.50 ng/g, respectively. As expected, when Flutamide was administered to the rat and the guinea pig, the levels of prostatic steroids remained undetectable while, in men, the DHT content in the prostate was further reduced to undetectable values. In summary, the plasma levels of DHEAS, DHEA, delta 4-dione are markedly different between men and both animal models used and furthermore, measurements of prostatic levels of androgens suggest that the high plasma levels of these steroids are likely responsible for the presence of important amounts of DHT in human prostate after castration.     

Evaluation of a new radioimmunological assay for the determination of the B subunit of creatine kinase enzyme

We used a new radioimmunological (RIA) kit for the assay of B subunit of creatine kinase enzyme (CK). This RIA system uses a specific antisera against the B subunit as ligant, human CK-BB labelled with 125I as tracer, and purified human CK-BB isoenzyme as standard. The mean (+/- SD) sensitivity obtained was 0.25 +/- 0.16 ng/tube and the between assay variability was about 9-10%. Serum levels of 113 normal subjects was not normally distributed. The 95% of values was found below 5 ng/ml. This new RIA is usefull in clinical practice when serum levels of CK-BB isoenzyme must be determined. This method is quickly and it is characterized by a good degree of precision, but the CK-MB isoenzyme cross-reacts for about 40% in this RIA system. Therefore, for the clinical diagnosis by means of this RIA it is necessary to rule out the concomitant elevations of serum CK-MB values.