Radioimmunoassay In Serum Research Articles

Midmolecular parathyroid hormone-related peptide in serum during pregnancy, lactation and in umbilical cord blood

Indications of an important physiological role of parathyroid hormone-related peptide (PTHrP) for fetal calcium homeostasis, maternal-fetal calcium transport and reproduction have accumulated over recent years. The PTHrP concentrations were measured by an earlier developed midregion radio-immunoassay in serum from lactating healthy females and umbilical cord blood and compared with levels in age-matched non-pregnant or lactating females. The PTHrP concentrations could be measured in all samples after silica cartridge C18 extraction of 10-12 ml of serum. The concentrations were significantly higher during lactation (mean +/- SD: 0.72 +/- 0.14 pmol/l, N = 22) and in umbilical cord blood (0.85 +/- 0.18 pmol/l, N = 12) compared with healthy age-matched women (0.48 +/- 0.09 pmol/l, N = 10, p < 0.001). The molecular forms of PTHrP were also studied in an age-matched control group, in pregnant women and in umbilical cord blood by gel chromatography in a fast protein liquid chromatography system of Sep-Pak-extracted pooled serum. In all three groups we found heterogeneity of the molecular forms with two predominant peaks. The smallest fragment had a molecular weight of 4-6 kD while the largest form appeared as a high-molecular-weight molecule. In conclusion, the concentrations of midmolecule PTHrP fragments in serum are elevated during lactation and in umbilical cord blood. Because the midregion of PTHrP has unique actions, our results indicate that PTHrP may play an important physiological role for the mother and for the maternal-fetal calcium transport.

Growth and the growth hormone axis in prepubertal children with asthma

To determine the influence of asthma and its treatment with inhaled corticoste-roids on growth, linear growth velocity, and the growth hormone axis in prepubertal children, we performed a longitudinal study for 12 months in 56 children with asthma, aged between 4.4 and 11.7 years. Height, weight, skin-fold thickness, and lung function were measured every 3 months and bone age at entry to and exit from the study. A 24-hour serum growth hormone concentration profile and fasting insulin-like growth factor I levels were measured halfway through the year. Seventy-four percent of boys and 62% of girls had heights below the 50th percentile. Growth velocity in the nonsteroid-treated control group (n = 13) was normal; 10 of 20 children taking beclomethasone grew slowly (14/20 used a dry powder device), and 4 of 19 children taking budesonide grew slowly (15/19 used a spacer). Three of four children using inhaled steroids and prednisolone grew slowly. In none of the treatment groups were measures of growth hormone secretion or levels of radioimmunoassayable serum insulin-like growth factor I affected. We conclude that slow growth in steroid-treated children with asthma does not appear to be associated with major perturbations in the growth hormone axis. (J P EDIATR 1995;126:297-303)

Eosinophil Cationic Protein in Perennial Allergic Rhinitis

For a quantitative investigation of eosinophil activation in perennial allergic rhinitis, eosinophil cationic protein (ECP) concentrations were measured by a radioimmunoassay in serum, nasal secretions (ECPWN) and in the supernatant of these nasal secretions (ECPsup) obtained from normal subjects and allergic patients. Levels of ECPWN and ECPsup were higher than that of ECPserum. ECPsup showed a positive correlation with clinical severity, despite the lack of a significant correlation with eosinophilia in nasal smears. ECPWN and ECPserum showed no significant correlation with any of these clinical parameters. There was a weak tendency toward an increase in histamine sensitivity of the nasal mucosa of allergic patients with higher ECPsup although this was not statistically significant. These results suggest accumulation and activation of eosinophils in the allergic nasal mucosa, and also indicate that ECPsup may be a clinical parameter of perennial allergic rhinitis.

Development and validation of a radioimmunoassay for follistatin in human serum.

Follistatin (FS/FSH-suppressing protein/activin-binding protein) is a single-chain glycoprotein, structurally distinct from inhibin, that has been shown to have inhibin-like activity in suppressing FSH secretion both in vivo and in vitro. The aim of these studies was to develop and validate a radioimmunoassay (RIA) for FS in human serum, and to describe the physiological variations of serum FS in humans. Serum was collected from normal men, and normal women in the follicular and luteal phases of the menstrual cycle. Clinical samples were also collected from male patients with hypogonadism, post-menopausal women and pregnant women in the first, second and third trimesters. A RIA for FS in human serum was developed using antisera raised against purified bovine FS and using bovine FS as tracer and standard. Serial dilutions of serum were non-parallel to purified bovine FS standard in the RIA. The addition of sodium dodecyl sulphate (SDS 0.05%) to the assay buffer resolved the non-parallelism suggesting that the interference of serum in the RIA was an assay matrix effect. To characterize the serum FS immunoactivity further, serum was fractionated by gel filtration on Sephadex G-100. At neutral pH, FS immunoactivity eluted as a major peak in the molecular weight range > 200 kDa. In 0.1 M HCl, a second peak of FS immunoactivity eluted in the molecular weight range 30-60 kDa consistent with the known sizes of FS. This suggested that FS was dissociating from a larger complex. Fractions taken from the low molecular weight region diluted in parallel with bovine FS in contrast with fractions from the high molecular weight region which were non-parallel. It is concluded that the non-parallelism of human serum in the FS RIA is due to the binding of serum FS to an unknown high molecular weight factor. This interference is eliminated by the inclusion of 0.05% SDS in the assay buffer. The RIA in 0.05% SDS has been applied to the description of the normal and pathophysiological variations of FS in human serum. There were no significant differences between FS levels in serum from hypogonadal men, normal women in the follicular phase of the menstrual cycle, post-menopausal women and pregnant women from the first, second and third trimesters. FS levels in serum of women in the luteal phase of the menstrual cycle were significantly lower than all other groups. FS levels in normal men were significantly higher than those in both luteal phase women and women in the first trimester of pregnancy (P < 0.05). These findings argue against a role for circulating follistatin in the control of gonadotrophin secretion and suggest that the gonads and/or conceptus are not the primary source of follistatin immunoactivity in serum. The lower follistatin levels in the luteal phase of the menstrual cycle remain unexplained.

Elevation of serum eosinophil cationic protein in primary biliary cirrhosis

It has recently been reported that in patients with primary biliary cirrhosis (PBC), eosinophils are not only increased in the peripheral blood, but also infiltrted in the liver portal tracts. There is general agreement that eosinophils and eosinophil cationic protein (ECP) released from eosinophils contribute to cellular damage, particularly in allergic inflammation. In the present study, ECP was measured by the radioimmunoassay in sera of patients with a variety of liver diseases including PBC. Serum ECP levels were significantly higher in patients with PBC than in those with chronic viral hepatitis, in those with liver cirrhosis, and in healthy subjects. There were no significant differences in serum ECP levels between symptomatic and asymptomatic PBC. The present study suggests that high levels of serum ECP may reflect high grades of eosinophil infiltration around the septal and interlobular bile ducts characterized by chronic non-suppurative destructive cholangitis, particularly in the early stages of PBC.

Serum lactogens possessed normal bioactivity in patients with lactation insufficiency

Insufficient breast milk is the most common reason for premature termination of breast-feeding. The causes of lactation insufficiency are usually multifactorial; in a small percentage of cases it is due to primary lactation failure of unknown origin. The aim of this study was to investigate whether lactation insufficiency of unknown origin could be caused by serum lactogens that had reduced biological activity. Women with lactation insufficiency of unknown origin and normal lactating controls were subjected to a standardized breast-feeding test for assessment of milk production. Thirty minutes later, serum samples were obtained for determination of total lactogen bioactivity, using an in-vitro bioassay, and levels of prolactin (PRL) and growth hormone (GH) using radioimmunoassay (RIA). Twelve lactating mothers with a clinical diagnosis of lactation insufficiency of unknown origin were compared with 12 matched mothers with normal lactation. The Nb2 lymphoma cell bioassay was used to measure total lactogen bioactivity in sera. Conventional RIA kits were used to estimate serum PRL and GH concentrations. Mean milk yield on standardized test feed was 21.6 ml for patients and 146.5 ml for controls. In both patient and control groups the total serum lactogen bioactivity ranged from about 150 to 5000 mIU/l, while the serum RIA (PRL+GH) levels ranged from about 350 to over 7000 mIU/l. There was no evidence of lactogens with reduced bioactivity in the patients' sera. Lactation insufficiency in the women studied cannot be explained by serum lactogens that possess unusually low bioactivity.

Monoclonal antibodies to bovine growth hormone potentiate hormonal activity in vivo by enhancing growth hormone binding to hepatic somatogenic receptors.

Administration of GH complexed with monoclonal antibodies (MABs) potentiates the in vivo actions of the hormone. In particular, growth and serum IGF-I concentrations of GH-treated hypophysectomized rats are increased by concomitant injection of anti-GH MABs. Among 37 anti-bovine GH (bGH) MABs, we selected one MAB with the most potentiating effects to investigate the mechanisms responsible for this phenomenon. Hypophysectomized rats were killed 18 h after a single s.c. injection of bGH (100 micrograms/rat), alone or complexed with increasing doses of MAB (4, 40, 400 micrograms/rat; MAB:bGH molar ratio: 0.005, 0.05, 0.5). IGF-I was measured by radioimmunoassay in acid-extracted sera and livers, whereas liver IGF-I mRNA was quantified by Northern blot hybridization. The in vivo occupancy of liver somatogenic (GH) receptors was derived from the determinations of total and free 125I-labelled bGH binding to liver homogenates treated with 4 mol MgCl2/l or water. Injection of MAB-bGH complexes enhanced body weight gain and raised serum IGF-I, liver IGF-I and liver IGF-I mRNA more than bGH alone (1.6-, 6-, 10- and 7-fold increases at the highest dose of MAB, compared with bGH alone; P < 0.001). These potentiating effects of the MAB were dose-dependent and significant potentiation of the growth response was already observed with the lowest dose of MAB. In vivo occupancy of liver GH receptors was markedly higher 18 h after injection of MAB-bGH complexes than after bGH alone, and this effect was also dose-dependent (receptor occupancy of 28%, 37% and 83% after 4, 40 and 400 micrograms of MAB respectively compared with 6% after bGH alone; P < 0.05, 0.05 and 0.001 respectively). In contrast, the in vitro binding of 125I-labelled bGH to liver homogenates was decreased in the presence of high doses of MAB. We conclude that low amounts of MABs complexed with bGH potentiate the stimulation by the hormone of liver IGF-I synthesis and secretion in a dose-dependent manner. These effects are mediated, at least in part, through changes in hormone-receptor interaction in vivo, leading to enhanced and/or prolonged binding of bGH to its somatogenic receptors.

Platelet-Derived Growth Factor Release and Antiplatelet Treatment with Low-Dose Acetylsalicylic Acid

Platelet-derived growth factor (PDGF) and beta-thromboglobulin (beta-TG) are released from alpha granules during platelet activation. PDGF may play a role in the development of atherosclerosis and the late restenosis after percutaneous transluminal coronary angioplasty (PTCA). The effect of acetylsalicylic acid (ASA) on PDGF release was studied in healthy volunteers before and twelve hours after ingestion of 300 mg ASA. PDGF, beta-TG, and thromboxane B2(TxB2) were measured by radioimmunoassay (RIA) in serum and in platelet rich plasma (PRP) after submaximal stimulation with collagen. TxB2 decreased significantly from 0.9 +/- 0.3 ng/(mL x 10(6) platelets) to 0.006 +/- 0.005 ng/(mL x 10(6) platelets) (mean +/- SD) in serum after ASA ingestion while PDGF and beta-TG remained unchanged. Measurements in PRP after stimulation with collagen showed a significant decrease in PDGF (from 21.5 +/- 1.4 pg/(mL x 10(6) platelets) to 1.8 +/- 4.1 (pg/mL x 10(6) platelets), in beta-TG (from 21.0 +/- 13.3 ng/(mL x 10(6) platelets) to 2.2 +/- 1.4 ng/(mL x 10(6) platelets)) and in TxB2 (from 143.6 +/- 80.7 pg/(mL x 10(6) platelets) to 0.5 +/- 0.6 pg/(mL x 10(6) platelets)) after treatment with ASA. In conclusion low-dose ASA inhibits collagen-induced release of both beta-TG and PDGF in PRP and TxB2-synthesis in PRP and serum.

INSULIN-LIKE GROWTH FACTOR-1: A PROGNOSTIC FACTOR OF KNEE OSTEOARTHRITIS

During a population survey in 1975-1978 persons with radiological osteoarthritis (ROA) of the knee were identified. After 12 years a follow-up study was conducted to study the effect of circulating insulin-like growth factor-1 (IGF-1) on cartilage loss, osteophyte growth and overall progression in 141 persons with confirmed ROA of the knee. The outcome measures were scored by comparing the radiographs taken at baseline and at follow-up. Insulin-like growth factor-1 was measured by radioimmunoassay in serum taken at follow-up and in 79% of the baseline sera. After adjusting for age, gender and body mass index at baseline, IGF-1 concentration at follow-up was related to osteophyte growth and overall progression. The adjusted odds ratio of the highest vs the lowest tertile was 2.96 (95% CI: 1.15-7.60) for osteophyte growth and 2.58 (1.01-6.60) for overall progression. No clear relationship was found with cartilage loss. These results were confirmed when baseline IGF-1 was studied. We conclude that the circulating IGF-1 concentration has an effect on the course of knee OA by influencing osteophyte formation but a preventive effect on cartilage loss could not be shown.

Myocardial damage following coronary bypass surgery: assessment with antimyosin antibody uptake.

To assess the role of 111In antimyosin antibody (AbAm) in the delineation of myocardial damage following coronary bypass surgery, we studied 51 consecutive patients who underwent coronary surgery, 27 of whom had a history of prior myocardial infarction. All patients underwent a diagnostic protocol comprising: (1) 99Tcm pyrophosphate (PYP) and AbAm injection 48 h after surgery (AbAm imaging 24 and 48 h post-injection) (myocardial/background and myocardial/lung ratios were obtained respectively from the computer image); (2) Radioimmunoassay (RIA) serum CK-B levels from samples obtained immediately before surgery, and 24 and 48 h later; (3) clinical and ECG follow-up. Twenty-five patients showed positive AbAm studies, 10 had positive PYP images, and 21 had CK-B levels above normal limits at 24 h. One patient with abnormal AbAm, PYP and CK-B studies had new Q waves on the ECG after surgery. This patient was considered to have sustained a peri-operative myocardial infarction. The large number of positive AbAm studies probably reflects myocardial damage frequently associated with coronary bypass surgery.

Transpulmonary gradient of type III procollagen peptides: acute effects of cardio-pulmonary bypass.

Type III procollagen N-peptides (PIIINPs) are believed to be released in stoichiometric amounts as type III collagen molecules are secreted from cells. We hypothesized that if the human lung actively produces type III collagen a detectable transpulmonary gradient in PIIINPs would exist in normal individuals that might be altered following a pulmonary insult. PIIINPs were therefore measured by radioimmunoassay in serum taken simultaneously from the pulmonary artery (PA) and left ventricle/aorta (LV) in 11 patients undergoing routine cardiac catheterisation. Mean PIIINP levels +/- SEM in LV were 66.8 +/- 5.4 micrograms.ml-1 and 59.9 +/- 4.1 micrograms.ml-1 in PA (p less than 0.04). In 6 patients, repeat measurements taken 4 h after cardiopulmonary bypass revealed a significant fall in PA values to 43.8 +/- 2.6 micrograms.ml-1 (p less than 0.001) and abolition of the transpulmonary gradient. These results suggest the adult human lung actively synthesis type III collagen and that, in the short term, cardiopulmonary bypass inhibits this process.

Effect of murine retroviral infection on hair and serum levels of cocaine and morphine

LP-BM5 retrovirally infected female C57BL 6J mice were administered cocaine, morphine or both by daily intraperitoneal injection for 9 weeks. Drug concentrations were measured by radioimmunoassay in serum and in hair extracts. Hair samples obtained from all drug-treated mice were positive for the drug injected, while none of the saline-treated mice had detectable drug levels in hair or serum. The average morphine concentration obtained from non-infected mice was 11 ng/mg hair whereas the amount found in the LP-BM5-infected mice was significantly higher (20 ng/mg hair). Mice injected with both morphine and cocaine were given 50% of the regular dose of each drug and drug levels in the hair of these animals were approximately half that of mice injected with the full dose of the single drug. Non-infected mice treated with both drugs had a mean value of 7 ng morphine/mg hair and 374 ng cocaine/mg hair while retrovirus -infected mice had significantly higher concentrations, 10 ng morphine/mg hair and 1160 ng cocaine/mg hair ( P < 0.001). Serum concentrations of cocaine and morphine were significantly higher ( P < 0.01) in the retrovirus-infected animals from 40 min to 1.5 h. The increased concentrations of cocaine and morphine in serum during retrovirus infection are accompanied by a significant increase in the amount of drug incorporated into the hair matrix. This change indicates that retroviral infection may influence the disposition of these drugs in the systemic circulation.

Immune inhibition of repair of canine skull trephine defects implanted with partially purified bovine morphogenetic protein

The healing of 14-mm trephine skull defects was observed in ten adult mongrel dogs. First and second set trephine operations were performed to determine whether xenogeneic bovine bone morphogenetic protein (bBMP) and associated bone matrix water-insoluble noncollagenous proteins (iNCP) incite an immunological humoral response inhibiting bone repair. The effects of immunization to BMP/iNCP were observed by serum radioimmunoassay, and by correlated roentgenographic and histological analysis of deposits of new bone. The first set implants of bBMP/iNCP induced 96% healing while the regeneration of the second set trephines was 34% less than the first set. The second set was associated with a significant increase in serum anti-BMP antibodies. While xenogeneic bBMP induced complete healing of trephine defects when implanted without previous immunization, and repair in response to a second set of bBMP/iNCP was always incomplete, further research with high purified recombinant BMP is required to measure immune effects in a statistically significant number of pure bred recipients.

Serum Concentrations of Extracellular Matrix Components

Laminin, hyaluronic acid (HA), procollagen III amino-terminal propeptide (PIIINP), and procollagen I carboxy-terminal propeptide (PICP) concentrations were measured by radioimmunoassay or radiometric assay in sera from 21 patients with glycogen storage disease (GSD). Laminin was increased in 16 of 29 samples from the six children with GSD I, 25/43 from the seven with GSD III, and 3/19 from the eight with GSD IX; laminin correlated with serum aspartate aminotransferase and gamma-glutamyltransferase but not with serum triglycerides, cholesterol, lactate, or urate. HA was increased in 20% of samples from GSD I, 58% from GSD III, and in none from GSD IX; HA correlated with serum lactate and urate but not with liver function tests, serum cholesterol, or triglycerides. Serum PIIINP was increased in only eight samples and PICP in only one; children with poor linear growth had low PIIINP and PICP. Immunostaining studies of nine liver biopsies taken at diagnosis showed increased laminin and PIIINP staining in portal tracts, fibrous septa, and around sinusoids in periportal regions; children with a greater degree of immunostaining did not always show significantly higher values of serum laminin or PIIINP. We speculate that raised serum laminin may reflect fibrogenesis (but not necessarily established fibrosis) in response to tissue damage in GSD, raised HA may reflect disturbed sinusoidal endothelial cell function, and low PIIINP and PICP impaired somatic growth rather than intrahepatic pathology.

Serum Tumor Necrosis Factor- Concentrations in Children Hospitalized for Acute Lower Respiratory Tract Infection

Tumor necrosis factor-alpha (TNF alpha) concentrations were measured by radioimmunoassay in sera of 118 children (median age, 1.7 years; range, 2 months-15 years) hospitalized for acute lower respiratory tract infection (ALRI). Both viral and bacterial ALRI were associated with elevated concentrations of TNF alpha. Concentrations greater than 40 ng/l were seen in children with bacterial or mixed ALRI in 64% and with viral ALRI in 50% of cases. Elevated concentrations were associated with longer duration of fever before admission (P less than .05) and with a higher serum C-reactive protein concentration (P less than .05). There were no significant differences in TNF alpha concentrations between gram-positive and gram-negative infections, nor was there an association with clinical severity of ALRI. TNF alpha concentrations decreased in most patients to normal within 5 days of hospitalization, irrespective of the etiology of the infection.

Correlation of squamous cell carcinoma antigen levels and treatment response in cervical cancer

Squamous cell carcinoma antigen (SCC) was measured by serum radioimmunoassay in 82 patients with primary (55) or recurrent (27) squamous carcinoma of the cervix, to determine correlation with the clinical presence of tumor and response to radiation and chemotherapy. Sixty-seven percent of those with newly diagnosed cancer and eighty-one percent of those presenting with recurrence had values above 2.5 ng/ml. In the newly diagnosed patients, 17% of those with stage I and 80% of those with stages II and above had abnormal values. Mean titers for stages I through IV were 2, 9, 19, and 144 ng/ml, respectively, confirming an association with tumor burden. Twenty-four patients underwent radiation therapy; eighteen of these had elevated pretreatment levels and were assessed for response. Titers accurately predicted regression and progression in all cases. Thirty patients received chemotherapy; twenty-seven of them had elevated pretreatment levels and were evaluable for response correlation. Titers predicted response in 96% of cases. An additional group of 20 patients successfully treated and with no evidence of disease was followed with SCC levels every 3 months. In 19 of these 20 patients (95%), the marker accurately predicted the status of disease. In 2 of these patients, who recurred while on the study, the SCC level increased before the tumor became clinically apparent. We conclude that SCC levels are useful in evaluating the response of primary and recurrent squamous cell carcinoma of the cervix to radiation and chemotherapy.

Placental Protein 4 as a Possible Tumor Marker in Ovarian Tumors

Placental protein 4 (PP-4), a recently characterized glycoprotein from human placenta, was studied using a specific double-antibody radioimmunoassay in sera of 130 volunteers, 76 ovarian tumor patients and in ovarian tumor cyst fluid and ascites of 21 patients. Elevated levels (greater than 3 micrograms/l) were found in 45 of 52 ovarian cancer patients (86.5%). PP-4 levels correlated significantly with staging. 31 patients with malignant ovarian tumor were monitored on 2-9 occasions during 5-82 weeks. Rising or falling levels of PP-4 correlated with progression or regression of disease in 25 of 31 instances (80.6%). Elevated levels were found in 10 of 24 benign and borderline ovarian tumors. Elevated PP-4 level does not indicate malignancy in each case. PP-4 can be regarded as tumor-associated antigen and an tumor marker in oncological practice.

Increased hGH Production Rate after Low-Dose Estrogen Therapy in Prepubertal Girls with Turner's Syndrome

Low-dose estrogen therapy significantly increases radioimmunoassayable serum hGH concentrations in the prepubertal hypogonadal female. In this study, we have examined the effects of short- and long-term low-dose ethinyl estradiol therapy on the endogenous production rates and metabolic clearance rates of hGH. We used deconvolution mathematical modeling to provide quantitative estimates of individual secretory parameters and to calculate subject-specific hGH metabolic clearance rates, by using all serum hGH concentrations and their variances considered simultaneously. Nine girls with Turner's syndrome (mean age 7.7 +/- 0.5 y) were studied on three separate nights by drawing blood every 20 min from 2200 to 0800 h before (I), after 1 wk (II), and 5 wk (III) of 100 ng/kg/d ethinyl estradiol therapy orally. We found that the endogenous hGH production rate more than doubled in all patients studied after 5 wk of ethinyl estradiol therapy (194 +/- 22 (I), 290 +/- 43 (II), and 412 +/- 66 (III) micrograms/L/12 h; p less than 0.05 for I and III). The half-life of endogenous hGH was not altered in the estrogen treatment paradigm with a mean of 19 +/- 1.6 min in study I and 18 +/- 1.2 min in both studies II and III. Our results suggest that even prepubertal concentrations of gonadal steroids in the hypogonadal female may be physiologically relevant to the maintenance of normal somatotrope secretory function.

Serum of patients with prostatic cancer or benign prostatic hypertrophy contains nonpolar testosterone

We have previously described a nonpolar form of radioimmunoassayable serum testosterone (NPT) not measured by available antitestosterone antibodies. It is detected by mild alkaline hydrolysis of the petroleum ether extract of serum and subsequent radioimmunoassay. The properties of NPT are consistent with that of a fatty acid ester of testosterone or dihydrotestosterone. The serum of young males contains 1 to 3 ng/ml NPT, but it is not detected infemale serum. We measured serum testosterone and NPT levels in 36 men between 58 and 87 years of age. Seventeen subjects with advanced prostatic cancer (NPT 1.70 ± 1.44 ng/ml) were compared with a control group consisting of six patients with benign prostatic hypertrophy (BPH) and 13 patients with no prostatic disease (NPT 0.72 ± 0.46, P = 0.07). There was no significant difference between BPH patients and patients with no prostatic disease; the results were pooled. The concentration of NPT in prostatic cancer patients but not in controls was inversely correlated with that of testosterone. Immunoassayable testosterone was present in the serum of two orchiectomized patients and, therefore, cannot derive solely from the testes. (Steroids 55:491–494, 1990)

Selenium and amino acid composition of selenoprotein P, the major selenoprotein in rat serum.

Selenoprotein P is the second plasma selenoprotein to be purified. It is a glycoprotein and has been shown to be distinct from plasma glutathione peroxidase. This study characterizes selenoprotein P further. Deglycosylation of the protein shifts its migration on sodium dodecyl sulfate-polyacrylamide gel electrophoresis from Mr 57,000 to Mr 43,000, indicating it has a substantial carbohydrate component. Measurement of selenium indicates a selenium content of 7.5 +/- 1.0 atoms/molecule based on a polypeptide weight of 43,000. Amino acid analysis accounts for all the selenium as selenocysteine. The protein is also rich in cysteine (17 residues) and histidine (23 residues). Fragmentation of selenoprotein P by trypsin and by cyanogen bromide produces peptides with varying selenium content. This indicates that selenium-rich regions of the protein exist. The concentration of selenoprotein P determined by radioimmunoassay in serum from control rats is 26.3 +/- 4.5 micrograms/ml and in serum from selenium-deficient rats it is 2.7 +/- 0.8 micrograms/ml. Depletion of selenoprotein P from control serum using an immunoaffinity column indicates that over 60% of serum selenium in the rat is contained in this protein. These results demonstrate that selenoprotein P is the major form of selenium in rat serum. It is the first selenoprotein described which has more than one selenium atom/polypeptide chain.