RESUMO
BACKGROUND: Placental growth factor (PlGF) is a proposed first-trimester screening marker for pre-eclampsia. This study investigates the stability of PlGF in serum and whole blood at typical routine storage temperatures. METHODS: Serum pools were stored at refrigerator temperature, room temperature or 30 °C for up to 30 days, or exposed to up to six freeze-thaw cycles. Whole blood was stored at room temperature or 30 °C for up to 6 days. PlGF was quantified using a DELFIA Xpress analyser. RESULTS: Placental growth factor levels increased over time, seemingly because of the dissociation of PlGF bound to a soluble binding protein, sFlt-1. Increase was slow in serum at refrigerator temperature, remaining stable (less than 10% change from start point) for at least 30 days. At room temperature PlGF was stable for 3.3 days and at 30 °C for 1 day. Serum PlGF remained stable for at least six freeze-thaw cycles. In whole blood, instability was worse, being stable for only 19.4 h at room temperature and just 3.3 h at 30 °C. CONCLUSION: Routine screening of sample handling requires careful monitoring. However, no extra precautions need to be taken when PlGF is used for pre-eclampsia screening run alongside existing first trimester aneuploidy screening programs that include hCGß.
Assuntos
Pré-Eclâmpsia/diagnóstico , Proteínas da Gravidez/sangue , Biomarcadores/sangue , Feminino , Humanos , Programas de Rastreamento , Fator de Crescimento Placentário , Gravidez , Primeiro Trimestre da Gravidez , Estabilidade Proteica , Análise de RegressãoRESUMO
Parenteral administration of human chorionic gonadotropin (hCG) or luteinizing hormone (LH) stimulates the production of testosterone in males and these gonadotropins can therefore be used by athletes to enhance muscle strength. However, they are more expensive and less efficient than testosterone and anabolic steroids. Therefore their main use is probably to stimulate gonadal testosterone production during and after self-administration of testosterone or anabolic steroids. A positive effect of hCG on muscle strength has not been demonstrated in women and elevated concentrations of hCG in females are often caused by pregnancy. The use of gonadotropins is therefore prohibited only in males but not in females. HCG occurs at low but measurable concentrations in plasma and urine of healthy males and can be measured by sensitive methods. However, the characteristics of the method to be used for doping control have not been defined. Virtually all commercially available hCG assays have been designed for determination of hCG in serum rather than urine, which is used for doping control. Methods based on mass spectrometric detection of fragments derived from hCG extracted from urine by immunoadsorption have been developed but their suitability for doping control remains to be determined. The concentrations of LH in serum and urine are variable and more then 10-fold higher than those hCG. It is therefore difficult to detect illicit use of LH. The characteristics and reference values for hCG and LH assays used in doping control and the cutoff values need to be defined.
Assuntos
Gonadotropina Coriônica/farmacologia , Dopagem Esportivo , Hormônio Luteinizante/farmacologia , Feminino , Humanos , Masculino , Força Muscular/efeitos dos fármacos , Fatores Sexuais , Detecção do Abuso de Substâncias/métodos , Testosterona/biossínteseRESUMO
We conducted a retrospective seroepidemiological study to evaluate the relationship between past chlamydial infection and primary fallopian tube carcinoma (PFTC). Postoperative serum samples were drawn from 79 consecutive patients treated for PFTC in 1985-2000. For each case two controls were selected. Serum samples were analysed for IgG antibodies to different C. trachomatis serotype pools and to C. pneumoniae. Seropositivity in general or serum antibody levels to different C. trachomatis serovars or C. pneumoniae did not differ between PFTC patients and controls. The lack of association between anti-chlamydial antibodies and PFTC suggests that past chlamydial infection does not play a role in the etiopathogenesis of PFTC. However, because chlamydial infection is common at young age and PFTC develops decades later, we cannot definitively exclude the possibility that C.trachomatis contributes to the development of PFTC.
Assuntos
Infecções por Chlamydia/complicações , Neoplasias das Tubas Uterinas/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Bactérias/análise , Estudos de Casos e Controles , Chlamydia trachomatis/imunologia , Chlamydophila pneumoniae/imunologia , Feminino , Humanos , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
Human choriogonadotropin (hCG), its free beta subunit (beta hCG), and the core beta hCG fragment (c beta hCG) were measured by highly sensitive time-resolved immunofluorometric assays in the serum and urine of 29 patients with pancreatic cancer, 7 patients with biliary cancer, and 45 patients with benign pancreatic or biliary diseases. The results were compared with those of an age- and sex-matched reference population of nonpregnant women and men. Of the various forms of hCG assayed in serum, beta hCG showed the best diagnostic accuracy, and c beta hCG was the best marker in urine. Elevated serum concentrations of beta hCG were observed in 72% of the patients with pancreatic cancer, in 6 of 7 patients with biliary cancer, and in 9% of those with benign disorders. The serum concentrations of c beta hCG were elevated in 45%, 57%, and 2%, respectively, and those in urine in 55%, 71%, and 11%, respectively. The molar concentrations of c beta hCG in serum were mostly lower than those of beta hCG. Thus beta hCG secreted into serum appears to be the main source of c beta hCG in urine. Provided that they are measured by sufficiently sensitive and specific assays, beta hCG in serum and c beta hCG in urine appear to be useful markers for pancreatic and biliary cancer.
Assuntos
Neoplasias dos Ductos Biliares/sangue , Neoplasias dos Ductos Biliares/urina , Carcinoma Intraductal não Infiltrante/sangue , Carcinoma Intraductal não Infiltrante/urina , Gonadotropina Coriônica/sangue , Gonadotropina Coriônica/urina , Neoplasias Pancreáticas/sangue , Neoplasias Pancreáticas/urina , Adulto , Idoso , Colestase/sangue , Colestase/urina , Gonadotropina Coriônica/química , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pancreatite/sangue , Pancreatite/urina , Fragmentos de Peptídeos/sangueRESUMO
We have studied the forms of prostate-specific antigen (PSA) in serum of patients with prostatic cancer and benign prostatic hyperplasia. Fractionation of serum by gel filtration and assay of the fractions for PSA showed that a considerable part of the PSA immunoreactivity in serum consisted of complexes that were larger than PSA. The complexes were assayed by time-resolved immunofluorometric assays based on an antibody against PSA on the solid phase and europium-labeled antibodies against various protease inhibitors as indicator antibodies. In addition to its monomeric form, PSA was found to occur in complex with alpha 1-antichymotrypsin. The proportion of the alpha 1-antichymotrypsin complex was a major form of PSA and it increased with increasing PSA concentrations, being over 85% at PSA levels exceeding 1000 micrograms/liter. A complex with alpha 1-protease inhibitor was also observed in serum of patients with prostatic cancer and very high levels of PSA. Complexes with alpha 2-macroglobulin and inter-alpha-trypsin inhibitor were detected, but their concentrations were low and similar in sera of cancer patients, normal men, and normal women, suggesting that they were not prostate derived. Commercial immunoradiometric assays for PSA were found to measure free PSA and its complexes with alpha 1-antichymotrypsin but not the complexes with alpha 2-macroglobulin and inter-alpha-trypsin inhibitor. The proportion of the PSA-alpha 1-antichymotrypsin complex was higher in patients with prostatic cancer than in those with benign hyperplasia. Therefore, assay of the complex had a higher sensitivity for cancer than assay of total PSA immunoreactivity.
Assuntos
Antígenos de Neoplasias/sangue , Neoplasias da Próstata/sangue , alfa 1-Antiquimotripsina/sangue , Antígenos de Neoplasias/análise , Humanos , Imunoensaio/métodos , Substâncias Macromoleculares , Masculino , Peso Molecular , Antígeno Prostático Específico , Neoplasias da Próstata/imunologia , alfa 1-Antiquimotripsina/análiseRESUMO
The activation of protein kinase C and adenylate cyclase in the regulation of human CG (hCG) synthesis by cultured BeWo choriocarcinoma cells was studied. Both cholera toxin (CT), which activates adenylate cyclase, and 12-O-tetradecanoyl phorbol-13-acetate (TPA), a protein kinase C activator, stimulated the secretion of hCG in a dose-dependent manner. During a 72-h culture, stimulation with the maximal effective concentration of CT (1.0 ng/ml) exerted a more pronounced increase (16-fold) in hCG synthesis than TPA (10 ng/ml) (2.8-fold), whereas the inactive phorbol ester 4 alpha-phorbol 12,13-didecanoate (100 ng/ml) was ineffective. When added together, TPA potentiated the effect of CT on hCG secretion (from 16- to 27-fold) and cAMP accumulation (from 36- to 54-fold) in the medium. TPA (1.0 ng/ml) also caused a 2.0-fold increase in basal cAMP production after 72 h. Time-course studies indicated that the effect of TPA on CT-induced cAMP and hCG productions became significant at 45 min and 6 h, respectively, from the beginning of stimulation. Proliferation of cells was not responsible for the responses, since the treatments only slightly increased the total protein content and did not alter the rate of incorporation of C3H3-methylated thymidine of the cells. Our results demonstrate that TPA potentiates CT-induced cAMP and hCG production in cultured human choriocarcinoma cells.
Assuntos
Coriocarcinoma/metabolismo , Gonadotropina Coriônica/metabolismo , AMP Cíclico/biossíntese , Acetato de Tetradecanoilforbol/farmacologia , Neoplasias Uterinas/metabolismo , Adenilil Ciclases/metabolismo , Linhagem Celular , Toxina da Cólera/farmacologia , Sinergismo Farmacológico , Ativação Enzimática/efeitos dos fármacos , Feminino , Humanos , Cinética , Gravidez , Proteína Quinase C/metabolismoRESUMO
We have developed a time-resolved immunofluorometric assay (IFMA) for quantification of the core fragment of the beta-subunit (c beta hCG) of hCG. The assay uses two monoclonal antibodies. One antibody is immobilized onto the wall of a microtiter strip well, and the other one is labeled with a europium chelate. The assay is sensitive (0.44 pmol/L = 4.6 ng/L), but cross-reaction with free beta-subunits of hCG (beta hCG) prevents direct determination of c beta hCG in serum. To circumvent this limitation we separated hCG and beta hCG from c beta hCG by gel chromatography and quantified each component in the fractions by specific IFMAs. The high sensitivity of the newly developed IFMA enabled us to demonstrate that serum from pregnant women contains c beta hCG and elutes at the same position in gel chromatography as c beta hCG purified from urine. The proportion of c beta hCG to hCG in pregnancy serum was 0.012-0.045% (mean +/- SD, 0.028 +/- 0.01) on a molar basis. Our finding of c beta hCG in serum confirms earlier reports suggesting that proteolytic degradation of beta hCG in the kidneys may not be the only pathway by which c beta hCG is formed.
Assuntos
Gonadotropina Coriônica/sangue , Fragmentos de Peptídeos/sangue , Gravidez/sangue , Adulto , Gonadotropina Coriônica/isolamento & purificação , Gonadotropina Coriônica/urina , Gonadotropina Coriônica Humana Subunidade beta , Cromatografia em Gel , Reações Cruzadas , Feminino , Fluorimunoensaio , Meia-Vida , Humanos , Rim/metabolismo , Fragmentos de Peptídeos/isolamento & purificação , Fragmentos de Peptídeos/urina , Gravidez/urinaRESUMO
The increases in serum immunoreactive (RIA) LH and FSH concentrations during puberty are small and of limited value in the evaluation of pubertal development. We, therefore, used highly sensitive time-resolved immunofluorometric assays to evaluate the changes in LH and FSH during female puberty. The sensitivity of the LH assay was 0.02 IU/L, and that of the FSH assay was 0.01 IU/L. Fifty normal premenarcheal girls, 7-12 yr old, 15 postmenarcheal girls, 16 to 17 yr old, and 15 adult women, 24-29 yr old, were studied. In postmenarcheal women, the blood samples were taken on cycle days 4-7. Serum estradiol concentrations were measured by RIA, and pubertal stages were graded. Serum LH levels in prepubertal girls were very low; the mean concentration was 0.05 IU/L. All girls less than 10 yr of age had serum LH concentrations below 0.2 IU/L, while FSH levels varied from 0.3-2.0 IU/L. The earliest significant changes in serum LH, FSH, and estradiol levels took place simultaneously at 9-10 yr of age. The increase in serum FSH was gradual, but the increase in serum LH was sudden and very steep, coinciding with the increase in serum estradiol and the onset of physical puberty. The increase in the mean LH concentrations between the 7-yr-old and the adult group was 116-fold, that for estradiol was 12-fold, and that for FSH was 6.7-fold. These results suggest that the increase in serum LH is important at the onset of puberty, and LH concentrations are a sensitive indicator of pubertal development.
Assuntos
Hormônio Luteinizante/sangue , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Criança , Estradiol/sangue , Feminino , Fluorimunoensaio , Hormônio Foliculoestimulante/sangue , Humanos , Pessoa de Meia-Idade , Fatores de TempoRESUMO
To explore the diurnal variation and the relationship between serum hCG levels and thyroid function during pregnancy, 26 women with an uncomplicated early pregnancy were studied before and after interruption of pregnancy. The high serum hCG levels in early pregnancy were accompanied by an increase in serum thyroid hormone and a decrease in serum TSH levels. Nevertheless, serum TSH exhibited diurnal variation similar to that in nonpregnant women. The nocturnal surge of TSH exceeded the daytime nadir by 112% and was distinctly different from the normal serum cortisol variation. The diurnal serum T4 and hCG variations were similar to the variation in serum protein concentrations. After pregnancy interruption, serum hCG levels decreased by 95% within 10 days, and TSH levels rose concomitantly from 0.80 to 1.48 mIU/L (P less than 0.001). In individual women serum hCG correlated negatively with TSH (r = 0.322; P = 0.005) and positively with free T3 (r = 0.388; P less than 0.001). These results suggest that hCG has thyrotropic activity, which, through rises in thyroid hormone levels, suppresses TSH secretion. In this regard, 27,000-128,000 IU hCG correspond to 1 mIU TSH. Pregnancy-induced changes in thyroid function, however, do not affect the circadian TSH rhythm.
Assuntos
Gonadotropina Coriônica/fisiologia , Ritmo Circadiano , Gravidez/fisiologia , Glândula Tireoide/fisiologia , Adulto , Gonadotropina Coriônica/sangue , Feminino , Humanos , Hidrocortisona/sangue , Gravidez/sangue , Glândula Tireoide/metabolismo , Hormônios Tireóideos/sangue , Tireotropina/sangueRESUMO
To elucidate the role of the testis in the control of LH and FSH secretion before puberty, we examined pulsatile LH and FSH secretion in six prepubertal boys with primary testicular failure (two boys with masculine pseudohermaphroditism, two boys with the Klinefelter's syndrome, and two boys with anorchia) and eight normal prepubertal boys. Plasma LH and FSH levels were measured every 15 min for 6 h during the day and night with ultrasensitive (0.019 and 0.014 IU/L) time-resolved immunofluorometric assays. In all six hypogonadal boys the mean FSH level was above the range of the normal prepubertal boys, whereas the LH level was elevated in only one boy. All boys had LH and FSH pulses. The FSH pulse interval in the anorchid boys was shorter than that in the normal boys, but this was not observed in the other hypogonadal boys. The LH pulse interval in the anorchid and other hypogonadal boys was the same as that in the normal boys. The FSH pulse amplitudes were higher in the anorchid and other hypogonadal boys than in the normal boys, but the LH pulse amplitudes were higher only in the anorchid boys. We conclude that in prepuberty the testes have little effect on LH secretion, but that they are involved in the regulation of FSH levels. In primary testicular failure, the elevation of FSH levels is associated with an increase in FSH pulse amplitude and, in the absence of testicular steroids, possibly also with an increase in FSH pulse frequency.
Assuntos
Hormônio Foliculoestimulante/sangue , Hipogonadismo/sangue , Hormônio Luteinizante/sangue , Puberdade , Testículo/fisiologia , Fatores Etários , Criança , Pré-Escolar , Fluorimunoensaio , Hormônio Foliculoestimulante/metabolismo , Humanos , Hormônio Luteinizante/metabolismo , Masculino , Periodicidade , Software , Testosterona/sangue , Fatores de TempoRESUMO
To establish the pubertal changes in gonadotropin secretion, 24-h secretory profiles of LH and FSH were studied in 10 healthy boys by ultrasensitive (sensitivity, 0.019 and 0.014 IU/L, respectively) time-resolved immunofluorometric assays 21 times. Five of the 10 boys were sampled on 2-6 occasions over a time interval of 0.95-6.4 yr. When sampled, 6 boys were prepubertal (testicular volume, less than 3 mL), 8 boys were early pubertal (testicular volume, 3-5 mL), and 7 boys were midpubertal (testicular volume, 10-25 mL). Plasma was taken every 20 min for 24 h. All boys had LH and FSH pulses. In prepuberty, the mean LH level was much lower than the mean FSH level, and neither showed significant diurnal variation. In early puberty, the mean LH level increased much more than that of FSH. For LH, the increase in mean levels was due to an increase in both pulse amplitude and frequency. During early and midpuberty, these changes were most marked at night, leading to the appearance of diurnal variation. For FSH, the mean levels increased progressively from prepuberty to midpuberty, with a slight increase in the mean pulse amplitude at the onset of puberty, whereas no change in pulse frequency was found. In contrast to LH, no diurnal variation was found for FSH at any of the pubertal stages. Thus, at the onset of puberty, gonadotropin secretion undergoes specific changes, which are different for LH and FSH, involving changes in pulse amplitudes and frequencies and development of diurnal variation for LH.
Assuntos
Envelhecimento/sangue , Hormônio Foliculoestimulante/sangue , Hormônio Luteinizante/sangue , Puberdade/sangue , Adolescente , Criança , Ritmo Circadiano , Humanos , Masculino , RadioimunoensaioRESUMO
Serum hCG levels were measured in apparently healthy nonpregnant women and men using a highly sensitive and specific time-resolved immunofluorometric assay. The sensitivity of the assay was 0.03 IU/L. The levels were low in women of fertile age (median, 0.05 IU/L) and in men less than 60 yr of age (median, 0.04 IU/L). In women the median level increased to 1.1 IU/L after the menopause (range, 0.17-4.8 IU/L), and a similar but smaller increase occurred in men after 60 yr of age (median, 0.20 IU/L; range, less than 0.03-2.3). Stimulation with GnRH caused a 2- to 3-fold increase in the hCG level in both men and women. Chronic treatment of postmenopausal women with a combination of estrogen and progestagen lowered their serum hCG levels to about 50% of the pretreatment values. The hCG in serum could be separated from LH by gel chromatography, and the hCG immunoreactivity measured by direct assay of serum corresponded to the immunoreactivity eluted in the hCG fractions after gel chromatography. Thus, the results were not due to cross-reaction with LH. We conclude that serum of nonpregnant women and men contains hCG-like material, whose production is modulated by GnRH and sex steroids.
Assuntos
Gonadotropina Coriônica/sangue , Hormônios Esteroides Gonadais/farmacologia , Hormônios Liberadores de Hormônios Hipofisários/farmacologia , Adulto , Idoso , Cromatografia em Gel , Estradiol/farmacologia , Feminino , Humanos , Imunoensaio , Hormônio Luteinizante/sangue , Masculino , Medroxiprogesterona/farmacologia , Menopausa/sangue , Pessoa de Meia-IdadeRESUMO
Ovarian hyperstimulation caused by a gonadotroph adenoma in premenopausal women has been described only twice before this report. A 28-yr-old woman presented with menstrual disturbances and pelvic pains that began after stopping the use of contraceptive pills. Transvaginal ultrasound revealed enlarged ovaries with multiple cysts. The patient had elevated serum estradiol (up to 2900 pmol/L; normal, 80-300 pmol/L in the follicular phase) and inhibin (6.4 kU/L; normal, 0.5-2.5 kU/L) levels. Serum LH was appropriately suppressed (0.6 IU/L), but serum FSH varied from 4.9-8.1 IU/L. Both gonadotropins as well as the free alpha-subunit showed a paradoxical response to the stimulus by TRH. A nuclear magnetic resonance study unraveled a pituitary tumor, 12-14 mm in diameter, extending up to the suprasellar cistern. After pituitary surgery, all hormone values normalized, and the patient resumed regular ovulatory cycles. In immunostaining, 20-30% of the cells of the tumor stained positively for FSHbeta. We conclude that a gonadotropin-producing adenoma must be considered in the differential diagnosis of a patient presenting with large multicystic ovaries and high estradiol levels in the absence of exogenous gonadotropins.
Assuntos
Adenoma/metabolismo , Hormônio Foliculoestimulante/metabolismo , Cistos Ovarianos/etiologia , Neoplasias Hipofisárias/metabolismo , Adenoma/complicações , Adenoma/cirurgia , Adulto , Anticoncepcionais Orais Hormonais/administração & dosagem , Estradiol/sangue , Etinilestradiol/administração & dosagem , Feminino , Hormônio Foliculoestimulante/sangue , Humanos , Inibinas/sangue , Hormônio Luteinizante/sangue , Distúrbios Menstruais/etiologia , Norpregnenos/administração & dosagem , Cistos Ovarianos/sangue , Cistos Ovarianos/diagnóstico por imagem , Neoplasias Hipofisárias/complicações , Neoplasias Hipofisárias/cirurgia , Tireotropina/sangue , Hormônio Liberador de Tireotropina , Tiroxina/sangue , UltrassonografiaRESUMO
Determinations of serum gonadotropin concentrations by ultra-sensitive methods have improved the diagnosis of pubertal disorders. The onset of puberty can be estimated by measuring serum gonadotropin pulsation, but as this requires serial nocturnal blood sampling, it is not a routine investigation. Gonadotropin measurements in first morning voided (FMV) urine samples could reflect the integrated nocturnal gonadotropin secretion and predict pubertal development earlier than daytime serum measurements. We studied the value of urinary LH (U-LH) measurements in FMV urine with reference to serum LH (S-LH) levels using an ultrasensitive time-resolved immunofluorometric assay in samples from 297 children and adolescents (145 boys and 152 girls, aged 5-15 yr) with known pubertal stages (Tanner 1-5). Stage 1 subjects (prepubertal) were divided into 5 age groups to assess whether there is an increase in LH before clinical signs of puberty can be detected. The correlation between FMV urine and S-LH values was good (r = 0.64; P < 0.0001). The 2 oldest groups of prepubertal subjects (11 and 12 yr) had significantly higher (P < 0.001) U-LH concentrations than the 3 younger groups. This difference was less marked for S-LH. A significant increase in FMV U-LH concentration occurs before the first clinical signs of puberty in a sex-independent fashion. Our data indicate that FMV U-LH measurement is a clinically relevant, noninvasive method for the evaluation of pubertal development, and it may be helpful in the investigation of pubertal disorders.
Assuntos
Ritmo Circadiano , Hormônio Luteinizante/urina , Puberdade/urina , Envelhecimento/sangue , Envelhecimento/urina , Criança , Pré-Escolar , Feminino , Humanos , Hormônio Luteinizante/sangue , Masculino , Puberdade/sangue , Caracteres SexuaisRESUMO
Earlier studies have demonstrated, that an extremely rapid and sensitive assay for human chorionic gonadotropin (hCG) can be obtained by utilizing time-resolved fluorometry in combination with an immunometric assay. In this assay 2 different monoclonal antibodies are used. One antibody, directed against the beta-subunit of hCG, is immobilized by coating onto a microtitre well whereas the other antibody is labelled with a europium chelate. This assay was compared with a similar assay, which differed only by using 125I instead of europium as a label. The results demonstrate, that the high sensitivity of the immunofluorometric assay was mainly the result of the high antibody concentration used since almost equal sensitivity was obtained using the same amounts of radiolabelled antibody. However, this implied the use of 10-100 times more radioactivity per tube than usually applied in immunoassays. For practical purposes the time-resolved immunofluorometric assay was superior because the measuring time was shorter and the reagents non-radioactive and stable.
Assuntos
Gonadotropina Coriônica/análise , Anticorpos Monoclonais , Európio , Imunofluorescência , Humanos , Radioisótopos do Iodo , Radioimunoensaio/métodosRESUMO
Quantitation of human chorionic gonadotropin (hCG) in maternal serum is widely used for screening of fetal trisomy-21 (Down's syndrome). When using immunometric assay, the sample usually has to be prediluted in order to quantitate the high hCG concentrations occurring during weeks 10-17 of pregnancy. Utilizing a streptavidin-biotin system we have developed an immunofluorometric assay (IFMA) for the quantitation of hCG in pregnancy serum that does not require predilution of the sample. The sample is added to a streptavidin coated microtiter strip well together with a biotinylated 'capture' antibody. About 1-2% of the capture antibody is biotinylated whereas most of the antibodies are unbiotinylated and thus unable to bind to the solid phase, while still binding antigen. This displaces the assay range from 0.5-5000 IU/l of the standard assay to 20-250,000 IU/l. The method shows good correlation with the standard procedure (r = 0.96). By eliminating a dilution step the assay procedure is both simplified and reinforced. The principle of this method should be readily applicable to other antigens and detection methods.
Assuntos
Gonadotropina Coriônica/sangue , Fluorimunoensaio/métodos , Proteínas de Bactérias , Biotina , Feminino , Humanos , Gravidez , Sensibilidade e Especificidade , EstreptavidinaRESUMO
Human chorionic gonadotropin (hCG), its subunits and fragments are widely used for diagnostic purposes. In addition to the diagnosis of pregnancy and pregnancy related disorders, hCG determinations are used for diagnosis of trophoblastic and recently also nontrophoblastic tumors. The use for diagnosis of nontrophoblastic tumors requires highly specific and ultrasensitive assays. With these, it is possible to measure the concentrations of both hCG, the free beta-subunits and the so called beta-core fragment in healthy subjects. Therefore it is important to establish reference values for these and also to be aware of the influence of physiological factors on the serum and urine concentrations. Improved standardization of the assay methods is also essential for these novel applications of hCG determinations to become useful.
Assuntos
Gonadotropina Coriônica/química , Gonadotropina Coriônica/análise , Gonadotropina Coriônica/fisiologia , Gonadotropina Coriônica/normas , Feminino , Humanos , Neoplasias/diagnóstico , Gravidez , Complicações na Gravidez/diagnóstico , Testes de GravidezRESUMO
Some pituitary hormones are expressed in leukocytes and are thought to play a role in the regulation of leukocyte function. We studied the expression of the mRNA for the beta-chains of luteinising hormone (LHbeta) and chorionic gonadotropin (CGbeta) and their translation into protein in various leukocyte subsets. Monocytes, granulocytes, B and T-cells from peripheral blood were separated. Lymphocytes were stimulated with various mitogens, prolactin and mixed lymphocyte culture. LHbeta and CGbeta mRNA expression was determined by reverse transcriptase polymerase chain reaction. LH, LHbeta, CG and CGbeta protein were determined in the culture medium by immunofluorometric assays. LHbeta mRNA expression was detected in all cell fractions and cultures and stimulation with prolactin induced LH protein in the culture medium. CGbeta mRNA expression appeared after culture of lymphocytes, but mitogens and prolactin had no clear stimulating effect. The LH expression in leukocytes shown here suggests an autocrine function of this hormone in blood cells.
Assuntos
Gonadotropina Coriônica Humana Subunidade beta/sangue , Gonadotropina Coriônica Humana Subunidade beta/genética , Leucócitos/metabolismo , Hormônio Luteinizante/sangue , Hormônio Luteinizante/genética , RNA Mensageiro/sangue , RNA Mensageiro/genética , Adulto , Sequência de Bases , Linhagem Celular , Gonadotropina Coriônica Humana Subunidade beta/metabolismo , Primers do DNA/genética , Feminino , Expressão Gênica , Humanos , Técnicas In Vitro , Hormônio Luteinizante/metabolismo , Masculino , RNA Mensageiro/metabolismoRESUMO
A novel time-resolved immunofluorometric method is described for the estimation of human luteinizing hormone (LH) in serum and urine. The method utilizes two monoclonal antibodies: one reacting with the beta-subunit is adsorbed to the wall of a microtiter well, and the other is labeled with a fluorescent europium chelate and reacts with the alpha-subunit. The method is ultrarapid (15-30 min) and highly sensitive (1 IU/L). A large linear measuring range allows measurement of LH levels from 1 to 250 IU/L. For the monitoring of urinary LH, IFMA gives the same information as the Hi-Gonavis assay, but it has the advantages of greater sensitivity and a shorter assay time. For the determination of serum LH levels, an acceptable correlation was observed between radioimmunoassay and IFMA. Furthermore, IFMA has a greater sensitivity, and is more rapid and not dependent on the handling of radioactive materials.
Assuntos
Hormônio Luteinizante/metabolismo , Anticorpos Monoclonais , Feminino , Humanos , Imunoensaio/métodos , Hormônio Luteinizante/sangue , Hormônio Luteinizante/urina , Microquímica , Espectrometria de Fluorescência/métodosRESUMO
The view that light affects the mammalian circadian clock only through the eyes was recently challenged by a study in which the phases of human circadian rhythms were shifted by extraocular light exposure. This finding has not been confirmed, however. We studied the effects of light exposure (3 h, broad spectrum fluorescent white light, 13000 lux) on abdomen and chest on the circadian rhythms of serum melatonin, cortisol and thyrotropin in six subjects. The protocol consisted of two 3-day sessions in a dimly lit (< 10 lux) experimental unit. In both sessions hourly serum samples were collected for hormone analysis on days 1 and 3. The skin light exposure was delivered on day 2 from 22.00 to 01.00h in one of the two sessions in a randomized order. In both sessions all three rhythms tended to delay, presumably due to the endogenous circadian cycle length being slightly longer than 24 h. However, the phase shifts did not differ significantly between the sessions. Thus, the present study does not support the existence of extraocular photic regulation of the circadian rhythms in humans.