A radioimmunoassay for porcine intrauterine folate binding protein.
(65/8213)
A RIA was developed for porcine intrauterine folate binding protein (FBP). Displacement of [125I]FBP caused by increasing dilutions of uterine flushings collected from either d-15 pregnant or nonpregnant gilts or media from culture of endometrial tissue from d-15 pregnant or nonpregnant gilts was parallel to the displacement caused by the standard curve. Addition of known amounts of purified allantoic fluid FBP to dilutions of either intrauterine flushings or endometrial culture medium were measured accurately with the RIA. To test specificity, 2-mL samples of uterine flushings collected from d-15 pregnant and nonpregnant gilts were preincubated with 10 microCi of [3H]folic acid and then chromatographed using Sephadex G-100 (Sigma Chemical Co., St. Louis, MO). The fractions were subsequently assayed for radioactivity by liquid scintillation spectrophotometry and for FBP by RIA. The [3H]folic acid and FBP peaks coincided, indicating that the RIA is specific for FBP. Uterine flushings were collected on d 10, 11, 12, 13, 14, and 15 of the cycle or pregnancy from 1) White crossbred, 2) progesterone-treated White crossbred (200 mg of progesterone at 48 and 72 h after estrus), and 3) Meishan gilts and assayed for FBP. Total FBP increased 140-fold from d 10 to 15, and the pattern of change across day did not differ between pregnant and nonpregnant gilts. Progesterone treatment increased intrauterine FBP content on d 10 and 11. No difference in FBP concentrations was detected between White crossbred and Meishan gilts. These results indicate that the RIA for FBP is valid, allowing measurement of this protein in uterine flushings and endometrial culture medium. The onset of FBP secretion by the uterus between d 10 and 15 of the cycle or pregnancy is influenced by the timing of onset of progesterone influence in a manner similar to the endometrial proteins uteroferrin and retinol binding protein. In contrast to these endometrial proteins, FBP concentrations are similar in Meishan and White crossbred gilts. (+info)
Improved immunoradiometric assay for plasma renin.
(66/8213)
BACKGROUND: Our renin IRMA overestimated renin in plasmas with high prorenin-to-renin ratios. We suspected that the overestimation of renin was caused less by cross-reactivity of the renin-specific antibody with prorenin than by a conformational change of prorenin into an enzymatically active form during the assay. METHODS: Because the inactive form of prorenin converts slowly into an active form at low temperature, we raised the assay temperature from 22 degrees C to 37 degrees C, simultaneously shortening the incubation time from 24 to 6 h. The former IRMA was performed in <1 working day with these modifications. RESULTS: The comeasurement of prorenin as renin was eliminated. Reagents were stable at 37 degrees C, and the new and old IRMAs were comparable in terms of precision and accuracy. The functional lower limit of the assay (4 mU/L) was below the lower reference limit (9 mU/L). The modified IRMA agreed closely with the activities measured with an enzyme-kinetic assay. Results were not influenced by the plasma concentration of angiotensinogen. At normal angiotensinogen concentrations, the IRMA closely correlated with the classical enzyme-kinetic assay of plasma renin activity. CONCLUSION: The modified IRMA, performed at 37 degrees C, avoids interference by prorenin while retaining the desirable analytical characteristics of the older IRMA and requiring less time. (+info)
Endothelin concentrations in monochorionic twins with severe twin-twin transfusion syndrome.
(67/8213)
The objective of this study was to determine endothelin (ET-1) concentrations in monochorionic twin fetuses with and without twin-twin transfusion syndrome (TTTS). Fourteen monochorionic twin pregnancies complicated by TTTS and six without TTTS were studied. Matched maternal and fetal blood samples were obtained both in utero and at birth. Amniotic fluid samples were also collected from twin pairs. ET-1 concentrations were measured by radio-immunoassay. ET-1 concentrations in recipient fetuses were higher than in the donors both in utero(P < 0.001) and at birth (P < 0.01). Fetal concentrations of ET-1 in donors were similar to non-TTTS twins. Plasma ET-1 concentrations were significantly higher (P < 0.01) in recipient fetuses with severe hydrops than those with mild/no hydrops. Maternal concentrations of ET-1 were comparable in the two groups. Endothelin concentrations in recipient twins were 2(1/2) times higher than in their co-twins and this was related to the severity of hydrops. (+info)
Physical activity, body mass index, and prostaglandin E2 levels in rectal mucosa.
(68/8213)
BACKGROUND: Evidence suggests a relationship between prostaglandin levels in colonic mucosa and risk of colon cancer. Physical inactivity and a higher body mass index (BMI; weight in kilograms divided by [height in meters]2) have been consistently shown to increase risk of this cancer. We investigated whether higher levels of leisure-time physical activity or a lower BMI was associated with lower concentrations of prostaglandin E2 (PGE2) in rectal mucosa. METHODS: This study was conducted in 41 men and 22 women, 42-78 years of age, with a history of polyps, who participated in a randomized clinical trial testing the effects of piroxicam on rectal mucosal PGE2 levels. An [125I]PGE2 radioimmunoassay kit was used to determine PGE2 levels in samples of extracted rectal mucosa collected before randomization. Leisure-time physical activity was assessed through a self-administered questionnaire collected at baseline. The reported time spent at each activity per week was multiplied by its typical energy expenditure, expressed in metabolic equivalents (METs), to yield a MET-hours per week score. A repeated measures model was used to assess the effect of BMI and physical activity as predictors of PGE2 concentration. All statistical tests were two-sided. RESULTS: After adjustment for age, a higher BMI was associated with higher PGE2 levels (P = .001). A higher level of leisure-time physical activity was inversely associated with PGE2 concentration (P<.03). An increase in BMI from 24.2 to 28.8 kg/m2 was associated with a 27% increase in PGE2. An increase in activity level from 5.2 to 27.7 MET-hours per week was associated with a 28% decrease in PGE2. CONCLUSIONS: Physical activity and obesity may alter the risk of colon cancer through their effects on PGE2 synthesis. (+info)
Preparation and application of anti-idiotypic antibody against anti-gibberellin A4 antibody.
(69/8213)
A monoclonal anti-idiotypic antibody was raised against anti-gibberellin A4 (GA4) antibody, which recognizes biologically active gibberellins such as GA1 and GA4 specifically. Amino acid sequences of variable regions of both anti-GA4 and anti-idiotypic antibodies were analyzed. By using the property of the anti-idiotypic antibody to compete with GA1/4 in binding to the anti-GA4 antibody, we successfully applied the anti-idiotypic antibody to ELISA as a tracer for measuring GA1/4. The single-chain Fv (scFv) gene of the anti-idiotypic antibody was constructed, and scFv expressed in E. coli showed binding activity to anti-GA4 antibody. These results suggest the possible application of anti-idiotypic antibody as a handy and stable source of an enzymatic tracer for ELISA by production of fusion protein of the scFv and an appropriate enzyme. (+info)
Nail analysis for drugs of abuse: extraction and determination of cannabis in fingernails by RIA and GC-MS.
(70/8213)
Fingernail clippings were evaluated as analytical specimens for the detection and quantitation of cannabinoids. Specimens were obtained from consenting adults attending a drug clinic, along with information concerning the drugs which they had used over the previous six months. Methods for the surface decontamination and extraction of the specimens were evaluated. Detergent, water, and methanol washes followed by alkaline hydrolysis and liquid-liquid extraction were selected for use in the study. Extracts were analyzed by radioimmunoassay (RIA) and gas chromatography-mass spectrometry (GC-MS) to detect and quantitate cannabinoids present in fingernail clippings. Positive RIA results were obtained from specimens from six known cannabis users. The mean cannabinoid concentration in fingernail clippings determined by RIA was 1.03 ng/mg. Using GC-MS, the mean delta9-tetrahydrocannabinol concentration in fingernail clippings from a further 14 known cannabis users was 1.44 ng/mg. Using GC-MS, the average 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid concentration in fingernail clippings from three known cannabis users extracted in acidic pH was 19.85 ng/mg. Based on these results, fingernails are potentially useful biological specimens for the detection of past cannabis use in cases of medicolegal interest. (+info)
Radioimmunoassay of human cardiac acidic isoferritin: a new index for hepatic cancer.
(71/8213)
OBJECTIVE: To investigate human cardiac acidic isoferritin as a specific index of hepatic cancer. METHODS: Acidic isoferritin was isolated and purified from human heart muscle. A radioimmunoassay for the acidic isoferritin in human serum has been developed, on the equilibrium method. The antiserum was obtained from rabbits immunized with purified acidic isoferritin. The 125I-acidic isoferritin was prepared by the chloramine-T method. The data were processed using the automated smoothed spline function data processing program. RESULTS: The intra- and inter-assay CV of acidic isoferritin RIA were 1.65% and 9.71%, respectively, and the recovery rate was 102%. The antiserum provided a linear response from 7.0 to 369.6 micrograms/L with ED50 of 27.50 micrograms/L. The cross reactivity with AFP, CEA, lactoferrin and transferrin was negligible, and that with ferritin was 1.74%. The serum acidic isoferritin concentration showed a considerable variation in different sex and age groups. The serum acidic isoferritin was measured in liver diseases including hepatic cancer, hepatic cirrhosis and acute and chronic hepatitis. Its sensitivity for diagnosis of hepatic cancer was 73.05%, independent from the severity of hepatic injury. In 8 malignant tumors studied, acidic isoferritin appeared the most valuable in the diagnosis of hepatic cancer, with its positive, negative, false positive and false negative rates all being ideal. CONCLUSIONS: Acidic isoferritin may turn to be a rather specific index of hepatic cancer. Combination of monitoring both acidic isoferritin and AFP would raise the positive detection and specificity in the diagnosis of hepatic cancer. (+info)
In vivo activity of a mixture of two human monoclonal antibodies (anti-HBs) in a chronic hepatitis B virus carrier chimpanzee.
(72/8213)
A 35-year-old female hepatitis B virus carrier chimpanzee was infused with one dose of a mixture of human monoclonal antibodies 9H9 and 4-7B (antibodies against hepatitis B virus surface antigen; HBsAg). Blood samples were taken before and up to 3 weeks after infusion. HBsAg and antibodies against HBsAg (anti-HBs) were quantified by radioimmunoassay and enzyme immunoassay. Free anti-HBs was never detected. Thirty min after the start of the infusion the HBsAg level was minimal with maximum loading of the chimpanzee HBsAg with human immunoglobulin. HBsAg complexes could be dissociated by acid treatment. The HBsAg level was completely restored on day 7. Similar results were obtained for the preS1-containing particles that may represent the infectious viral particles in the chimpanzee serum. A mouse monoclonal anti-HBs (HBs.OT40) was found to compete with 9H9 in artificial immune complexes with the pre-treatment HBsAg from the chimpanzee. Used as a conjugate, HBs.OT40 yielded a maximum decrease in the signal in the 30 min sample compared to non-competing anti-HBs conjugates. This indicates binding of HBsAg with 9H9 in the circulation of the chimpanzee. Immune-complexed 4-7B could not be detected by its corresponding 4-7B peptide conjugate, probably due to its low concentration in the complexes. It is concluded that human monoclonal anti-HBs can effectively reduce the level of HBsAg in serum from this chronic carrier. Monoclonals 9H9 and 4-7B may complement each other due to their different mechanisms of inactivation, probably with higher efficiency than that monitored by our HBsAg screening assays. (+info)