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Biomedical subjects

E Ishikawa

Publications and source records attributed to E Ishikawa.

At least 163 records · Page 9Linked to original sources
PubMed1988

Urinary growth hormone levels measured by ultrasensitive enzyme immunoassay in patients with renal insufficiency.

An ultrasensitive enzyme immunoassay was used to measure urinary GH levels in patients with renal insufficiency and normal subjects. Urinary GH excretion varied widely, but was significantly higher (P less than 0.01) in patients with renal insufficiency (median, 339; range, 2-17,000 ng/day) than in normal subjects (5.4; 1.2-15 ng/day). Urinary GH excretion correlated positively with urinary beta 2-microglobulin excretion (r = 0.79; P less than 0.001) and negatively with creatinine clearance (r = -0.83; P less than 0.001). Gel chromatography of urine from patients with renal insufficiency revealed a major peak of urinary GH corresponding to a mol wt of 22K, that of pituitary GH. These findings suggest that the kidneys play an important role in the catabolism of GH and that urinary GH may reflect, at least in part, renal function as well as hypothalamo-pituitary function.

Adolescent↗
PubMed1988

Urinary growth hormone (GH) measurements are useful for evaluating endogenous GH secretion.

Daily (24-h) urinary GH excretion was measured using a highly sensitive sandwich enzyme immunoassay in 10 normal adults, 6 patients with hypopituitarism, 25 normal but short children who had normal plasma GH responses (peak plasma GH level, greater than 10 micrograms/L) to provocative tests, and 8 patients with acromegaly. The mean urinary GH values in the normal adults, patients with acromegaly, and patients with hypopituitarism were 13.8 +/- 4.0 (+/- SE) and 431.1 +/- 149.1 ng/g creatinine (Cr) (1.56 +/- 0.45 and 48.77 +/- 16.87 ng/mmol Cr) and undetectable, respectively; these mean values were significantly different from each other. In the normal but short children the urinary values ranged from undetectable to 55.8 ng/g Cr (6.31 ng/mmol Cr). All of the normal but short children and 4 patients with hypopituitarism participated in a 24-h endogenous GH secretion study. The urinary GH values correlated significantly with the mean 24-h plasma GH concentrations as an index of endogenous GH secretion (r = 0.81; P less than 0.001) and plasma somatomedin-C levels (r = 0.67; P less than 0.001), respectively. In 6 patients with acromegaly whose plasma GH levels were constant throughout a 4-h period, the urinary GH values also significantly correlated with the mean plasma GH levels (r = 0.95; P less than 0.01). These data indicate that urinary GH measurements reflect endogenous GH secretion and that measurement of urinary GH excretion is a useful, simple, and practical method for evaluating endogenous GH secretion.

Acromegaly↗
PubMed1988

Sensitive sandwich enzyme immunoassay of human growth hormone (hGH) in serum and urine using monoclonal antibody-coated polystyrene balls.

A sensitive sandwich enzyme immunoassay for human growth hormone (hGH) using monoclonal antibody is described. A monoclonal anti-hGH IgG-coated polystyrene ball was incubated with hGH and subsequently with affinity-purified rabbit anti-hGH Fab'-horseradish peroxidase conjugate. Peroxidase activity bound to the polystyrene ball was assayed by fluorimetry using 3-(4-hydroxyphenyl) propionic acid as a substrate. The detection limits of hGH in serum and urine were 1.5 ng/l using 20 microliters of serum and 0.2 ng/l using 0.15 ml of urine, respectively. The specificity and assay precision were satisfactory. hGH levels in serum and urine determined by the present sandwich enzyme immunoassay using monoclonal anti-hGH IgG-coated polystyrene balls were well correlated to those determined by the previous sandwich enzyme immunoassay using rabbit anti-hGH IgG-coated polystyrene balls. Levels of hGH in urine collected as first morning voids from healthy subjects aged 19-28 yr were 6.4 +/- 3.2 (SD) ng/g creatinine. However, the present assay gave lower hGH levels than the previous assay. This was at least partly explained by the fact that hGH in urine was less efficiently bound to monoclonal anti-hGH IgG-polystyrene balls than standard hGH, while the binding of hGH in urine and standard hGH to rabbit anti-hGH IgG-coated polystyrene balls was equally efficient. In addition, gel filtration showed that 22K hGH, a major component, in urine was less efficiently bound to monoclonal anti-hGH IgG-coated polystyrene balls than standard 22K hGH. The nature of hGH in serum and urine remains to be investigated.

Adult↗
PubMed1988

[Experimental study of drug-induced acute renal failure. Recovery after renal impairment caused by the administration of low molecular weight dextran and kanamycin in water-deprived rats].

Various risk factors for the development of acute renal failure (ARF) have been pointed out clinically. Hypotension, excessive aminoglycoside exposure, and dehydration were identified as highly significant acute insults. Adenine nucleotide level in the renal tissue and morphological changes were investigated to clarify the recovery of renal tissue after gradual impairment caused by the administration of low molecular weight dextran (LMWD) and Kanamycin (KM) in water-deprived rats. Male SD rats, deprived of water for 2 days, received various dosages of LMWD (10, 15, 20, 30 ml/kg, i.p.) and KM (300 mg/kg, s.c.) concomitantly. The rats were killed on the 5th day and the level of adenine nucleotide in the renal tissue was determined by high pressure liquid chromatography (HPLC). The kidney was examined by electron microscopy. For comparison, LMWD and KM were administered to a second group of rats in tap water ad libitum, using the same dosages. The group dehydrated rats was more liable to have ARF. Electron microscopy disclosed changes in the kidney, indicating a disorder of the lysosome of the epithelial cells, not the mitochondria, of the proximal tubules. In this group, a decrease in Energy Charge value (EC = (ATP + 1/2ADP)/(ATP + ADP + AMP)) occurred later than both functional and morphological impairments. Both the EC values and the microscopic examination of the rats with the LMWD.KM-induced ARF showed that water deprivation greatly increased the potential for ARF. The degree of renal impairment depended on the dosage of LMWD. Both excessive aminoglycoside exposure and water deprivation were significant risk factors.

Acute Kidney Injury↗
PubMed1988

Radioimmunoassay of cholecystokinin in plasma.

A sensitive and specific radioimmunoassay for cholecystokinin (CCK) has been developed. Synthetic unsulfated carboxy-terminal fragment, CCK-8, was radioiodinated by the conventional Chloramine-T method. Antibodies were raised against sulfated CCK-8 covalently coupled to bovine thyroglobulin via 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide. By purification, highly immunoreactive 125I-labeled CCK-8 was obtained. The antiserum was highly avid, and plasma could be assayed directly. The detection limit of the assay was 5 pmol of sulfated CCK-8 per liter. The assay measured fragments CCK-8, CCK-33, and CCK-39 with equimolar potency. CCK-4, gastrin, and vasoactive intestinal polypeptide were not detected, even at higher concentrations. The concentration of CCK, as the sum of these CCK peptides, in plasma during fasting was low (10.5 +/- 2.1 pmol/L, mean +/- SEM) but still detectable in all normal subjects examined (range, 6.4-20.1 pmol/L). After ingestion of a test meal, CCK in plasma increased rapidly, peaking at 41.3 (SEM 5.7) pmol/L at 40 min and remaining high for 3 h after the meal. This supports the concept that CCK has important roles in digestion and absorption.

Adult↗
PubMed1987

A novel enzyme immunoassay of anti-insulin IgG in guinea pig serum.

A novel enzyme immunoassay of anti-insulin IgG in guinea pig serum is described. Guinea pig anti-insulin serum diluted with nonspecific guinea pig serum was incubated with dinitrophenyl biotinyl nonspecific rabbit IgG-insulin conjugate and a rabbit (anti-dinitrophenyl bovine serum albumin) IgG-coated polystyrene ball. After washing to eliminate nonspecific guinea pig IgG in the diluted serum, the polystyrene ball was incubated with dinitrophenyl-L-lysine to elute the complex of anti-insulin IgG and the conjugate. The eluate was incubated with an avidin-coated polystyrene ball. Finally, the amount of guinea pig anti-insulin IgG in the complex trapped onto the avidin-coated polystyrene ball was measured by incubation with rabbit (anti-guinea pig IgG) Fab'-peroxidase conjugate. This enzyme immunoassay was 10,000-fold more sensitive than the conventional enzyme immunoassay using insulin-coated polystyrene ball and rabbit (anti-guinea pig IgG) Fab'-peroxidase conjugate.

Animals↗
PubMed1987

A highly sensitive enzyme immunoassay of anti-insulin antibodies in human serum.

A highly sensitive enzyme immunoassay of anti-insulin antibodies in human serum is described. Serum samples were subjected to successive processes of incubation with insulin, dextran-charcoal treatment to remove free insulin, precipitation of insulin-anti-insulin antibodies by polyethylene glycol, acid-treatment of the precipitates to inactivate anti-insulin antibodies and measurement of insulin by sandwich enzyme immunoassay technique. The detection limit of anti-insulin IgG in human serum was 50 pg/assay or 450 ng/l of serum. This was 1,000- to 3,000-fold less than that obtained by a conventional enzyme immunoassay, in which an insulin-coated polystyrene ball was incubated with diluted serum and subsequently with (anti-human IgG gamma-chain) Fab'-horseradish peroxidase conjugate. By the present enzyme immunoassay, anti-insulin antibodies were demonstrated in most (89%) of serum samples from diabetic patients who had been treated with porcine insulin and porcine insulin plus bovine insulin for 0.6-10 mth, while only a small proportion (3%) of serum samples from the same patients was positive by the conventional enzyme immunoassay. Similar results were obtained with serum samples from diabetic patients who had been treated with human insulin for 0.5-8.2 mth. The present enzyme immunoassay may be useful for the measurement of antibodies not only for insulin but also other antigens which can be removed by dextran-charcoal treatment and are not precipitated by polyethylene glycol.

Adult↗
PubMed1987

Sandwich enzyme immunoassay for human interleukin-1 beta (hIL-1 beta) in urine.

A highly sensitive sandwich enzyme immunoassay for human interleukin-1 beta (hIL-1 beta) in urine is described. Rabbit anti-hIL-1 beta IgG-coated polystyrene balls were incubated with urine samples and subsequently with affinity-purified rabbit anti-hIL-1 beta Fab'-horseradish peroxidase conjugate. After washing, the peroxidase activity bound to the polystyrene ball was assayed by fluorimetry using 3-(4-hydroxyphenyl)propionic acid as a substrate. The detection limit of hIL-1 beta was 0.5 pg/tube (30 amol/tube) or 5 ng/1 of urine when 0.1 ml of urine sample was used. The molecular weight of hIL-1 beta in normal urine was shown to be 17,000 by gel filtration, and levels of urine hIL-1 beta in healthy subjects aged 19-87 yr were 0-146 ng/1 or 0-88 ng/g of creatinine.

Adult↗
PubMed1987

Clinical application of enzyme immunoassay in the analysis of citrullinemia.

We have developed a sensitive enzyme immunoassay (EIA) for the quantification of argininosuccinate synthetase (ASS) in the range of 0.05-1 ng/tube using a Fab'-peroxidase conjugate prepared with maleimide compound devised by Ishikawa et al. Amounts of hepatic ASS in control subjects were determined by this method with the purified enzyme as a standard and their specific activities were calculated to be 1.3-1.4 mumol product/min (U)/mg of ASS, which was quite similar to that of the purified enzyme. Amounts of ASS in the liver of patients with three types of citrullinemia, type I, II and III according to our arbitrary classification, were determined by the EIA method. The following results were obtained: hepatic ASS from a patient diagnosed as type I citrullinemia because of its abnormally large Km values for citrulline and aspartate was calculated to have a specific activity of 0.18 U/mg of ASS, indicating that the enzyme is a variant; specific activities of ASS in the hepatic extract of type II citrullinemic patients were 1.2-1.3 U/mg of ASS, confirming the finding obtained by single radial immunodiffusion method that the decrease of the activity in type II is caused by a decrease in the total amount of ASS protein; a very small amount of immune cross-reactive material was detected in the liver of a patient diagnosed as type III citrullinemia based on findings of undetectable ASS activity in the liver and cultured skin fibroblast.

Adult↗
PubMed1987

In vitro production of human interleukin 1 alpha and interleukin 1 beta by peripheral blood mononuclear cells examined by sensitive sandwich enzyme immunoassay.

The in vitro production of human interleukin 1 alpha (hIL 1 alpha) and interleukin 1 beta (hIL 1 beta) by peripheral blood mononuclear cells was examined by sensitive sandwich enzyme immunoassays which could discriminate hIL 1 alpha and hIL 1 beta without cross-reaction with human IL2. In culture supernatants of mononuclear cells, two components were detected by sandwich enzyme immunoassay for hIL 1 alpha or hIL 1 beta. The molecular weight of one component was shown to be equal to that of recombinant hIL 1 alpha or hIL 1 beta by gel filtration. The elution volume of the other component corresponded to a molecular weight of about 30,000. The sum of the two components for both hIL 1 alpha and hIL 1 beta in culture supernatants of peripheral blood mononuclear cells from healthy subjects increased 1.7 to 38-fold by Escherichia coli lipopolysaccharide. The sum of the two components for hIL 1 beta was 13 to 97-fold larger than that for hIL 1 alpha.

Adult↗
PubMed1987

Development and clinical application of sensitive enzyme immunoassay for macromolecular antigens--a review.

Radioimmunoassay has been a powerful tool to measure haptens and antigens which are important for the investigation and diagnosis of diseases, especially endocrine disorders. However, the use of radioisotopes in radioimmunoassay suffers from serious disadvantages. Radioisotope-labeled reagents are unstable and hazardous to health. The disposal of radioactive wastes is not easy. Furthermore, the sensitivity of radioimmunoassay is limited by the detection limit of radioisotope that depends upon the half-life. The detection limit of the most widely used radioisotope, 125I, with a half-life of 60 days is 5 to 10 amol, when it is carrier-free. By contrast, the use of enzymes has obvious advantages. Some enzymes are very stable and cause no health hazards or waste disposal problems, provided that appropriate substrates are chosen. The detection limits of some enzymes are lower than that of 125I and will be further improved in the future. Therefore, enzyme immunoassay is potentially more sensitive than radioimmunoassay. This article reviews the development and clinical application of sensitive enzyme immunoassay for macromolecular antigens, which has been replacing radioimmunoassay.

Antigens↗
PubMed1987

A highly specific and sensitive sandwich enzyme immunoassay for human hemoglobin A.

A highly specific and sensitive sandwich enzyme immunoassay for human hemoglobin A (Hb A) is described. A rabbit anti-human Hb A IgG-coated polystyrene ball was incubated with human Hb A and subsequently with affinity-purified anti-human Hb A Fab'-horseradish peroxidase conjugates, which had been prepared before and after absorption with Japanese monkey Hb-Sepharose 4B and dog Hb-Sepharose 4B. Bound peroxidase activity was measured by fluorimetry using 3-(4-hydroxyphenyl)propionic acid as a substrate. The detection limits of human Hb A using the conjugate before and after the absorption were 0.65 pg/tube (3 X 10(10)-fold dilution of whole blood) and 2.0 pg/tube (1 X 10(10)-fold dilution of whole blood), respectively. Human Hb A could be discriminated from Hb of animals such as Japanese monkey, dog, cat, pig, horse, sheep, chicken and cow by measuring bound peroxidase activity in the presence and absence of the conjugates prepared before and after the absorption. Human Hb A in bloodstains on cotton gauze could be discriminated from Hb of animals described above even after seven washings. Human Hb A in 220,000-fold diluted bloodstains on cotton gauze could also be discriminated from Hb of animals described above.

Animals↗
PubMed1987

Clinical usefulness of a highly sensitive enzyme-immunoassay of TSH.

Using a recently developed sensitive enzyme immunoassay (EIA) for TSH, serum TSH levels in normal subjects and patients with various thyroid disorders were measured. The minimum detectable concentration was less than 0.005 mU/l. An excellent correlation was observed between TSH values measured by EIA and by sensitive radioimmunoassay (r = 0.932). In 26 normal subjects, serum TSH ranged from 0.30 to 2.65 mU/l (geometric mean 0.97 mU/l). In 27 untreated hyperthyroid Graves' patients, serum TSH was detected in all but one, ranging from 0.005 to 0.19 mU/l (geometric mean 0.040). These values were proved to be specific for human TSH by absorption, recovery, cross-reactivity and dilution studies; non-specific serum effects were also ruled out. In 10 patients with destructive thyroiditis, similar TSH values (0.005-0.20 mU/l, mean 0.032) were observed. However, no significant correlations between TSH and circulating thyroid hormone levels were observed in these thyrotoxic conditions. Furthermore, 9 untreated Graves' patients did not respond to a single i.v. injection of TRH. In 13 hypothyroid patients with hypothalamo-pituitary disorders, 10 had basal TSH levels of less than 0.4 mU/l, and TRH tests in five gave peak TSH values of more than 0.8 mU/l, which were associated with significant increases in serum T3. In three patients with Sheehan's syndrome, elevated basal TSH levels were observed. Two of them responded to TRH, but these TSH elevations were not associated with T3 increases. In conclusion, this sensitive EIA could measure TSH levels in most patients with thyrotoxicosis and central hypothyroidism. However, the true significance of these measured values needs to be elucidated by future studies.

Adolescent↗