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Biomedical subjects

E Ishikawa

Publications and source records attributed to E Ishikawa.

At least 145 records · Page 8Linked to original sources
PubMed1989

Novel and sensitive noncompetitive enzyme immunoassay for peptides.

A novel and sensitive noncompetitive enzyme immunoassay for peptides is described. Peptides were biotinylated using sulfosuccinimidyl-6-(biotinamido)hexanoate and were trapped onto anti-peptide IgG-coated polystyrene balls. After washing the polystyrene balls to eliminate other biotinylated substances, the biotinylated peptides were eluted with HC1 and were reacted with anti-peptide Fab'-peroxidase conjugate. The complex formed was trapped onto streptavidin-coated polystyrene balls. Peroxidase activity bound to the polystyrene balls was assayed by fluorimetry. The detection limit of angiotensin I as a model peptide was 13 fg (10 amol)/tube and 0.8 ng/l of plasma, which was 80 to 480-fold lower than those previously reported by competitive radioimmunoassay and competitive enzyme immunoassay. And other peptides could also be measured more sensitively by the present noncompetitive enzyme immunoassay method than by competitive immunoassays.

Angiotensin I↗
PubMed1989

Comparative study of plasminogen activators in cancers and normal mucosae of human urinary bladder.

We have developed a highly sensitive sandwich enzyme immunoassay for determination of urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA) antigen levels in extracts of human tissues. We determined antigen levels of PAs in extracts of 31 primary cancers and 15 normal mucosal tissues of the urinary bladder using this method. U-PA antigen levels in extracts of bladder cancers were significantly higher than those in normal tissues (p less than 0.005). U-PA antigen levels significantly increased as histological grading of malignancy advanced. There was no correlation between t-PA antigen level and malignancy. These results indicate that an increase of u-PA antigen level may be a parameter of malignant transformation and may play an important role in invasiveness of cancer.

Aged↗
PubMed1989

More sensitive and simpler immune complex transfer enzyme immunoassay for antithyroglobulin IgG in serum.

A more sensitive and simpler immune complex transfer enzyme immunoassay for antithyroglobulin IgG in serum and its use for the measurement of antithyroglobulin IgG in healthy subjects and patients with thyroid diseases are described. Antithyroglobulin IgG in test serum was reacted simultaneously with dinitrophenylthyroglobulin and thyroglobulin-beta-D-galactosidase conjugate. The complex formed of antithyroglobulin IgG, dinitrophenyl thyroglobulin, and thyroglobulin-beta-D-galactosidase conjugate was trapped onto two polystyrene balls coated with affinity-purified rabbit (antidinitrophenyl bovine serum albumin) IgG. The polystyrene balls were washed to eliminate nonspecific IgG in the test serum, and the complex was eluted from the polystyrene balls with dinitrophenyl-L-lysine and transferred to two polystyrene balls coated with affinity-purified rabbit (anti-human IgG gamma-chain) IgG. beta-D-Galactosidase activity bound to the polystyrene balls was assayed by fluorimetry using 4-methylumbelliferyl-beta-D-galactoside as substrate. As compared with the previous immune complex transfer enzyme immunoassay, the procedure was simplified by reducing one incubation step and one washing step, and the detection limit of antithyroglobulin IgG in serum (0.1 microgram/liter) was lowered 100-fold. More careful and extensive examination than in a previous study revealed the presence of antithyroglobulin IgG in a large proportion of healthy subjects and in all patients with Graves' disease and chronic thyroiditis.

Adolescent↗
PubMed1989

Development and applications of sensitive enzyme immunoassay for antibodies: a review.

The sensitivity of the conventional enzyme immunoassay methods is seriously limited by the presence of nonspecific immunoglobulins at high concentrations in test serum. In order to overcome this difficulty, various methods have been developed, and the detection limit of serum antibodies has been lowered to 50-100 ng/l, which is 300,000-fold lower than that by the conventional enzyme immunoassay method using antigen-coated solid phases and anti-immunoglobulin antibody-enzyme conjugates. As a result, anti-insulin antibodies have been demonstrated in most of patients treated with insulin for 0.5-10 months, and anti-thyroglobulin IgG has been demonstrated not only in all patients with Graves' disease and chronic thyroiditis, but also in a large proportion of healthy subjects.

Antibodies↗
PubMed1989

Measurement of anti-thyroglobulin IgG in serum by novel and sensitive immune complex transfer enzyme immunoassay.

Anti-thyroglobulin IgG in human serum was measured by a novel and sensitive immune complex transfer enzyme immunoassay. Anti-thyroglobulin IgG in human serum was reacted with dinitrophenyl thyroglobulin, and the complex formed between human anti-thyroglobulin IgG and dinitrophenyl thyroglobulin was trapped onto an affinity-purified rabbit (anti-dinitrophenyl bovine serum albumin) IgG-coated polystyrene ball. The polystyrene ball was washed to eliminate most nonspecific IgG in test serum, and the complex was eluted from the polystyrene ball, to which nonspecific IgG had been adsorbed, with dinitrophenyl-L-lysine and transferred to a clean polystyrene ball coated with rabbit anti-thyroglobulin IgG. The amount of human anti-thyroglobulin IgG in the complex on the rabbit anti-thyroglobulin IgG-coated polystyrene ball was estimated using rabbit (anti-human IgG gamma-chain) Fab'-horseradish peroxidase conjugate. The present enzyme immunoassay was 1,000 to 3,000-fold more sensitive than the conventional enzyme immunoassay, in which a human thyroglobulin-coated polystyrene ball was incubated with serum containing human anti-thyroglobulin IgG and, after washing, with rabbit (anti-human IgG gamma-chain) Fab'-horseradish peroxidase conjugate. By the present enzyme immunoassay, anti-thyroglobulin IgG was demonstrated in about 10% of healthy subjects and in all patients with Graves' disease and chronic thyroiditis. The principle of the present method may make it possible to sensitively measure other autoantibodies including anti-thyroid peroxidase and anti-thyrotropin receptor antibodies to aid diagnosis of thyroid diseases.

Adult↗
PubMed1989

Bioactivity of human growth hormone in urine from acromegalic patients.

Bioactivity of hGH in urine from five acromegalic patients was determined by Nb2 bioassay and IM-9 receptor modulation assay (RMA). Urine samples were concentrated by immunoadsorbent chromatography, dialysis and lyophilization. The ratio of the bioactivity by Im-9 RMA and the immunoactivity by RIA was between 0.81 and 1.24 (1.01 +/- 0.19, mean +/- SD). The ratio of the bioactivity by Nb2 bioassay and the immunoactivity ranged from 0.45 to 0.60 (1.37 +/- 0.85). Gel chromatography of hGH in urine samples measured by sensitive sandwich enzyme immunoassay showed that more than 97% of hGH activity was the 22K form. Urinary hGH from acromegalic patients was bioactive in Nb2 bioassay and IM-9 RMA and the bioactivity showed a close correlation with the immunoactivity. The major immunoactive form of hGH (22K) seems to correspond to the bioactivity.

Acromegaly↗
PubMed1989

The pathogenesis of graft-versus-host reaction in the intrahepatic bile duct. An immunohistochemical study.

Although intrahepatic bile duct injury following bone marrow transplantation is considered to be one feature of graft-versus-host disease, its developmental mechanism has not been clarified. In order to elucidate this aspect, an immunohistochemical study of the liver following human allogeneic bone marrow transplantation was made. Cytotoxic T lymphocytes (Tc) and natural killer cells (NK) were found in contact with intrahepatic bile duct epithelial cells showing degeneration and necrotic changes. These findings suggested a cytotoxic effect of these cells on bile duct epithelial cells. Abnormal expression of HLA class II (DR) antigen was recognized in intrahepatic bile duct epithelial cells following bone marrow transplantation. Cell injury was prominent in cells with weak DR antigen expression, whereas the cells demonstrating conspicuous expression appeared almost normal. There results suggest that abnormal expression of DR antigen plays an important role in the development of GVHD of the intrahepatic bile duct.

Bile Duct Diseases↗
PubMed1989

Comparative in vivo and in vitro studies on abnormal GH secretion in patients with acromegaly.

Eight patients with active acromegaly due to GH-producing pituitary adenoma were studied. GH secretory dynamics in vitro was evaluated by adding GRF, CRF, or a somatostatin analog, SMS 201-995 to the perifusate of dispersed cells from tumors. A comparison was made between the data obtained in preoperative tests for GH secretion and those obtained in experiments in vitro. Before operation, the GRF test (100 micrograms, iv) resulted in no GH response in three of six patients examined. The CRF test (100 micrograms, iv) resulted in a paradoxical GH increase in two of the same six patients. In vitro studies performed on adenoma cells revealed that exposure to GRF (100 ng/ml) elicited an increase in GH in seven of eight patients examined. Exposure to CRF (100 ng/ml) caused an enhanced GH secretion in four of the same eight patients. There were cases in which GH response to these hypothalamic hormones was observed in vitro but not in vivo, whereas there was only one case in which CRF caused an increase in GH in vivo but not in vitro. Thus, GH secretory dynamics was not always the same in vivo and in vitro. The discrepancy could be ascribed to the different secretory status of hypothalamic hormone (e.g., GRF or somatostatin) in vivo in each acromegalic patient.

Acromegaly↗
PubMed1989

Measurement of nocturnal urinary growth hormone values.

Nocturnal urinary growth hormone values were measured by a sensitive enzyme immunoassay in normal adults, patients with GH deficiency, patients with Turner's syndrome, normal but short children who had normal plasma GH responses to provocative tests, and patients with acromegaly. The mean nocturnal urinary GH values in patients with acromegaly were significantly greater than those in normal adults (1582.3 +/- 579.8 vs 53.5 +/- 8.6 pmol/mmol creatinine (+/- SEM); p less than 0.05). In the normal but short children and patients with Turner's syndrome, the mean nocturnal urinary GH values were 83.1 +/- 5.2 and 79.8 +/- 29.5 pmol/mmol creatinine, respectively. In patients with GH deficiency, the nocturnal urinary GH values were undetectable (less than 5.3 pmol/mmol creatinine) except in one patient where the value was 6.3 pmol/mmol creatinine. The nocturnal urinary GH values of the patients with GH deficiency were significantly lower than those of the other groups (p less than 0.05). In normal but short children, the nocturnal urinary GH values correlated significantly with mean plasma nocturnal GH concentrations (r = 0.76, p less than 0.001), and 24-hour urinary GH values (r = 0.84, p less than 0.001), respectively. In 4 patients with GH deficiency who had circulating anti-hGH antibody, the urinary GH values were also undetectable. These data indicate that nocturnal urinary GH value reflects endogenous GH secretion during collection time, and that measurement of the nocturnal urinary GH values is a useful method for screening of patients with GH deficiency and acromegaly.

Acromegaly↗
PubMed1989

A highly sensitive sandwich enzyme immunoassay of urinary growth hormone in children with short stature, Turner's syndrome, and simple obesity.

GH values were determined by a highly sensitive sandwich enzyme immunoassay in the 1st morning and/or 24-h accumulated urine samples in 94 children (short stature 70, including 14 with complete GH deficiency, 9 with partial GH deficiency, and 47 with GH-normal short stature; Turner's syndrome, 10, and simple obesity, 14). GH values were also determined in the 2nd to 4th urine samples taken on the same day together with the 1st morning urine in 5 of them. GH values in the 1st morning urine correlated significantly with those of the 24-h urine and with serum peak and mean GH values during nocturnal sleep as a physiological GH secretion test. The 2nd to 4th urines had lower GH concentrations than the 1st morning urine. The GH value of the 1st morning urine in complete GH deficiency was significantly lower than those in GH-normal short stature, partial GH deficiency and Turner's syndrome. However, no significant difference was detected in urinary GH values between complete GH deficiency and simple obesity. We conclude that 1st morning urinary GH estimation may be useful for differentiation of complete GH deficiency from other causes of short stature, but may be difficult for the distinction between complete GH-deficiency and obesity with normal GH secretory ability.

Adolescent↗
PubMed1989

[Serological classification of Bacteroides gingivalis and purification of group specific antigens].

Bacteroides gingivalis were serologically investigated by the immunodiffusion technique using autoclave-extracted antigens and antisera against whole cells. Fifteen strains of B. gingivalis used in this study, were classified in two distinct serological groups. Eleven strains include strain 381, ATCC 33277 were classified Group A and four strains include strain Su63 were classified in Group B. The group specific antigens from B. gingivalis strain 381 (Group A) and Su63 (Group B) were partially purified from autoclave extracts by 75% ethanol precipitation and anion exchange chromatography, and were identified as polysaccharides composed of glucose (82.3%, 23.7%), rhamnose (7.1%, 12.7%), galactose (4.9%, 18.2%), mannose (2.3%, 1.7%), glucosamine (0.4%, 28.0%) and galactosamine (3.0%, 15.3%).

Antigens, Bacterial↗
PubMed1989

A highly sensitive and specific sandwich enzyme immunoassay for water soluble A and B substances.

A highly sensitive and specific sandwich enzyme immunoassay for water soluble A and B substances is described. An affinity-purified rabbit anti-A IgG-coated polystyrene ball was incubated with water soluble A substance and subsequently with affinity-purified rabbit anti-A IgG-horseradish peroxidase conjugate. In addition, an affinity-purified goat anti-B IgG-coated polystyrene ball was incubated with water soluble B substance and subsequently with affinity-purified goat anti-B IgG-horseradish peroxidase conjugate. Peroxidase activity bound to the polystyrene balls was assayed by fluorimetry. The detection limits of artificial A and B antigens (synthetic A and B trisaccharides coupled to bovine serum albumin) were both 0.1 fmol/tube, which were 30- and 300-fold lower, respectively, than those by the hemagglutination inhibition method. In addition, A and B substances in secretor saliva could also be measured 3- to 10-fold and 100- to 1,000-fold more sensitively, respectively, by the present method than by the hemagglutination inhibition method.

ABO Blood-Group System↗
PubMed1988

Comparative study of plasminogen activator antigens in colonic carcinomas and adenomas.

The correlation between malignant transformation and increased PA synthesis or secretion has been examined in a variety of cell lines. To study the relationship between content and composition of PAs and colorectal neoplasms, we measured u-PA and t-PA antigen levels in normal mucosa, tubular adenoma, adenocarcinoma in adenoma, and adenocarcinoma, using a sensitive sandwich enzyme immunoassay. This assay was very sensitive and was not hindered by the presence of serine protease inhibitors. Both adenomas and carcinomas had higher u-PA antigen levels than normal mucosa. The u-PA antigen level of adenomas was lower than that of carcinomas. Antigen level of t-PA, however, was lower in both adenomas and carcinomas compared with that in normal mucosa, the values being lowest in carcinomas. Two cases of carcinoma in adenoma had PA contents similar to those of carcinomas. u-PA antigen level of adenomas with dysplastic epithelium was higher than that of adenomas without dysplastic epithelium. Therefore, the increase of u-PA content in adenomas could be a parameter of malignant changes in adenomas.

Adenoma↗
PubMed1988

Existence of anti-thyroglobulin IgG in healthy subjects.

An autoantibody, anti-thyroglobulin IgG, was detected in a large proportion of healthy subjects. Sera were collected from 232 healthy subjects aged 7-83 yr, who had no apparent symptoms with normal serum levels of thyroid-stimulating hormone, confirming the absence of Graves' disease and chronic thyroiditis. Anti-thyroglobulin IgG in serum was measured by a novel enzyme immunoassay, the principle of which has been shown to provide 3,000 to 10,000-fold higher sensitivity than the conventional methods. Anti-thyroglobulin IgG was demonstrated in 38% of the healthy subjects (15% of those aged 7-19 yr and 69% of those aged 20-39 yr), and the serum concentration of anti-thyroglobulin IgG was assessed to be 2 micrograms/l - 38 mg/l.

Adolescent↗
PubMed1988

Enzyme immunoassay for alpha-human atrial natriuretic polypeptide--direct measurement of plasma level.

A sandwich enzyme immunoassay for alpha-human atrial natriuretic polypeptide (alpha-hANP) was developed. Polystyrene balls were coated with monoclonal IgG1 specific for the N-terminal half of the ring structure of alpha-hANP, and rabbit Fab' specific for the C-terminal (17-28) of alpha-hANP was conjugated to horseradish peroxidase. The polystyrene ball was incubated with alpha-hANP standards or plasma and subsequently with the conjugate. Bound peroxidase activity was measured by fluorimetry. No addition of hANP-free plasma for the standard curve was required when less than 50 microliters of plasma sample was used. The detection limit of alpha-hANP was 30 fg (10 amol). Using 50 microliters of plasma, we could detect 0.6 ng (0.2 pmol)/l without extraction. The basal plasma alpha-hANP level of healthy men in a supine position after an overnight fast was 24.5 +/- 13.2 (SD) ng/l and tended to decrease (15.3 +/- 8.5 (SD) ng/l) after intravenous administration of furosemide (40 mg) and subsequent one hour walking. There was a highly significant correlation between the plasma alpha-hANP concentrations measured by this enzyme immunoassay and by radioimmunoassay (r = 0.92). This enzyme immunoassay for alpha-hANP allows measurement of plasma alpha-hANP without extraction even in the volume-contracted state.

Adult↗
PubMed1988

[Bioactivity and heterogeneity of human growth hormone in urine from acromegalic patients].

Bioactivity of hGH in urine from five acromegalic patients was determined by Nb2 rat lymphoma bioassay (Nb2BA) and IM-9 receptor modulation assay (RMA). One urine sample (case 1) was concentrated by dialysis and lyophilization. Bioactivity by IM-9 RMA showed close correlations with immunoactivity by RIA in serial dilutions (RMA/RIA; 0.66-1.77). The estimates of bioactivity by Nb2BA were higher in all diluted samples except undiluted one than those of immunoactivity. Four samples were concentrated by affinity chromatography, dialysis and lyophilization. The ratio of RMA and RIA estimates was between 0.81-1.24, and that of Nb2BA and RIA estimates was between 0.46-2.62. Heterogeneity of urinary hGH was analyzed by gelfiltration using TSK-G-3000SW column by high performance liquid chromatography (HPLC) and by sensitive sandwich enzyme immunoassay. In addition to the main peak of 22K molecular weight (more than 97%), very small peaks of 40-60K (big hGH) and 150K (big-big hGH) were detected in chromatographic profile. Urinary hGH from acromegalic patients was bioactive in IM-9 RMA and Nb2BA. The predominant form of urinary hGH from acromegalic patients was little hGH, but big and big-big hGH were detected by sensitive sandwich enzyme immunoassay.

Acromegaly↗
PubMed1988

A comparative pathological study of liver injury after different combination chemotherapies for leukemia.

A form of liver injury characterized by bile ductule-like transformation of hepatocytes, prominent hemosiderin deposition and fibrosis, was occasionally encountered in autopsy cases we examined during the period from 1973 to 1981. It was found that this liver injury was irreversible and intimately related to combination chemotherapy with the DCMP regimen (daunorubicin, cytosine arabinoside, 6MP and prednisolone) for acute non-lymphocytic leukemia (ANLL). Its correlation was reconfirmed by the fact that this liver injury disappeared after withdrawal of the DCMP regimen in 1981. After 1978, the regimen of combination chemotherapy for adult ANLL was partly changed, 6-thioguanine being utilized (DCTP) instead of the 6MP in the DCMP regimen. The hepatic injury occurring after the change in the regimen was different from that produced with DCMP, showing reversible intrahepatic cholestasis. These facts indicated that substitution for one drug in a combination chemotherapy regimen could cause a different type of hepatic change.

Adult↗