Salivary bisphenol-A levels detected by ELISA after restoration with composite resin.

Bisphenol-A diglycidylether methacrylate (Bis-GMA), which is synthesized from bisphenol-A (BPA), a compound with exogenous endocrine disrupter action, is widely used as a dental material. During clinical filling with sealants and composite resins, the compounds are solidified by polymerization and then used. However, it has been noted that unpolymerized monomers may become dissolved in saliva. In this study using a competitive ELISA system, we investigated the changes in the BPA concentration in saliva after restoration with composite resins. Commercial composite resins from nine companies were tested. Mixed saliva was collected from 21 subjects. Based on the dynamics of salivary BPA detected by this ELISA system, we concluded that several tens to 100 ng/ml of BPA were contained in saliva after filling teeth with composite resin but that sufficient gargling can remove it from the oral cavity. Our data suggest that sufficient gargling after treatment is important for risk management.

X-ray crystallographic approach to the design of phenolic schiff base reagents for the mutual separation of lanthanoids.

A novel mutual selectivity of lanthanoids in the N,N'-bis(5-nitrosalicylidene)ethylenediamine (H2Nsalen)-KCl and N,N'-bis(5-nitrosalicylidene)-o-phenylenediamine (H2Nsaloph)-KCl extraction systems was evaluated by the X-ray analysis of similar model complexes for extracted species. Cerium(IV) complexes with N,N'-bis(5-chlorosalicylidene)ethylenediamine (H2Clsalen), N,N'-bis(salicylidene)-o-phenylenediamine (H2saloph) and N,N'-bis(3,5-dibromosalicylidene)-o-phenylenediamine (H2Br2saloph) were selected as the models. The result of the X-ray analysis suggested that [Ln(III)(Nsalen)2]- and [Ln(III)(Nsaloph)2]- are meridional type (two ligands are oriented perpendicular to each other) and sandwich type (two ligands are oriented parallel to each other), respectively. It was suggested that the selectivity of the meridional structure is superior to that of the sandwich structure in these extraction systems.

Selective extraction of gallium from aluminum and indium using tripod phenolic ligands.

Solvent extraction of trivalent group 13 metal cations such as aluminum, gallium and indium with tripod quadridentate phenolic ligand, tris(2-hydroxy-3,5-dimethylbenzyl)amine (H(3)tdmba), was investigated as fundamental study for their mutual separation. Gallium was extracted almost quantitatively as Ga(tdmba) (logK(ex)=-6.66+/-0.06 on using chloroform as extraction solvent), whereas aluminum and indium were hardly extracted due to steric hindrance on complexation of them with the ligand. The extracted Ga species was estimated as trigonal bipyramidal complex with one H(2)O molecule. Furthermore, extractability of Ga was increased by changing the ligand to more acidic tris(5-chloro-2-hydroxy-3-methylbenzyl)amine (H(3)tcmba) (logK(ex)=-6.18+/-0.18 on using dichloroethane as extraction solvent).

A new Lp(a) assay that is unaffected by apo(a) size polymorphism.

We developed sandwich ELISA methods in which anti-apo(a) kringle 4 type 5 through protease (K4 x 5-Pro) domain monoclonal antibody (clone: 203E2) is employed in each instance as the capture antibody and one of the three species of monoclonal antibody [Mab] (clones: 108B8, 202A9, 2B3) is used as the labeled antibody. Using serum containing apo(a) with 34 repeats of kringle 4 as the calibrator, a commercial kit using anti-Lp(a) polyclonal antibody (Pab) or anti-apo(a) Mab overestimated the Lp(a) concentration in samples containing apo(a) with more than 34 repeats of kringle 4 and underestimated the Lp(a) concentration in samples containing apo(a) with fewer than 34 repeats of kringle 4. Moreover, it was demonstrated that the ratios of commercial kit values to anti-apo(a) K4 x 5-Pro Mab-based method values increased as the size of apo(a) increased. The ratios of apo(a) K5 x Pro Mab-based method values to anti-apo(a) K4 x 5-Pro Mab-based method values, however, remained almost constant regardless of the size polymorphism. Thus, we suggest that apo(a) size heterogeneity can significantly affect Lp(a) measurement in the Lp(a) assay using anti-Lp(a) Pab. The novel Lp(a) assay method, using only anti-apo(a) K4 x 5-Pro Mab, is not subject to this phenomenon.

[A novel Lp(a) assay not affected by apo(a) size polymorphism].

Apo(a), the protein moiety of Lp(a), has size polymorphism that derives from the variable number of K4 type 2 repeats. We have developed three ELISA method with using monoclonal antibodies against the K4 type 2 repeats, the K4 type 5-protease domain, and the K5 protease domain of the apo(a) molecule. The commercial assay kits were correlated with the results from ELISA that utilized a monoclonal antibody against the K4 type 2 but indicated poor correlation with an ELISA method using anti Lp(a) K4 type 5-protease domain monoclonal antibody. Serum Lp(a) values were 1.8-fold overestimated by the commercial assay kits in samples with S4 type of the largest isoform, but our ELISA method in which the apo(a) K4 type 5-protease domain is used as labeled antibody and capture antibody was not affected by size polymorphism.

Serum soluble CD44 levels for monitoring disease states in acute leukemia and myelodysplastic syndromes.

To determine the clinical implications of soluble CD44 (sCD44) levels in hematologic neoplasias, we developed an enzyme-linked immunosorbent assay for sCD44 using two monoclonal antibodies to the standard 90 kDa form, and assessed the serum concentration of sCD44 in normal healthy volunteers, patients with acute leukemia, myelodysplastic syndromes (MDS), and those with chronic myeloid leukemia (CML). Compared to that in normal individuals (n=51; 145. 1 24.6 ng/ml), the serum sCD44 level was significantly elevated in patients with acute myeloid leukemia (AML; n=18; 331.9 99.0 ng/ml, P=0.0001), acute lymphoid leukemia (ALL; n=16; 551.3 427.8 ng/ml, P=0.0001) and CML (n=18; 262.0 97.5 ng/ml, P=0.0001). The sCD44 level was slightly elevated in patients with MDS (n=43; 173.8 54.9 ng/ml, P=0.0071). In patients with acute leukemia, serum sCD44 concentrations decreased significantly in response to treatment and reached nearly normal levels after complete remission (P=0.0005 in AML and P=0.0032 in ALL). The sCD44 levels in patients with MDS increased after they developed acute leukemia, whereas no significant difference in sCD44 levels was observed between the chronic and the blastic phases in patients with CML. Our results indicate that serum sCD44 levels may be a useful marker for monitoring response to treatment and disease progression, especially in acute leukemia.

Extraction behavior of divalent first row transition metal ions with N,N'-bis(2-hydroxyphenylmethyl)-N,N'-bis(2-pyridylmethyl)-1,2-ethanediamine and its derivatives.

For the fundamental study on the development of novel extractants, extraction behavior of divalent first row transition metals such as manganese, nickel, copper and zinc was studied with using N,N'-bis(2-hydroxyphenylmethyl)-N,N'-bis(2-pyridylmethyl)-1,2-ethanediamine (H(2)bbpen) and its derivatives as extractants. From the numerical analysis of the intricate extraction behavior of the metals, it was found that they were extracted into chloroform not only as uncharged chelate complexes but also as ion-pairs of charged complexes with a counter anion in aqueous solution. In other words, these ligands seemed to act not only as divalent hexadentate extractants but also as monovalent tri- or tetra dentate ligands, forming positively charged complexes. Furthermore, the order of the extractabilities between the metals did not coincide with the Irving-Williams series of stability because of the 'cavity size' in the ligands and the structural rigidity of them.

Ion-pair extraction system for the mutual separation of lanthanides using divalent quadridentate schiff bases.

For the selective extraction of trivalent lanthanides, the use of quadridentate divalent phenolic Schiff bases, such as N,N'-bis(5-nitrosalicilidene)ethylenediamine (H(2)Nsalen) and N,N'-bis(5-nitrosalicilidene)-o-phenylenediamine (H(2)Nsaloph), was investigated. Lanthanides made anionic 1:2 complexes with these ligands and could be extracted into nitrobenzene as ion-pairs with a suitable monovalent countercation in the aqueous phase, after which the ion-pair was dissociated almost perfectly. Although the order of the extractability of lanthanides between these ligands was H(2)Nsaloph > H(2)Nsalen, the mutual selectivity of them in the H(2)Nsalen system was higher than that in H(2)Nsaloph. Furthermore, with decreasing the size of the countercation, the mutual selectivity was enhanced although the extractability was lowered. A H(2)Nsalen-KCl extraction system, selected as an example, showed selectivity between lanthanides comparable with those of the better of other systems reported previously.

Formation of dinuclear copper(II) complex with N,N,N',N'-tetrakis(2-pyridylmethyl)-1,2-ethanediamine in aqueous solution.

The polynuclear complexation of divalent 3d transition metal cations with N,N,N',N'-tetrakis(2-pyridylmethyl)-1,2-ethanediamine (tpen) in aqueous solution was investigated. It was found that copper(II) forms a dinuclear complex with tpen in an aqueous solution containing chloride. The composition of the complex was determined as Cu(2)Cl(2)(tpen)(2+). Furthermore, the stability constant of the complex was determined and its structure was postulated to be (mu-Cl)(2).

[Time-resolved-fluoroimmunoassay for the determination of prostate-specific-antigen in capillary-blood spotted on filter paper and its clinical significance in prostate cancer patients].

Prostate-specific-antigen (PSA) has recently become widely recognized as an important tumor marker for prostate cancer. For relieving the examinee from pain and saving the time in blood collection, we developed a new method, based on time-resolved fluoroimmunoassay (TR-FIA), using a dried disc paper to collect capillary-blood. The standard curve ranged from 1.0 to 300 ng/ml, the intraassay (CV% was 4.09 to 6.36 (n = 10), while the interassay CV% was 3.45 to 8.87 (n = 2, k = 7). The diluted linearity and analytical recovery (91.5 to 98.2%) seemed to be satisfactory for routine use. All the samples from 30 healthy male adults had non-detectable serum PSA less than 1.0 ng/ml. When the cut-off value was set at 3.4 ng/ml (MEAN + SD of BPH), the positive rate in prostate cancer cases was 85.7% (12/14), in BPH cases was 20.0% (2/10) and in other benign urological disease was 0.0% (0/20). The clinical sensitivity was 85.7%, the specificity 93.3% and the efficacy 90.9%, accordingly. In conclusion, the newly developed TR-FIA method using dried disc paper to collect capillary-blood was quantitatively suitable as a routine test for prostate cancer, because of its simplicity in blood collection and less invasiveness to the examinee.

[A determination of anti-insulin receptor antibody in serum--a radioreceptor assay excluded the influence of insulin and anti-insulin antibody].

We studied an anti-insulin receptor antibody (IRAb) assay that excludes the influence of serum insulin and anti-insulin antibody in patients with anti-insulin antibody and normal subjects. The placental membranes strongly bound with 125I-insulin at 4 degrees C. The insulin specificity of the radioreceptor assay was confirmed by adding excess non-labeled insulin and other human hormones to the assay system. The strong correlation between the receptor binding reactivity (%) and the anti-insulin antibody levels was noted in the conventional direct IRAb assay (r = -0.95), but not in the present IRAb assay (r = 0.46) in 10 clinical samples. The placental membranes were stable as the target insulin receptor for IRAb assay at -80 degrees C for at least 4 months. A significant difference in IRAb levels was found between 10 patients with positive anti-insulin antibody and 20 normal controls (p less than 0.01, student t test). The coexistence of insulin and anti-insulin antibody were removed by absorbance to silicagel and by utilizing the two-step (indirect) assay, respectively. IRAb assay without the influence of serum cofactors showed excellent reproducibilities (CV 4.4% (N = 5) and 12.5% (N = 5) in within and between assay variations, respectively).

Correlation between X-prolyl dipeptidyl-aminopeptidase and serum amine oxidase in serum of patients with post-burn keloids.

Activities of X-prolyl dipeptidyl-aminopeptidase (EC 3.4.14.1) and amine oxidase (EC 1.4.3.6) in serum were assayed in two groups of patients, children two to nine years old and adults 23 to 60 years old, with hypertrophic scars after severe burn. The peptidase activity tended to be low initially for several months after the burn, but then returned to normal after six months. These changes were marked in the child group, less so in the adult group. Similar but less-pronounced changes were also observed in serum amino oxidase activity. The two serum enzyme activities showed a significant positive correlation (r = 0.668, p less than 0.001, n = 27) in the patients.