Thymolphthalein Monophosphate Research Articles

Dose- and time-dependent effects of taxifolin on viability and mineralization markers of osteoblast-like cells

The current study evaluated the effects of taxifolin treatments on the viability of osteoblast-like cells, and on the expression of early mineralization markers, as part of the ongoing search for new endodontic materials able to induce periapical healing without causing cytotoxicity. Saos-2 osteoblast-like cells were exposed to different concentrations of taxifolin (5 and 10 µM), applied as pretreatments either for 24h and 72h, or continuously throughout the experimental protocol. Cell viability using the methylthiazole tetrazolium (MTT) assay, alkaline phosphatase activity using thymolphthalein monophosphate assays, deposition of mineralized nodules using alizarin red staining, and expression of ALP and COL-1 by qPCR were determined after 6 and 13 days of treatment. The data were analyzed statistically (p<0.05). Taxifolin was not cytotoxic in the concentrations tested. Pretreatments with taxifolin for 24h and 72h at 10 µM stimulated ALP activity, and increased mineralized nodule deposition by Saos-2 cells. Continuous treatment with taxifolin was not effective in stimulating ALP activity and mineralization. ALP and COL-1 gene expression increased with taxifolin pretreatments, since the highest mRNA levels were observed after 72h of pretreatment with taxifolin at 10 µM on day 13. In conclusion, taxifolin was cytocompatible, and induced mineralization markers when applied for short periods in osteoblast-like cell culture.

Biostimulatory effects of simvastatin on MDPC-23 odontoblast-like cells

The aim of this study was to evaluate the bioactivity and cytocompatibility of simvastatin (SV) applied to MDPC-23 odontoblast-like cells. For this purpose, MDPC-23 cells were seeded in 96-well plates and submitted to treatments with 0.01 or 0.1 μM of SV for 24 h, 72 h or continuously throughout the experimental protocol. The negative control group (NC) was maintained in DMEM. Cell viability (MTT), ALP activity (thymolphthalein monophosphate), and mineralized matrix deposition (alizarin red) were analyzed at several time points. The data were submitted to ANOVA and Tukey's test (α = 0.05). Although cell viability was observed in the groups treated with SV, these groups did not differ from the NC up to 7 days. There was a reduction in cell viability for the groups treated with 0.1 μM of SV for 72 h, and submitted to continuous mode after 14 days. A significant increase in ALP activity occurred in the group treated with 0.01 μM of SV for 24 h, compared with the NC; however, only the group treated with 0.1 μM of SV in continuous mode reduced the ALP activity, in comparison with the NC. After 14 days, only continuous treatment with 0.1 μM of SV did not differ from NC, whereas the other experimental groups showed increased mineralized matrix deposition. Thus, it was concluded that low concentrations of simvastatin were bioactive and cytocompatible when applied for short periods to cultured MDPC-23 odontoblast-like cells.

In Vitro Proliferation and Osteoblastic Phenotype Expression of Cells Derived From Human Vertebral Lamina and Iliac Crest

Osteoblastic cells derived from vertebral lamina and iliac crest were isolated and cultured under the same conditions (osteogenic medium, pH, temperature, and CO2 levels). To compare proliferation and expression of osteoblastic phenotype of cells derived from vertebral lamina and iliac grafting. Many factors play a role in the success of bone graft in spinal fusion including osteoblastic cell population. Two common sources of graft are vertebral lamina and iliac crest, however, differences in proliferation and osteoblastic phenotype expression between cells from these sites have not been investigated. Cells obtained from cancellous bone of both vertebral lamina and iliac crest were cultured and proliferation was evaluated by direct cell counting and viability detected by Trypan blue. Alkaline phosphatase (ALP) activity was evaluated by thymolphthalein release from thymolphthalein monophosphate and matrix mineralization by staining with alizarin red S. Gene expression of ALP, osteocalcin, runt-related transcription factor 2, Msh homeobox 2, bone morphogenetic protein 7, intercellular adhesion molecule 1 precursor, osteoprotegerin, and receptor activator of NF-kB ligand was analyzed by real-time PCR. All comparisons were donor-matched. Proliferation was greater at days 7 and 10 in cells from vertebral lamina compared with ones from iliac crest without difference in cell viability. ALP activity was higher in cells from vertebral lamina compared with cells from iliac crest at days 7 and 10. At 21 days, mineralized matrix was higher in cells derived from vertebral lamina than from iliac crest. At day 7, gene expression of ALP, osteocalcin, runt-related transcription factor 2, Msh homeobox 2, bone morphogenetic protein 7, intercellular adhesion molecule 1 precursor, receptor activator of NF-kB ligand, and osteoprotegerin was higher in cells derived from vertebral lamina compared with iliac crest. Cell proliferation and osteoblastic phenotype development in cells derived from cancellous bone were more exuberant in cultures of vertebral lamina than of iliac crest.

Pulsed Field Gradient ‘Inverse’ HOESY Applied to the Isotope Pairs1H,31P and1H,7Li*

For many heteronuclei, inverse (1H detected) heteronuclear Overhauser effect spectroscopy (HOESY) should be superior in theory over heteronucleus detected HOESY. In practice, strong t1 noise artifacts completely cancel the benefits of inverse HOESY. However, pulsed field gradient (PFG)-enhanced inverse HOESY yields 2D spectra without any t1 noise, consisting only of genuine cross peaks. The applicability of the method is demonstrated for thymolphthalein monophosphate hydrate using 31P,1H PFG-HOESY and for n-butyllithium using 7Li,1H PFG-HOESY. In addition to the absence of t1 noise, the merit of PFG-inverse HOESY is improved digital resolution in the usually crowded 1H domain.

Diagnostic Value of Prostatic Acid Phosphatase (Pap) and Prostate-Specific Antigen (Psa) in Patients with Prostatic Cancer

Prostatic acid phosphatase (PAP) and specific prostatic antigen (PSA) in the serum of patients with prostatic carcinoma were determined and their clinical value compared. 128 patients were examined, 60 (46.9%) of whom had prostatic carcinoma (4 in stage T2, 27 in T3 and 29 in T4) and 68 with benign prostatic pathology. ROC (receiver operating characteristic) curves were plotted from resulting data and the underlying areas calculated to evaluate the clinical accuracy of the two markers. The area for PSA (0.90 +/-0.03) was significantly greater than that for PAP (0.71 +/-0.05), showing that PSA was better at detecting patients with or without prostatic carcinoma. Maximum clinical accuracy was 0.883 obtained with discriminating values of 0.8 U/L for PAP and 10 ug/L for PSA, confirming the superiority of PSA. However, PAP determination using thymolphthalein monophosphate as the specific substrate for the prostatic isoenzyme, showed greater clinical specificity (98.5%) so that association of the two markers made it possible to eliminate false positive results. In conclusion, results suggest the possibility of using PSA for diagnosing prostatic carcinoma in a selected high risk population. However, the simplicity of the method used for determining PAP and the greater clinical specificity of PAP and PSA combined, suggest determining both parameters for diagnostic purposes.

Effects of Presumptive Test Reagents on the Ability to Obtain Restriction Fragment Length Polymorphism (RFLP) Patterns from Human Blood and Semen Stains

Some of the commonly used presumptive test reagents for identification of blood and semen could potentially affect the recovery of intact high-molecular-weight deoxyribonucleic acid (DNA) from evidentiary samples. Thus, the capability of performing restriction fragment length polymorphism (RFLP) analysis on evidentiary samples could be compromised. In order to investigate the potential effects of presumptive test reagents on the DNA present in these samples, bloodstains on cotton and glass were exposed directly to luminol, benzidine, phenolphthalein, o-tolidine, and leucomalachite green, while semen stains and vaginal swabs containing semen were exposed directly to bromochloroindolyl phosphate (BCIP) and sodium thymolphthalein monophosphate (STMP) reagents. The yield gels for DNA quality and quantity and RFLP results indicated that bloodstains exposed to luminol, benzidine dissolved in ethanol, and phenolphthalein, as well as semen stains and vaginal swabs exposed to BCIP and STMP yield RFLP patterns consistent with that of the uncontaminated control. Except for the phenolphthalein treatment, the quantity of extractable, high-molecular-weight DNA obtained was comparable with that of untreated stains. Therefore, evidentiary material purposely or inadvertently contaminated with these reagents can be successfully typed. However, stains exposed to benzidine dissolved in glacial acetic acid, leucomalachite green, and o-tolidine failed to yield high-molecular-weight DNA or to produce any RFLP patterns.

Diagnosis and staging of prostate cancer.

This paper reviews current concepts and recent advances in the diagnosis and staging of carcinoma of the prostate (CaP). Open perineal biopsy is seldom performed because of unnecessary morbidity and a substantial risk of impotence. Transurethral biopsy is inaccurate in detecting both early CaP and residual Cap following transurethral resection of the prostate. Transrectal needle biopsy is contraindicated because of a high rate of sepsis. Transperineal needle biopsy is accurate, has a low risk of complications, and can be performed in an out-patient setting. Transrectal needle aspiration of the prostate appears very accurate with even less morbidity and may replace core needle biopsy in the future. The staging of CaP includes measurement of prostatic acid phosphatase (PAP), radionuclide bone scan, and pelvic lymphadenectomy. PAP should be measured enzymatically using thymolphthalein monophosphate as the substrate. Patients with elevated serum levels of PAP (0.8 IU/L) generally have advanced disease and are not candidates for radical prostatectomy. Radioimmunoassay measurements of PAP have not proved useful because of low specificity. Prostate specific-antigen (PSA) levels are similar in CaP and BPH, and PSA does not appear useful in either the diagnosis or staging of CaP. PSA is, however, extremely sensitive in detecting residual disease after radical prostatectomy. Alkaline phosphate and other biochemical markers have not been reliable because of low specificity. Flow cytometry is a new technique that may be useful in selecting patients for radical prostatectomy. The value of transrectal ultrasound both in diagnosis and staging remains uncertain. Radionuclide bone scan is extremely accurate in detecting osseous metastasis with a false negative rate of two percent. Pelvic lymphadenectomy with frozen section diagnosis is highly accurate in detecting lymphnode metastases with a false negative rate of three point five percent.

Correlation of Clinical Stage, Serum Prostatic Acid Phosphatase and Preoperative Gleason Grade with Final Pathological Stage in 275 Patients with Clinically Localized Adenocarcinoma of the Prostate

The usefulness of clinical stage, serum prostatic acid phosphatase and preoperative Gleason grade in predicting final pathological stage in patients with adenocarcinoma of the prostate remains controversial. To determine the predictive value of these 3 preoperative variables we reviewed 275 patients with clinically localized disease who were treated between April 1982 and February 1986. All patients were examined preoperatively and subsequently were operated upon by 1 urologist. Serum prostatic acid phosphatase was determined in all patients by the Roy method using thymolphthalein monophosphate as the substrate. The Gleason grade of each prostatic biopsy specimen was determined preoperatively by 1 pathologist, who also examined the final pathological specimen with respect to capsular penetration, and seminal vesicle and pelvic lymph node involvement.Using logistic regression analysis with the likelihood ratio chi-square test, clinical stage and Gleason grade had a direct correlation with capsular penetration (p less than 0.0001 and less than 0.0001, respectively), seminal vesicle involvement (p less than 0.0001 and less than 0.0001, respectively) and positive lymph nodes (p less than 0.0001 and less than 0.0002, respectively). Within the normal range of values (0.0 to 0.8IU/1.) serum prostatic acid phosphatase correlated directly with capsular penetration (p less than 0.003) and seminal vesicle involvement (p less than 0.01) but not with lymph node involvement (p equals 0.08). Again with logistic regression analysis we determined that the best predictors of final pathological stage are not individual variables but models that use combinations of preoperative variables. The models generated are as follows: capsular penetration—serum prostatic acid phosphatase and Gleason grade (p less than 0.00001), seminal vesicle involvement—clinical stage and Gleason grade (p less than 0.00001), and lymph node involvement—clinical stage and Gleason grade (p less than 0.00001). With these models probability plots have been constructed so that the final pathological stage in patients with clinically localized prostatic cancer can be predicted preoperatively.

Adverse Implications of Acid Phosphatase Levels in the Upper Range of Normal

A retrospective review of 102 consecutive patients with surgically staged, clinically localized prostatic carcinoma was performed to determine the relationship between pre-treatment enzymatic acid phosphatase values and histopathological extent of the tumor. Of 96 patients with normal pre-treatment acid phosphatase titers (thymolphthalein monophosphate substrate) 77 (80 per cent) had values in the lower and 19 (20 per cent) had values in the upper half of the normal range. Of the latter 19 patients 16 (84 per cent) had histological evidence of extraprostatic tumor extension. Similarly, 5 of 6 patients (83 per cent) with elevated pre-treatment acid phosphatase titers had extraprostatic extension and 1 had a persistent postoperative acid phosphatase elevation that normalized with megestrol acetate therapy. Thus, 22 of 25 patients (88 per cent) with acid phosphatase values in or above the upper half of the normal range had either histological or clinical evidence of extracapsular tumor extension. By contrast, 41 of the 77 patients (53 per cent) with acid phosphatase titers in the lower half of the normal range had extracapsular extension.The predictive value for extraprostatic tumor extension of an acid phosphatase level in the upper half of the normal range was 84 per cent. Furthermore, in the 96 patients with normal acid phosphatase titers the incidence of extraprostatic tumor extension was significantly greater (p less than 0.01, chi-square) in those with values in the upper rather than the lower half of the normal range. Acid phosphatase titers in the upper half of the normal range were proportionately more common among patients with high grade and high clinical stage tumors. However, among patients with low grade and low stage tumors an acid phosphatase value in the upper half of the normal range was an independent variable that correlated with the presence of extracapsular tumor extension. These results confirm previously reported adverse prognostic implications of enzymatic acid phosphatase titers in or above the upper half of the normal range.

Method for prostatic acid phosphatase (PAP) with sodium thymolphthalein monophosphate (STMP) as substrate, as adapted to the Cobas Bio.

Journal Article Method for prostatic acid phosphatase (PAP) with sodium thymolphthalein monophosphate (STMP) as substrate, as adapted to the Cobas Bio. Get access R Bonora, R Bonora Search for other works by this author on: Oxford Academic Google Scholar M Panteghini M Panteghini Search for other works by this author on: Oxford Academic Google Scholar Clinical Chemistry, Volume 32, Issue 8, 1 August 1986, Pages 1589–1590, https://doi.org/10.1093/clinchem/32.8.1589a Published: 01 August 1986

Effects of substrate concentration on results of determination of prostatic acid phosphatase with thymolphthalein monophosphate.

The relation between concentration of thymolphthalein monophosphate substrate and catalytic activity was investigated for the determination of prostatic acid phosphatase. This study, an extension of previously reported work (Clin. Chem. 27: 1372, 1981), shows that lot-to-lot variation in purity of thymolphthalein monophosphate preparations is reflected in substrate-velocity curves. Plateau regions in these curves at 1.5-2.5 g/L result from the combined effects of (a) substrate concentrations that are an order of magnitude below Km and (b) a further decrease in available substrate caused by formation of substrate aggregates in the presence of serum. To simplify the identification of superior lots of thymolphthalein monophosphate, we give a mixed-substrate protocol for testing different lots.

Presumptive Screening Test for Seminal Acid Phosphatase Using Sodium Thymolphthalein Monophosphate

The use of sodium thymolphthalein monophosphate for presumptive seminal acid phosphatase testing is discussed. Recent evidence reveals that the dye (o-dianisidine) used in conjunction with alpha-naphthyl phosphate for seminal acid phosphatase testing is a carcinogenic hazard. Numerous tests for seminal acid phosphatase were conducted with swabs from sexual assault kits to compare both substrates. Results of tests using sodium thymolphthalein or alpha-naphthyl phosphate correlated exactly. Sodium thymolphthalein is an effective alternative for preliminary seminal fluid analysis because of its high degree of selectivity and stability, and because it eliminates the potential health hazard.

Radioimmunoassay for prostatic acid phosphatase helps discriminate patients with prostatic cancer.

We measured serum prostatic acid phosphatase in ostensibly normal controls and a selected patient population, using both a modified radioimmunoassay and an enzymic method with thymolphthalein monophosphate as substrate. The upper limit of normal for the radioimmunoassay was 2.2 micrograms/L; its sensitivity and specificity for prostatic cancer were 71 and 95%, respectively, vs 51 and 99% for the enzymic method. For both methods the correlation between clinical staging and values for acid phosphatase was poor. Our data suggest that adjunctive use of the radioimmunoassay may help further discriminate those patients requiring needle biopsy.

Prostatic Cancer, Acid Phosphatase, Creatine Kinase-Bb and Race: A Prospective Study

To examine the effectiveness of prostatic acid phosphatase and creatine kinase-BB determinations in detecting prostatic cancer serum from 594 men more than 40 years old was assayed for prostatic acid phosphatase with the thymolphthalein monophosphate substrate and a radioimmunoassay kit. Creatine kinase-BB levels also were measured with a radioimmunoassay kit.Patients with benign prostatic hyperplasia had higher prostatic acid phosphatase levels than normal controls. Accordingly, to avoid a high incidence of false positives in patients with benign prostatic hyperplasia the 92.5 percentile level of the patients with benign prostatic hyperplasia (3.9ng./ml.) was chosen as the upper limit of normal. With this critical value elevated prostatic acid phosphatase levels were observed in 6 per cent of the patients with clinical stage A disease, 8 per cent with stage B, 35 per cent with stage C and 68 per cent with stage D. The radioimmunoassay was no more effective than the enzymatic assay in detecting prostatic cancer.A correlation between prostatic acid phosphatase levels and patient race was observed, with 80 per cent of the black patients with extracapsular prostatic cancer having elevated prostatic acid phosphatase levels compared to 34 per cent of the white patients with similar stage disease.Creatine kinase-BB was elevated only in patients with advanced disease and was of little value in the diagnosis of prostatic cancer.

A discrete analyzer (ABA-100) method for prostatic acid phosphatase with thymolphthalein monophosphate as substrate.

A discrete analyzer (ABA-100) method for prostatic acid phosphatase with thymolphthalein monophosphate as substrate.

Comparison of radioimmunological and conventional acid phosphatase assays in the serum of prostatic cancer patients.

Measurements of serum prostatic acid phosphatase concentrations (PAP) by radioimmunoassay (RIA) were compared with the conventional measurements of serum acid phosphatase activities using p-nitrophenylphosphate (pNPP, tot.) and magnesium thymolphthalein monophosphate (TMP) as substrates and L(+)-tartrate (pNPP, tr.) as inhibitor in five prostatic cancer patients before therapy and in 13 during therapy. Elevated serum acid phosphatase activities were detected in 2, 2 and 3 of the 5 untreated patients when using pNPP (tot.), pNPP (tr.) and TMP enzyme assays, respectively. RIA for PAP detected elevated concentrations of the enzyme in 4 of these patients' sera. Three of the patients without metastases and one patient with suspected metastases had elevated concentrations of PAP by RIA. Serum acid phosphatase isoenzyme 2, which is mainly of prostatic origin, was separated chromatographically from serum samples with increased acid phosphatase activity. It represented 60--92% of the total activity, when TMP was used as substrate. Significant correlations (beta less than 0.001) were observed between all conventional enzyme activity measurements used and PAP by RIA within the whole patient group (n = 18), but no correlations existed within the patient group (p = 6) of high normal, or low abnormal serum PAP (2.7--6.6 micrograms/l). In addition, PAP measured by RIA better reflected the clinical state of the 13 patients under treatment than the conventional enzyme assays investigated.

Serum and bone marrow radioimmunoassay of acid phosphatase in prostatic cancer

Use of radioimmunoassay (RIA) for determinations of prostatic acid phosphatase has recently received considerable attention because of reported higher sensitivity and specificity than previous enzymatic assays. We have compared the sensitivity and specificity of a commercially available RIA to a highly specific enzymatic assay (thymolphthalein monophosphate) using 37 patients with prostatic cancer and 34 patients with surgically proved benign prostatic hyperplasia. Seventeen of the cancer patients and all 34 of the BPH patients were studied prospectively. We further evaluated specificity by performing the RIA on 25 specimens of bone marrow from patients with nonprostatic disease. Our results indicate the radioimmunoassay is not, at this time, an adequate screening tool, and we question its accuracy in staging patients anymore reliably than by enzymatic assay.

Spectrophotometric and liquid-chromatographic studies of thymolphthalein monophosphate. Specifications for high-quality substrate for the measurement of prostatic acid phosphatase activity.

Fourteen lots of thymolphthalein monophosphate (TMP), disodium salt, obtained from 10 commercial suppliers were compared spectrophotometrically at 445 and 595 nm, liquid-chromatographically with monitoring at 254 nm, and enzymically by measurements of activity of prostatic acid phosphatase in human serum. Eight lots were classified as "unacceptable," six as "acceptable." Spectrophotometric testing revealed four lots with excessive thymolphthalein and three lots with grossly deficient amounts of TMP. In general, the chromatographic results paralleled those obtained by spectrophotometry, and both results correlated well with enzymic activity. Changing water content in this hygroscopic salt was a major problem, which resulted in great uncertainty as to the formula weight and therefore as to the moles of TMP actually taken. From these studies, specifications for high-quality TMP were determined. The critical importance of simultaneous enzymic activity measurements in comparisons with other "acceptable" lots in defining an adequate TMP substrate is stressed. Use of these specifications for selecting TMP for acid phosphatase activity measurements should improve intra- and inter-laboratory analytical performance.

Will Prostatic Acid Phosphatase Determination by Radioimmunoassay Increase The Diagnosis of Early Prostatic Cancer?

The determination of serum prostatic acid phosphatase by radioimmunoassay has received considerable attention as a more sensitive and specific test than its measurement by the usual enzymatic assay. It has been suggested that the increased sensitivity of the radioimmunoassay will improve greatly our ability to diagnose prostatic cancer in the early operative stages.Our study compares prostatic acid phosphatase determination by the radioimmunoassay and enzymatic techniques in 56 untreated and treated patients with prostatic cancer. In addition, 22 patients with biopsy-proved benign prostatic hyperplasia also were studied. The radioimmunoassay used was based on antibodies raised against prostatic acid phosphatase highly purified by 2-stage, ion-specific chromatography and isoelectric focusing techniques. The enzymatic assay used thymolphthalein monophosphate as the substrate.The patients were staged on the basis of digital examination, technetium bone scan and pelvic lymphadenectomy. There were 4 patients with stage A, 12 with stage B, 13 with stage C and 27 with stage D disease. The radioimmunoassay showed an elevated prostatic acid phosphatase in 13 per cent of the patients with intracapsular disease (stages A and B) and in 50 per cent of the patients with extracapsular disease (stages C and D). This finding compares with an enzymatic prostatic acid phosphatase elevation in 13 per cent of the patients with intracapsular and 53 per cent of those with extracapsular disease. There were fewer falsely elevated prostatic acid phosphatase levels by radioimmunoassay (2) than by enzymatic assay (6) in the patients with benign prostatic hyperplasia.Our data show that the radioimmunoassay method provides a reliable and specific measurement of prostatic acid phosphatase. However, the radioimmunoassay measurement of prostatic acid phosphatase does not appear to be more effective as a screening aid in the diagnosis of early stages of prostatic cancer and in our study it was no more helpful than the enzymatic assay.

Bone Marrow Acid Phosphatase in Prostate Cancer: An Assessment by Immunoassay and Biochemical Methods

Comparisons of the bone marrow and serum acid phosphatase values obtained by counter-immunoelectrophoresis and the Roy biochemical test were made in 72 patients with and in 13 patients without prostatic cancer. The counter-immunoelectrophoresis test, when positive at more than 1 international unit per liter, showed only 4.4 per cent falsely positive results. The Roy biochemical test, which uses sodium thymolphthalein monophosphate as the substrate, had 65 per cent falsely positive bone marrow acid phosphatase levels.Conflicting reports regarding the value of bone marrow acid phosphatase determinations in patients with prostatic cancer result from the use of non-specific substrates in biochemical methods for measurement and from the trauma incidental to bone marrow aspiration, which releases many non-prostatic acid phosphatase enzymes. The use of immunoassay such as counter-immunoelectrophoresis minimizes this source of error.