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G Valet

Publications and source records attributed to G Valet.

At least 37 records · Page 2Linked to original sources
PubMed1992

[Flow cytometric analysis in non-small-cell bronchial carcinoma and its prognostic significance].

Tumor and lymph node infiltration, and the DNA-ploidy status of a tumor contain prognostic information in addition to the information obtained by histological examination of surgical samples. Specimens from 112 patients with non-small-cell lung carcinoma obtained immediately after surgery were investigated by means of flow cytometry. DNA-aneuploidy was found in 43% of the primary tumors. Independent from tumor stage, patients with DNA-euploid tumors lived significantly longer (p less than 0.01) than with DNA-aneuploid carcinomas. In 29 cases the DNA-ploidy status of the primary tumor (PTU) could be compared with that of the N2 lymph node metastases (LM). 7 samples revealed a change from DNA aneuploidy in the PTU to DNA-euploidy in the LM. Patients with DNA-euploid PTU and DNA-euploid LM lived significantly longer than patients with DNA-aneuploid PTU/DNA-euploid LM, and patients with DNA-aneuploid PTU/DNA-aneuploid LM. In case of lymph node infiltration only the simultaneous measurement of DNA ploidy of PTU and LM offers an accurate prognostic evaluation. Local tumor recurrence exhibited stability of DNA ploidy, showing DNA euploidy in 12 out of 13 PTU and their corresponding recurrent tumor. Thus, the DNA-ploidy status offers additional prognostic informations which is useful for an extended tumor classification.

Carcinoma, Non-Small-Cell Lung↗
PubMed1992

Prognostic value of deoxyribonucleic acid aneuploidy in primary non-small-cell lung carcinomas and their metastases.

The ploidy status of the deoxyribonucleic acid of a malignant lung tumor provides additional information besides histologic grading and tumor staging according to lymph node infiltration and tumor metastasis. Ninety-nine surgical specimens from patients with non-small-cell lung carcinoma were investigated by flow cytometry. Deoxyribonucleic acid aneuploidy was found in 48% of the primary tumors. Patients with deoxyribonucleic acid-euploid tumors showed better survival (p < 0.01) than those with deoxyribonucleic acid-aneuploid carcinomas independent of tumor stage. Deoxyribonucleic acid ploidy status of the primary tumor was compared with that of N2 lymph node metastases in 29 cases. Seven samples showed a change from deoxyribonucleic acid aneuploidy in the primary tumor to deoxyribonucleic acid euploidy in the lymph node metastases. Survival was significantly better for patients with euploid primary tumors and lymph node metastases, followed by patients with deoxyribonucleic acid-aneuploid primary tumors and euploid lymph node metastases. Survival was poorest in patients with deoxyribonucleic acid-aneuploid primary tumors and lymph node metastases. It was observed that only the simultaneous determination of deoxyribonucleic acid ploidy of primary tumors and lymph node metastases permits accurate prognostic evaluation in case of lymph node infiltration.

Actuarial Analysis↗
PubMed1991

Inhibition of the oxidative burst response of N-formyl peptide-stimulated neutrophils by serum amyloid-A protein.

Strong binding of the acute phase protein serum amyloid-A (SAA) to human neutrophils was found using flow cytometry. This binding was shown to be functionally relevant with respect to the oxidative burst reaction assayed on N-formyl peptide-stimulated neutrophils by the intracellular oxidation of non-fluorescent dihydrorhodamine to fluorescent rhodamine 123. The results show reduction of the oxidative burst response by isolated SAA (and recombinant SAA2). Inhibition was also demonstrated by acute phase as compared to normal human serum. This inhibitory effect was abolished by the purified monoclonal anti-amyloid A antibody mc29, strongly suggesting that SAA counteracts neutrophil responses to cytokines or bacterial products. This newly recognized function of SAA may help to prevent oxidative tissue destruction.

Adult↗
PubMed1991

Analysis of unknown DNA sequences by polymerase chain reaction (PCR) using a single specific primer and a standardized adaptor.

A new procedure for the PCR amplification of unknown DNA sequences adjacent to a known sequence is described. The required but not readily available second primer sequence in the unknown DNA sequence is obtained by creating an overhanging restriction site in the unknown sequence to which a double-stranded oligonucleotide adaptor of known sequence is ligated.

Base Sequence↗
PubMed1991

Respiratory burst activity in brain macrophages: a flow cytometric study on cultured rat microglia.

A new flow cytometric method for the investigation of the respiratory burst of macrophages/microglia isolated from neonatal rat brain has been established. Respiratory burst activity was measured quantitatively in single viable cells by the intracellular oxidation of non-fluorescent dihydrorhodamine 123 (DHR) to fluorescent rhodamine 123. Cultured microglia exhibited high spontaneous respiratory burst activity already before stimulation. After maximal stimulation with phorbol myristate acetate, DHR oxidation rose by 40-95%. The respiratory burst activity in resident or inflammatory, i.e. thioglycolate elicited, peritoneal macrophages was significantly lower than in cultured brain macrophages suggesting a high potential of microglia for oxidative tissue destruction.

Animals↗
PubMed1990

Intracellular pH, esterase activity, and DNA measurements of human lung carcinomas by flow cytometry.

An important intention of flow cytometric investigations is to obtain biochemical and biophysical information about cells which is suitable for automated tumor diagnosis. In this study, the ploidy status, the intracellular pH value, the intracellular esterase activity, and the cell volume of vital cells and the DNA and cell volume of dead cells were measured in cancerous tissue and normal lung tissue of 30 patients by flow cytometry. The cell samples were simultaneously stained with the pH and esterase indicator dye 1.4-diacetoxy-2,3-dicyanobenzene (ADB) and propidium iodide (PI). The flow cytometric measurements were performed in three-parameter list mode. The data were evaluated on an AT-compatible personal computer with the DIAGNOS1 program system for automated diagnosis of flow cytometric list mode data. Significant differences were found between normal and malignant tissue in DNA ploidy, in the intracellular esterase activity, in the cell, volume and in the percentage of inflammatory cells and parameters of necrosis. DNA-aneuploidy was observed in 38% of the lung carcinomas. The simultaneous detection of DNA-aneuploidy and tumor-associated properties in a multifactorial analysis led to correct automatic tumor diagnosis in 85% of cases. DNA-aneuploidy was found at a significant higher frequency in advanced tumors. Adenocarcinomas displayed DNA-aneuploidy more often (80%) than squamous cell carcinomas (33%).

Adenocarcinoma↗
PubMed1990

Multiple end labeling of oligonucleotides with terbium chelate-substituted psoralen for time-resolved fluorescence detection.

A new procedure for the photochemical functionalization and the subsequent nonradioactive labeling of synthetic oligonucleotides with psoralen derivatives was developed where a double-stranded poly(A-T) tail is attached to the 5'- or 3'-end of the oligonucleotide to be labeled. The double-stranded poly(A-T) tail is covalently crosslinked by psoralen molecules which carry reactive thiol or amino groups for the attachment of labels. A NH2-specific terbium chelate exhibiting long-lived fluorescence was attached to the functional groups of the intercalated psoralen molecules. Oligonucleotides substituted in this way hybridize readily and can be sensitively detected by time-resolved fluorescence measurements.

Base Sequence↗
PubMed1990

Flow cytometric parameters of neutrophil function as early indicators of sepsis- or trauma-related pulmonary or cardiovascular organ failure.

Flow cytometric parameters of neutrophil function, such as phagocytosis and degradation of Escherichia coli, intracellular pH value, esterase activity, and cell volume, were evaluated as risk indicators for sepsis- and trauma-related pulmonary and cardiovascular organ failure in intensive care patients. Serial blood samples (n = 201) were obtained from 47 prospectively identified patients. Each patient's condition was classified daily within four categories: post-traumatic (n = 22) or septic (n = 28) organ failure, transition state (n = 119), and stable organ function after recovery (n = 27). Thirty-two parameters of neutrophil function were automatically calculated for each blood sample from several flow cytometric list mode measurements of cell samples vitally stained with acridine orange for intact and denatured DNA or with 1,4-diacetoxy-2,3-dicyanobenzene for intracellular pH and esterase activity. The DNA of dead cells was simultaneously counterstained with propidium iodide. The cell biochemical parameter pattern was significantly different among samples of patients from the four clinical categories (p less than 0.05). Hyperergic phagocytosis was observed after trauma, in contrast to hypoergic phagocytosis, increased neutrophil cell volume, and elevated intracellular pH during sepsis. The clinical categories were correctly identified in 82% of the samples by automated classification with the DIAGNOS1/SPSS program system from the flow cytometrically determined cell functions. The course of the disease was correctly predicted 3 days in advance to the clinical manifestation of pulmonary or cardiovascular organ failure in 92% of the samples. The multifunctional analysis of neutrophils by flow cytometry seems of interest for early medical intervention in preseptic and preshock patients.

Bacterial Infections↗
PubMed1990

Flow cytometric analysis of respiratory burst activity in phagocytes with hydroethidine and 2',7'-dichlorofluorescin.

Hydroethidine (HE) and 2',7'-dichlorofluorescin (DCFH) were used for the flow cytometric measurement of reactive oxygen metabolites in leukocytes. Hydroethidine and DCFH were both rapidly oxidized in a cell-free cuvette assay to ethidium bromide (EB) and 2',7'-dichlorofluorescein (DCF) by H2O2 and peroxidase, but not by H2O2 alone, while only HE was oxidized by KO2, a source of O2-. Quiescent lymphocytes, monocytes, and neutrophils spontaneously oxidized HE to EB, while DCFH was only oxidized to a low degree. Neutrophils increased 6.9-fold in EB red fluorescence and 12.5-fold in DCF green fluorescence during the respiratory burst induced by phorbol 12-myristate 13-acetate or 6.1-fold and 4.7-fold, respectively, during the respiratory burst induced by Escherichia coli bacteria. The HE or DCFH oxidation during the respiratory burst, unlike the spontaneous HE oxidation, was not inhibitable by 10 mM NaNe indicating a non-mitochondrial source of cellular oxidants during the respiratory burst such as NADPH oxidase, which produces O2-. The oxidation of DCFH, but not of HE, was decreased in stimulated neutrophils, which were simultaneously loaded with HE and DCFH. Intracellular DCFH oxidation induced by incubation of resting neutrophils with extracellular H2O2 was not influenced by the presence of HE. This indicates that HE is oxidized at an earlier step in the reactive oxygen metabolism of neutrophils than DCFH, i.e., by early oxygen metabolites like O2-, while DCFH is oxidized in part by H2O2 and phagosomal peroxidases. The differential oxidation of HE and DCFH during simultaneous cellular staining permits the analysis of up to three functionally different neutrophil populations in septic patients. This is of interest for the determination of disease-related alterations of oxygen metabolism in quiescent and stimulated leukocytes.

Adult↗
PubMed1989

How to build an inexpensive cyclotherm instrument for automated polymerase chain reaction.

The Cyclotherm instrument is a functionally fully equivalent but inexpensive alternative to commercial instruments for automated polymerase chain reaction (PCR). It can be rebuilt under conditions of a biochemical laboratory for less than +1000. A Peltier element is used for heating and cooling of the reaction vials and the temperature and timing of the PCR cycles are controlled by a BASIC program in a SHARP PC 1600 low cost computer.

Computer Systems↗
PubMed1989

Flow-cytometric determination of intracellular pH, esterase activity and cell volume in human leukemic cell lines following in vitro incubation with cytostatic drugs.

A recently developed flow cytometric assay method using patient tumor cells allows the determination not only of their sensitivity to cytostatic drugs but also of biochemical and biophysical parameters after treatment, such as esterase concentration and intracellular pH of the living cells. DNA-content of the dead cells and cell volume of living and dead cells. The T-cell lines CEM, Molt4, Jurkat, the B-cell lines RPMI1788, Daudi, Raji and the promyelocytic line HL60 were incubated with: cytosine arabinoside (ara-C), L-asparaginase, daunorubicin, vincristine and prednisone for 48 h. Living cells then stained with esterase and pH-dye 1,4-diacetoxy-2,3-dicyanobenzene (ADB) and dead cells with DNA-dye propidium-iodide (PI). The esterase concentration, an index of metabolic activity, decreased in the T-cell lines under the influence of ara-C, daunorubicin and vincristine, whereas in the B-cell lines smaller changes in esterase concentration were observed (P less than 0.001). A decrease in intracellular pH was seen in the ara-C and daunorubicin-incubated cells Molt4, CEM and HL60, whereas in the B-cell lines no significant change in intracellular pH was found. In all lines except Jurkat the cell volume of the surviving cells increased under the influence of certain drugs (primarily ara-C and daunorubicin); B-cell lines showed a greater swelling than T-cell lines (P = 0.001).

Antineoplastic Agents↗
PubMed1988

Sensitive non-radioactive dot-blot hybridization using DNA probes labelled with chelate group substituted psoralen and quantitative detection by europium ion fluorescence.

A new labelling method for cloned DNA probes used in hybridization assays is described. The DNA insert of recombinant plasmid DNA was made partially single-stranded for the labelling reaction by a restriction enzyme digest, followed by a controlled exonuclease III incubation. A thiol-containing psoralen derivative was covalently bound through irradiation with UV-light to the remaining double-stranded region of the plasmid DNA. The psoralen-SH groups were labelled with a large number of metal chelators (diethylentriamine pentaacetic acid, DTPA) using poly-L-lysine as a macromolecular carrier. The main advantage of the labelling procedure is that a high degree of labelling is achieved without modification of the single-stranded DNA hybridizing sequences. The specific hybrids were labelled after filter hybridization with europium ions through the chelating groups of DTPA. The europium ions were quantitatively detected by time-resolved fluorometry. The sensitivity of the assay for target DNA detection was in the low picogram range, comparable to radioactively labelled DNA probes.

Chelating Agents↗
PubMed1988

Specific nuclear elimination in polyploid plasmodia of the slime mold Physarum polycephalum.

In growing plasmodia of the myxomycete Physarum polycephalum (G2-phase), three distinct classes of nuclei with a relative DNA content of 1x, 2x, and 4x are observed in the presumed haploid strain CL. The 2x and 4x species comprise up to 35% and 5% of the nuclei. Quantitative cytofluorometric studies of nuclei isolated in either G2- or S-phase or after FUDR treatment (G1 arrest) show that the three nuclear populations undergo a synchronous mitotic cycle and that the relative DNA content of the nuclear fractions in G-2 phase reflects the 2c, 4c, and 8c state. The heterogeneity of the nuclear population does, however, seem to be restricted to the growth phase. During a starvation period of 4 days that always preceeds sporulation (and also meiosis), the 4c nuclear population is reduced to 7%, 8c nuclei are no longer detected. These results suggest that a mechanism exists in Physarum for the selective detection and elimination of polyploid nuclei.

Cell Nucleus↗
PubMed1988

Phagocytosis, intracellular pH, and cell volume in the multifunctional analysis of granulocytes by flow cytometry.

Phagocytosis of Escherichia coli K12 strain bacteria was used to measure by flow cytometry the functional activities of human granulocytes in whole blood or buffy coat preparations. In a first measurement, the increase in electric cell volume and acridine orange (AO) green and red fluorescence were used to quantify the degree of phagocytosis. In a second measurement, the intracellular pH and esterase activity of each cell were determined with 1,4-diacetoxy-2,3-dicyanobenzene to obtain information on the metabolic activities during phagocytosis and degradation of bacteria. The DNA of dead cells was simultaneously counterstained with propidium iodide in both assays. The volume, the AO green and red fluorescence, the internal pH, and esterase activity were automatically averaged for all granulocytes or lymphocytes of a measurement. The calculated mean values were transferred into the self-learning database of the DIAGNOS1-program system. The functional granulocyte parameters of normal healthy individuals can be used as reference values for the automated diagnosis of abnormal granulocytes in various infectious disease states. The assays require 1 ml of heparinized whole blood and the results are available within 1 hour.

Acridine Orange↗