scholarly journals AUTOMATED ANALYSIS OF DEOXYRIBONUCLEIC ACID, PROTEIN AND NUCLEAR TO CYTOPLASMIC RELATIONSHIPS IN TUMOR CELLS AND GYNECOLOGIC SPECIMENS

1974 ◽  
Vol 22 (7) ◽  
pp. 616-621 ◽  
Author(s):  
J. A. STEINKAMP ◽  
H. A. CRISSMAN
Keyword(s):  
Tumor Cells ◽  
Optical Sensors ◽  
Deoxyribonucleic Acid ◽  
Pulse Height ◽  
Argon Laser ◽  
Total Protein Content ◽  
Fluorescence Signal ◽  
Green Fluorescence ◽  
Time Duration ◽  
Acid Protein

Quantitative two-color fluorescence staining techniques, coupled with flow system multiparameter cell analysis and sorting instrumentation, have been used for rapid, simultaneous determination of deoxyribonucleic acid, protein, nuclear (N) and cytoplasmic (C) diameters and N:C ratios in mammalian tumor cells and human gynecologic specimens. Cells stained in suspension for deoxyribonucleic acid and total protein content, respectively, with propidium iodide (red fluorescence) and fluorescein isothiocyanate (green fluorescence), enter a flow chamber and intersect an argon laser beam which excites cellular fluorescence. Optical sensors measure both red and green fluorescence plus the time duration of each fluorescence signal which is proportional to nuclear and cytoplasmic diameters, respectively. The resulting signals are processed and displayed as frequency distribution histograms using a multichannel pulse height analyzer. Cells are also sorted based on N:C ratios. Illustrative examples of preliminary two-color fluorescence analysis and sorting of mouse squamous tumor cells and human exfoliative vaginal cells are presented.

scholarly journals Construction and Evaluation of the Tumor-Targeting, Cell-Penetrating Multifunctional Molecular Probe iCREKA

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Li-juan Wang ◽  
Hong-sheng Li ◽  
Quan-shi Wang ◽  
Hu-bing Wu ◽  
Yan-jiang Han ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Tumor Targeting ◽  
Cyclic Peptide ◽  
Tumor Imaging ◽  
Tumor Stroma ◽  
Molecular Probe ◽  
Fluorescence Signal ◽  
Glioma Tumor

A novel tumor stroma targeting and membrane-penetrating cyclic peptide, named iCREKA, was designed and labeled by fluorescein isothiocyanate (FITC) and positron emitter 18F to build the tumor-targeting tracers. The FITC-iCREKA was proved to have significantly higher cellular uptake in the glioma U87 cells in the presence of activated MMP-2 than that in absence of activated MMP-2 by cells fluorescence test in vitro. The tumor tissue fluorescence microscope imaging demonstrated that FITC-iCREKA accumulated in the walls of the blood vessels and the surrounding stroma in the glioma tumor at 1 h after intravenous injection. While at 3 h after injection, FITC-iCREKA was found to be uptaken in the tumor cells. However, the control FITC-CREKA can only be found in the tumor stroma, not in the tumor cells, no matter at 1 h or 3 h after injection. The whole-animal fluorescence imaging showed that the glioma tumor could be visualized clearly with high fluorescence signal. The microPET/CT imaging further demonstrated that 18F-iCREKA could target U87MG tumor in vivo from 30 min to 2 h after injection. The present study indicated the iCREKA had the capacity of tumor stroma targeting and the membrane-penetrating. It was potential to be developed as the fluorescent and PET tracers for tumor imaging.

scholarly journals Study on Characteristics of Chemokine CXCL10 Gene Cloned from cDNA Expression Library of Ujumqin Sheep

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
P. F. Hu ◽  
X. C. Li ◽  
N. Lei ◽  
X. Y. Lan ◽  
Q. J. Zhao ◽  
...  
Keyword(s):  
Immune Responses ◽  
Transfection Efficiency ◽  
Peak Level ◽  
Fluorescence Signal ◽  
Cdna Expression Library ◽  
Fibroblast Cells ◽  
Green Fluorescence ◽  
Cdna Fragment ◽  
Expression Library ◽  
Cdna Expression

Chemokines were a major regulator of body’s inflammatory and immune responses. In this study, the cDNA fragment of chemokine CXC ligand 10 (CXCL10) was cloned from the Ujumqin sheep ear marginal tissue cDNA expression library; the CXCL10 gene had 103 amino acids and a molecular weight of 11.47 kDa, and it shared a high homology among cattle, sheep, and goat, while a low homology compared with mouse. The CXCL10 protein had 4 conservative cysteine residues, located in 28, 30, 55, and 72 sites. The expression pattern and intracellular distribution of recombinant CXCL10 proteins in Ujumqin sheep fibroblast cells showed that there were green fluorescence signals both in cytoplasm and nucleolus after 24 h of transfection, the number of positive cells was increased with time, the peak level of fluorescence signal was reached after 48 h of transfection and the transfection efficiency was 33.3%; there was a significant decrease in fluorescence intensity after 72 h of transfection. Expression of recombinant CXCL10 gene inEscherichia colihad a time- and temperature-dependency on the amount of protein expression, and a small quantity of inducer was needed.

scholarly journals AMINOACRIDINE DYES AND THE SECONDARY STRUCTURE OF DNA IN SITU

1971 ◽  
Vol 19 (12) ◽  
pp. 761-765 ◽  
Author(s):  
MANUEL DIAZ ◽  
JOSE HIERRO ◽  
GRACIELA DEMICHELI DE DIAZ
Keyword(s):  
Secondary Structure ◽  
Acid Solution ◽  
Nitrous Acid ◽  
Deoxyribonucleic Acid ◽  
New Method ◽  
Green Fluorescence ◽  
Double Stranded Dna

A new method is proposed to study the secondary structure of deoxyribonucleic acid (DNA) in situ in fixed chromatin. It is based on acriflavine staining and on differentiation with a nitrous acid solution of the fixed cytologic preparation. The presence of green fluorescence after this treatment is regarded as indicative of double stranded DNA. Experiments are described with DNA-acriflavine mixtures in solution, DNA-agar models and cytologic preparations submitted to different pretreatments. The feasibility and limitations of the method are discussed in the light of the results reported upon.

Novel Flowcytometry-Based Approach for Detection of Tumor Cells in Body Fluid Using Automated Hematology Analyzer

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5600-5600
Author(s):  
Yoko Tabe ◽  
Hiroyuki Takemura ◽  
Konobu Kimura ◽  
Aya Konishi ◽  
Takashi Horii ◽  
...  
Keyword(s):  
Positive Predictive Value ◽  
Negative Predictive Value ◽  
Tumor Cells ◽  
Chronic Inflammation ◽  
Predictive Value ◽  
Body Fluid ◽  
Mononuclear Cells ◽  
Fluorescence Signal ◽  
Hematology Analyzer ◽  
Automated Hematology Analyzer

Abstract Introduction: Nucleated cells differential analysis of body fluid (BF) samples is important diagnostic tool for several diseases including cancer metastasis. Detection of tumor cells in BF requires the manual morphological scanning of slides by the cytopathologists, which is time-consuming, labor-intensive and not always reliable because of a relatively low overall sensitivity rates (ranging 40-90%) with the higher false-negative rates for lymphomas and mesotheliomas. This study aimed to develop the scattergram gating analysis for detection of tumor cells in BF using the automated hematology analyzer Sysmex XN-1000 (Sysmex, Kobe, Japan). Methods: We used a total of 220 BF samples (53 cerebrospinal fluids, 73 pleural or ascitic fluids, and 94 peritoneal dialysis effluent) obtained from patients with cytological diagnoses (papanicolaou stain) including negative and positive of tumor cells, and chronic inflammation with an elevated lymphocyte and histiocyte fractions. As a reference method, morphological manual differential (200 cells counts) was performed by two experienced technologists using cytospin slides stained with the May-Grunwald Giemsa. The BF mode of XN-1000 (XN-BF) determines the differential cell counts of BF samples using side scatter and fluorescence intensity in WDF channel after the nuclear DNA/RNA stain by nucleic acid dye. The polymorphonuclear cells, mononuclear cells and high fluorescence cells (HF-BF), corresponding with a high amount of nucleic acids, are differentiated. Mesothelial cells and macrophages are theoretically categorized as HF-BF cells and included in the total nucleated cell count but not in the WBC count. We selected the tumor cells positive (n=18) and chronic inflammation positive samples (n=108) by morphological manual differential, and reviewed their scattergram patterns determined by XN-BF. Then the novel scattergram gating strategy targeting the tumor cells was evaluated. The gating criteria were based on the WDF scatter plots; #1: detect the cells with larger size and higher fluorescence signal in comparison with general leukocytes, which mainly derived from clustered tumor cells, #2: detect the middle sized mononuclear cells with less granules rather than neutrophils and similar fluorescence signal to monocytes, which targeting hematological malignant cells and solid tumor cells. BF samples that meet at least one criterion were interpreted as positive for tumor cells. Results: The malignant BF samples containing tumor cells showed the different scattergram patterns from the benign ones with chronic inflammation; the malignant BF formed the isolated cellular clusters in our gating system, and the inflammatory BF showed the continuous expansion into the HF-BF area. Our scattergram gating analysis achieved an overall sensitivity of 72.2% and specificity of 98.0% in detecting tumor cells positive samples when screening against all samples outcomes. The positive predictive value was 76.5% and the negative predictive value was 97.5%. For the samples with absence of tumor cells and inflammatory observations (n=94), no false positive was detected. Of notes, each of our gating criterion detected the different type of tumor cells. For example, the scattergram gating analysis #1 achieved an overall sensitivity of 72.7% and specificity of 99.0% in detecting adenocarcinoma with the positive predictive value of 80.0% and the negative predictive value of 98.6%. Conclusions: A simple measurement of BF by automated hematology analyzer in which cells are minimally handled has a potential to reduce costs and allow routine cell screening in clinical applications. For the BF malignancy diagnostics, the scattergram gating analysis is promising to (i) augment diagnostic routines without requiring additional sample preparation procedure, (ii) limit operator bias, and (iii) provide a standardized measurement. Disclosures No relevant conflicts of interest to declare.

A biodistribution of 5-fluorouracil co-administered with and without galactomannan derivative, in athymic mice implanted with COLO 205 human xenografts

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 12027-12027
Author(s):  
E. Zomer ◽  
A. Klyosov ◽  
D. Platt ◽  
R. Stevens
Keyword(s):  
Phase I ◽  
Tumor Cells ◽  
Phase I Study ◽  
Area Under The Curve ◽  
Systemic Exposure ◽  
Metastatic Tumor ◽  
Athymic Mice ◽  
Time Duration ◽  
Post Dose ◽  
Mediate Cell

12027 Background: The polysaccharide [(1–4)-linked β-D-mannopyranose]17- [(1–6)-linked-α-D-galactopyranose]10 (Davanat [D]), enhances activity of 5-Fluorouracil (5-FU) in 5-FU refractory murine colon tumor models. D binds to surface lectins and promotes transport of 5-FU into the tumor cells. Lectins are present in large amounts on the surface of metastatic tumor cells, and may mediate cell association, apoptosis and metastasis. Method: A preclinical biodistribution study was initiated to evaluate the intravenous (i.v.) administration of 5-FU (36 mg/kg/dose) in the absence and presence of D (60 mg/kg/dose) in female athymic mice with tumors. Forty mice either received 5-FU alone or with D on days 24–27 (Q1d × 4) after tumor implantation. On day 27, a (2-14C) labeled 5-FU was added to the regimen. After the injection, ten mice per time point (0.5, 1.5, 4 and 24 h) were bled, euthanized and then the plasma, tumor and tissues were assayed for radioactivity by a liquid scintillation analyzer. Objectives were to determine the biodistribution of 14C-5-FU in the absence and presence of D in plasma, tumor and tissues. Results: 14C-5-FU disappeared in a bi-phasic manner after i.v. injection both in the presence and absence of D. Addition of D increased the amount of 5-FU exposure as measured by area under the curve (AUC) from 1.5 to 24 hours post-dose. This finding confirms results from a Phase I study where administering escalating doses 30 to 280 mg/m2 D with fixed 500 mg/m2 5-FU also demonstrated an increase in 5-FU systemic exposure as the D dose increased to 280 mg/m2 Conclusions: In mice, 14C-5-FU dosed with D increased the overall exposure of 14C-5-FU during the 1.5 to 24 hr sampling time duration in plasma, tumor and kidneys, when compared to 14C-5-FU dosed alone. These results, and the results obtained from a Phase I study where D was co-administered with 5-, warrants further testing in a Phase II/III clinical trial. [Table: see text]

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