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Biomedical subjects

D M Benson

Publications and source records attributed to D M Benson.

54 records · Page 3Linked to original sources

Use of an encapsulated fluorescent probe to measure intracellular PO2.

The objectives of this investigation were to produce a reliable, sensitive probe to measure intracellular PO2 with a high degree of resolution and to apply this technique to biological systems. A fluorescent molecule, pyrene dissolved in paraffin oil, was encapsulated in polyacrylamide to form a probe of nanometer dimensions. The quantitative and microscopic oxygen values were determined by analyzing the quenching of the fluorescence of the probe by oxygen, as displayed on a television monitor by a silicon-intensified-target camera. The nanocapsules had a sensitivity of approximately 1 mm PO2, a spatial resolution of 0.5 micrometer, and a temporal resolution of milliseconds. Calibrated nanocapsules within nonrespiring Amoeba proteus responded to ambient partial pressures of oxygen. At two different ambient partial pressures, nanocapsules engulfed by respiring amoebas indicated an intracellular PO2 28 mm Hg less than extracellular PO2. The capsules retained their sensitivity to oxygen for at least 8 months.

Amoeba↗

Intracellular oxygen measurements of mouse liver cells using quantitative fluorescence video microscopy.

Our currently developed fluorescence video microscope can measure fluorescence intensities with an error of +/- 1.5% of full scale in 65536 different positions of a microscope field. With a video frame freeze acquisition time of 33 ms, time-dependent changes of this order of time or slower can be followed. Using cells which have absorbed pyrene-1-butyrate to an intracellular concentration of 0.05 to 1 mM, the changes in fluorescence intensity with oxygen concentration are easily measured. The spatial resolution for data collection is 0.5 micron when a 54X objective is used. The individual Stern-Volmer quenching constants of each individual pixel were measured for agar slices and mouse liver cells treated with pyrenebutyric acid. The distribution of quenching constants for agar follows a normal curve about a mean value of 16 . 10(-4) torr-1. The data for mouse liver cells gave a non-normal distribution of quenching constants with a mean value of 18 . 10(-4) torr-1. The greater spread of the data from cells is interpreted as evidence for a real biological variation in the solubility coefficient of oxygen in different locations within the cell. In all the cells examined, this distribution has been observed to be non-random and appears to be associated with specific cell structures.

Animals↗

Characterization of benign and malignant tissues of the thyroid gland. An ultrasonic approach using RF waveform analysis and pattern recognition.

A physical basis was developed for an accurate noninvasive technique to differentiate malignant thyroid tumors from benign lesions based upon a frequency-dependent attenuation model. The research effort utilized RF waveform analysis via a minicomputer based digital processing system. Data collection was performed using a standard clinical B-Mode diagnostic instrument with a transducer specially designed for imaging the thyroid gland. Differences between the frequency spectra of the RF waveforms for benign and malignant pathologies could be demonstrated in accordance with the proposed model.

Diagnosis, Differential↗

Fixation of testicular tissue for immunohistochemical and ultrastructural examination.

The fixation of testicular tissue with glutardialdehyde destroys the antigenicity of cell epitopes in many cases. To obtain both, morphological and immunohistochemical examination, a fixation which could preserve the antigenicity and condition of the testicular structure was sought. The solution obtained was a mixture of 3.7% formalin with 0.2% glutardialdehyde and 0.05% saponin in a phosphate buffer of pH 7.4. Blocks of human tests were immersed in this fixative, embedded in Epon 812 or paraffin for conventional light and electron microscopy and immunohistochemical staining. The immunohistochemical examination was focused on the lamina propria of the human seminiferous tubules. Good preservation of the structure was observed both in light and electron microscopy. Cytological details were seen by light microscopy, especially the recognition of single tumour cells. Electron microscopically, all cells of the seminiferous tubules and the lamina propria showed a well-preserved internal structure. At the same time the peritubular cells of the lamina propria exhibited a well-expressed vimentin and desmin immunoreactivity, thus providing evidence that the corresponding epitopes retain their antigenicity under these fixation conditions. The applied fixation procedure provides comparable results in preservation of the structure to glutardialdehyde but does not destroy the antigenicity of epitopes.

Acetates↗