Protistiary - Microscopy

<%@LANGUAGE="JAVASCRIPT" CODEPAGE="1252"%> Protistiary - Microscopy

Examples of contrast enhancement
	Paramecium bursaria imaged with an array of contrast enhacement techniques. All cells point towards the center of the circle. The most visible components of the cell are the large macronucleus, an ajacent micronucleus, symbiotic algae, and the mouth. Bright field (Koehler illumination - (1 o'clock) shows the algae and major organelles, and these are made more strongly visible when the condenser iris is closed (2 o'clock). Phase contast tends to generate a lot of noise inside the cell (9 o'clock), whereas Nomarski (DIC or differential interference contrast - 10 o'clock) creates an image that is like a slice through the cell. Interference optics (3 o'clock)convert elements with different optical properties into different colours. Polarising optics (8 o'clock) only show refringent crystals within the cell. In dark ground (7 o'clock), the object is illuminated obliquely and only refracted light is visible. Illumination with ultraviolet light causes autofluorescence of the algae and the nucleus shows up because of the addition of DAPI (5 o'clock). The nucleus is also clearly visible in the cell that has been stained with Feulgen stain (for DNA - 12 o'clock). Cells dried in indian ink have ink collected in the depressions on the cell surface and the ink collects around the cell - this is referred to as negative staining (4 o'clock). Silver stains reveal the bases of the cilia (11 o'clock). The dowload file contains intermediate sized images of each technique.
Briht field. Algae, location of macronucleus and mouth are evident.	Closed condenser iris generates enhanced contast. The contractile vacuole, cilia and more details of the cell can be seen.	Phase contrast. Differences in optical densities are converted into darkness, very good for fine details such as the cilia.	Nomariski or differential interference optics creates the effct of an optical slice through the cells, and discontinuities in optical density emerge as boundaries of light and shadow.
Intereference optics. Elements with different optical properties are converted into different colors.	Polarised light. The cell was illuminated with polarised light, and a crossed polarised placed above the speciment to eliminate any light with the original polarization. Any refringent particles in the cell which changed to poalrization of the light are visible.	Dark ground illumination. The cell was illuminated obliques, so that the only light that entered the objective was light that refracted off edges in the cell.	Fluorescence. The cell was stained with DAPI (associated with DNA) and illuminated with ultraviolet light. The chlorophylls in the algae and the DAPI radiate lower energy (longer wavelength) radiation. The ultraviolet light is filtered out to show only the fluorescence from the algae and nucleus.
This cell was suspended in the presence of Indian Ink and the preparation allowed to dry out. The ink settled around the cell and into the depressions from which the cilia emerge.	This cell has been stained with Feulgen, a stain that associated with DNA. The macronucleus and micronuclei are clearly visible, but also small nuclei can be seen in many of the symbiotic algael cells.	Silver staining is used to show the bases of the cilia. Shows the arrangement of the kineties 9 rows of cilia) on the cell.	All images b y D J Patterson
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Examples of contrast enhancement

Paramecium bursaria imaged with an array of contrast enhacement techniques. All cells point towards the center of the circle. The most visible components of the cell are the large macronucleus, an ajacent micronucleus, symbiotic algae, and the mouth. Bright field (Koehler illumination - (1 o'clock) shows the algae and major organelles, and these are made more strongly visible when the condenser iris is closed (2 o'clock). Phase contast tends to generate a lot of noise inside the cell (9 o'clock), whereas Nomarski (DIC or differential interference contrast - 10 o'clock) creates an image that is like a slice through the cell. Interference optics (3 o'clock)convert elements with different optical properties into different colours. Polarising optics (8 o'clock) only show refringent crystals within the cell. In dark ground (7 o'clock), the object is illuminated obliquely and only refracted light is visible. Illumination with ultraviolet light causes autofluorescence of the algae and the nucleus shows up because of the addition of DAPI (5 o'clock). The nucleus is also clearly visible in the cell that has been stained with Feulgen stain (for DNA - 12 o'clock). Cells dried in indian ink have ink collected in the depressions on the cell surface and the ink collects around the cell - this is referred to as negative staining (4 o'clock). Silver stains reveal the bases of the cilia (11 o'clock). The dowload file contains intermediate sized images of each technique.

Briht field. Algae, location of macronucleus and mouth are evident.

Closed condenser iris generates enhanced contast. The contractile vacuole, cilia and more details of the cell can be seen.

Phase contrast. Differences in optical densities are converted into darkness, very good for fine details such as the cilia.

Nomariski or differential interference optics creates the effct of an optical slice through the cells, and discontinuities in optical density emerge as boundaries of light and shadow.

Intereference optics. Elements with different optical properties are converted into different colors.

Polarised light. The cell was illuminated with polarised light, and a crossed polarised placed above the speciment to eliminate any light with the original polarization. Any refringent particles in the cell which changed to poalrization of the light are visible.

Dark ground illumination. The cell was illuminated obliques, so that the only light that entered the objective was light that refracted off edges in the cell.

Fluorescence. The cell was stained with DAPI (associated with DNA) and illuminated with ultraviolet light. The chlorophylls in the algae and the DAPI radiate lower energy (longer wavelength) radiation. The ultraviolet light is filtered out to show only the fluorescence from the algae and nucleus.

This cell was suspended in the presence of Indian Ink and the preparation allowed to dry out. The ink settled around the cell and into the depressions from which the cilia emerge.

This cell has been stained with Feulgen, a stain that associated with DNA. The macronucleus and micronuclei are clearly visible, but also small nuclei can be seen in many of the symbiotic algael cells.

Silver staining is used to show the bases of the cilia. Shows the arrangement of the kineties 9 rows of cilia) on the cell.

All images b y D J Patterson

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