Circular oblique illumination (COL) and the examination of blood.

Editor's note: COL is circular oblique illumination.
The author's four part series describing its creation and use begins here.

A typical full field view of blood cells in COL. The colour of both the cells and background

can vary depending on the objective in use, it's na., and the width and 'thickness' of

the annulus.

A crop of the prime CCD image above ( 2560 x 1920 pixels = x 3000 on-screen ).
For screen size of 1024x768 pixels.

x 3000 (on-screen) Wild x 50 oil Fluorite 1.0 na Objective

DF/COL using a Heine condenser. Showing red corpuscles and

two neutrophils which display their protoplasmic granularity.

The subtle positioning of the Heine accounts for wide

variation of effects of which this image is just one.

Note the neutrophil's nuclei revealing typical 'Skull Eyes' look.

x 2600 (on-screen) Wild x40 Fluotar 0.75 na

This greyscaled image shows a neutrophil and lymphocyte plus fragments of waxy looking clotting material.

The cast shadow 'electron microscope look' has been brought about by

offsetting the COL annulus, which often enhances contrast and helps to define

imagery (and also DUST) but does not suit all subject matter. This image certainly shows what can be done

with a mere 0.75na of objective aperture.

x 1850 (on-screen) using Wild x 40 0.75 na Fluotar Objective in COL. This is a sample of discharge from a

wound. Many depleted neutrophils with their characteristic nuclei in the field, and one or two lymphocytes

and red corpuscles are present too. A filament of 'micropore' dressing can be seen near the top border.

The blood smears were simple untouched preparations below a coverslip which was pressed firmly to reduce the film depth to allow COL to work at its best. Magnification estimates were based on a simple calculation using the average width of a red corpuscle which is about 7.5 microns. Thus if the image of a typical corpuscle measures 22 mm then when divided by 0.0075mm ( 7.5 microns ) the amplification on-screen is approximately x 3000.

The images were captured using a Minolta F300 digital camera above a Zeiss Photomic stand. The camera's x 3 zoom and also the x 2 'Zeiss Optovar' internal amplication unit was brought into use to represent as large an image as possible on the CCD of the camera. This effectively reduces the inherent noise of an image which tends to be exacerbated by the lower light intensity of COL.

The magnification in most cases is empty and excessive, but this was necessary to illustrate the potential of COL using a difficult subject matter without any preparation. As usual the live imaging was distinctly superior to these images where the unique colouring effects of COL and its inherently high contrast inducing properties excel.

The dreaded dust spots in eyepieces are all too evident above and also unfortunately the ringing problem with the Minolta lens, which is exagerrated by COL........ an observation but no excuse!

The list of specimens suitable for observation with COL is slowly growing..........diatoms, butterfly scales, protozoans, bacteria and blood films........enough to keep the observer busy for a lifetime.

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