LAB 2: MEASUREMENT AND COUNTING OF CELLS USING MICROSCOPE

2.1: Ocular Micrometer

INTRODUCTION

Ocular micrometer

Ocular micrometer is use in order to measure and compare size of prokaryotic and eukaryotic microorganisms. Suitable scale for their measurements should be somewhere in the microscope itself. An ocular micrometer is a glass disk that attaches to a microscope's eyepiece. For this an ocular micrometer serves as a scale or rule to measure the size of magnified objects. Ocular micrometer is simply a disc of glass upon which is etched lines. The ruler on a typical ocular micrometer has between 50 to 100 individual marks, is 2 mm long and has a distance of 0.01 mm between marks.

Technicians can easily make calculations of object size after measuring an object against the dimensions of an ocular micrometer, which are calibrated using a stage micrometer, a microscope slide with its own surface scale visible when viewed through the microscope. Because the micrometer measures a flat dimension, consider the three-dimensional aspect of the measured object. For example, the length of a curved surface may be longer than the ocular micrometer measured length.

The ocular micrometer was located inside the ocular lens placed with eyes over the microscope eyepiece. The ocular micrometer is the visible scale shown as you look through the eyepiece. Each line represents 1 micrometer.

Ocular micrometer calibrated using stage micrometer    

Figure 1.1: ocular micrometer

RESULTS

Each division of the stage micrometer = 1 m

i.                    For 40x magnification,

10 eyepiece division  = 15  stage division  = 15 µm

1 eyepiece division = 15 µm / 10 = 1.5 µm

Size of yeast   : long x width

                                                    : 7.5 µm x 9 µm

ii.                  For 100x magnification,

10 eyepiece division  = 11  stage division  = 11 µm

1 eyepiece division = 11 µm / 10 = 1.1 µm

Size of yeast   : long x width

                                                    : 11 µm x 13.2 µm

iii.                For 400x magnification,

11 eyepiece division  = 40  stage division  = 40 µm

1 eyepiece division = 40 µm / 11 = 3.6 µm

Size of yeast   : long x width

                                                    : 144 µm x 162 µm

DISCUSSION

An ocular micrometer is a small glass disk with thin lines and numbering etched in the glass.

An ocular micrometer was placed into one ocular on your microscope so that the lines superimposed on the image will allow to measure the specimen.

For each magnification, must compare the lines on the ocular micrometer to the lines on a stage micrometer.

The stage micrometer is a glass slide with precisely space lines etched at known intervals.

The vertical distance of an object that is in focus.  When magnification is increased, less of the object is in focus (depth of field decreases) – but greater detail of the area in focus can be seen.

You may have to adjust the focus of your eyepiece in order to make the scale as sharp as possible. If you do that, also adjust the other eyepiece to match the focus. Any ocular scale must be calibrated, using a device called a stage micrometer

The ocular lenses usually magnify 10X.  Thus the total magnification observed is the multiplication of the power of magnification of the ocular times the objective.  For example an object magnified by the ocular and the 40X high-dry objective is viewed at 4002 times its real size.  Most ocular lenses can be moved back and forth to adjust to the interpupillary distance of the student.  When first using the microscope, adjust the ocular lenses back and forth until a circular field is viewed with both eyes open.  Additionally, many microscopes allow the ocular lenses to be adjusted up and down (mechanical tube length adjustment) and there is a scale alongside the tube.  After adjusting the interpupillary distance, read the distance off the scale and adjust the tube length of the ocular lens to the same value.  Now the ocular lenses are adjusted to your eyes.