Importance

1. For identifying mycobacteria.

2. It is used extensively in fluorescent antibody techniques

used in parasitology and bacteriology. FIG. 1.11: The principle of dark ground illumination

22 Concise Book of Medical Laboratory Technology: Methods and Interpretations 3. It is also used widely in histopathology of kidney, skin,

etc. where immune/autoimmune basis of disease is

expected. In fact, anything can be confirmed with

high degree of sensitivity and specificity, if antibodies

against it (later tagged with a fluorescent dye) can be

produced.

4. Used widely in cytogenetics.

Electron Microscope

Basic Principle

The resolution of the light microscope has been shown

to be limited by the NA and the wavelength of light

employed. As the degree of correction in glass lenses is

very high, the main limitation is imposed by the light

(e.g. half wavelength of light), giving a normal resolution

of approximately 250 nm; and when UV light is used,

a resolution of about 100 nm. By the substitution of an

electron beam for light rays, a much greater degree of

resolution can be obtained; since at an acceleration of

50,000 volts, electrons have a wavelength of only 0.001 nm;

therefore, a theoretical resolving power of 0.0005 nm could

be attained, which would enable molecules to be seen.

Unfortunately, the degree of correction that is currently

feasible with transmission electron microscope (TEM)

lenses will permit a resolution of only 0.25 nm, but this is

still a thousand times greater than that possible with the

light microscope. A further difficulty with the TEM is that,

since electrons have poor penetrating power, the sections

to be examined must be very thin, less than 50 nm thick.

This necessitates the use of special hard embedding media

(plastics) and special ultra-microtomes to cut such thin

sections. Steel knives cannot be used to cut these sections;

either glass or diamond knives are used.

Weighing Scales or Analytical Balance

Weighing scales: For weighing large quantities.

Analytical balance: For accurate weighing of smaller

quantities.

Use and Care

1. The weighing equipment must be placed on a firm

bench, away from vibration, draughts, direct sunlight

and dust.

2. It should be kept perfectly horizontal by altering the

screws on which the equipment stands.

3. Chemicals, etc. should never be placed directly on the

pans. Weigh them in a container.

4. Never touch the weights with hands, handle them with

forceps.

5. The balance should be at rest before adding or

removing the weights or chemicals.

6. Before taking the reading, the glass window of the

instrument should be closed.

Electronic analytical balances are also available. Made

by various companies, these are very accurate.

Centrifuge

Centrifuge is used to sediment or deposit rapidly particles

such as cells which may be suspended in a fluid. The speed

is expressed as rpm, i.e. revolutions per minute.

Relative Centrifugal Force (RCF)

More important than rpm is relative centrifugal force

(RCF). RCF is expressed as the acceleration due to gravity

or G (dynes per cm). The formula is:

G = 0.00001118 × (r) × (n)2

where r = radius in centimeters

and n = revolutions per minute.

The time of centrifugation is equally important. The tubes

should be spun for a definite period to obtain the desired

effect.

Types of Centrifuge

Hand Centrifuge

Fixed to the bench, the handle is rotated manually. It gives

low speeds only.

FIG. 1.12: Components of fluorescence system

Laboratory 23

Motor-driven Centrifuge

Operated through mains electricity supply. The tubes

may be kept in a fixed angle head or in a swing out head

(Figs 1.13 and 1.14).

Microhematocrit Centrifuge

Also motor driven for finding out packed cell volume

(PCV) of red blood cells (RBCs). In this, blood-filled

capillary tubes are spun and later the percentage of

RBC-filled column is estimated (Figs 1.15 and 1.16).

Use and Care

1. Use centrifuge tubes made of strong glass and they

should not be too long.

2. The opposite tubes should be balanced properly.

3. The centrifuge speed should be increased gradually.

4. The instrument should be kept clean. If something

spills over inside, it should be cleaned and the

instrument disinfected, if necessary.

FIG. 1.13: Swing out head centrifuge

(Courtesy: Yorco Sales Pvt. Ltd)

FIG. 1.14: Motor driven centrifuge with rpm. indicator and auto

(timed) shut off

(Courtesy: Yorco Sales Pvt. Ltd)

FIG. 1.15: Dual centrifuge routine centrifuge with microhematocrit

attachment

(Courtesy: Yorco Sales Pvt. Ltd)

FIG. 1.16: Microhematocrit centrifuge and its parts

(Courtesy: Yorco Sales Pvt. Ltd)

24 Concise Book of Medical Laboratory Technology: Methods and Interpretations Glassware (Many Items are now Made of Plastic)

1. Flasks—are of different sizes and shapes.

a. Erlenmeyer or conical flasks—for heating and

boiling liquids (Fig. 1.17).

b. Volumetric flasks—are graduated for getting

exact volume of liquids (Fig. 1.18).

c. Round and flat-bottomed flasks for preparing

solutions (Figs 1.19A and B).

2. Beakers—available in different sizes (Fig. 1.20).

3. Bottles