Sterilization

C H A P T E R

The terms sterilization and disinfection are used to indicate

the treatment of material so as to destroy or otherwise

eliminate any living organisms present. However, the

term sterilization is used where physical methods are

used and disinfection is used where chemical agents are

made use of.

METHODS COMMONLY USED FOR STERILIZATION

The methods used commonly in practice are:

1. Killing organisms by heat: Heat may be dry or moist

2. Destroying organisms by employing chemical antiseptics,

e.g. lysol, phenol, perchloride of mercury, etc.

3. Removing organisms mechanically by filtration, e.g.

Seitz, unglazed porcelain.

Sterilization by Heat

Adequate heat is the most certain and rapid method for

sterilization. The time needed for sterilization is inversely

related to the temperature of exposure—the higher the

temperature, the shorter the time needed. High temperature

kills bacteria by coagulating their proteins. Different

types of bacteria show considerable differences in heat

susceptibility. In general, vegetative forms are destroyed

at lower temperatures, whereas high temperatures are

needed for sporing organisms.

Dry Heat

This is the preferred method for sterilizing glassware, e.g.

of glass syringes and of materials such as oils, jellies and

powders which are impervious to steam. Dry heat requires

a much higher temperature or a much longer time at the

same temperature than does moist heat. Dry heat can be

used in the following ways:

Flaming

The articles are passed through the Bunsen flame, without

letting them become red hot. It is used for scalpels, needles,

mouths of culture tubes, glass slides, coverslips and points

of forceps. Only the surfaces actually touched by the flame

are sterilized.

Red Heat

Platinum loops, inoculating wires and needles are heated

in the Bunsen flame until red hot.

Hot Air Oven

These are electrically heated and thermostatically

controlled. The oven itself is a double-walled steel chamber

with a stout door. The top or side contains a ventilator

which is left open during sterilization to disperse any

moisture or volatile matter. Air circulates within the oven

by convection currents. Suitable sterilizing times in the

hot air oven are 3 hours at 140°C, 1 hour at 160°C and

30 minutes at 180°C. All dry glassware, such as test tubes,

petri-dishes, flasks, pipettes and throat swabs, etc. are

made sterile by using hot air oven.

This method is not suitable for sterilizing culture

media, liquids, rubber connections, glass to metal fitting

and fabrics, e.g. masks, towels or gowns.

Moist Heat

Temperature

A temperature of 60 to 65°C kills most vegetative bacteria

(made use of in pasteurization of milk and preparation of

vaccines).

Boiling

Boiling is frequently used for sterilizing syringes, etc. but is

not adequate as many spores withstand this temperature.

30 Concise Book of Medical Laboratory Technology: Methods and Interpretations

Steam

Steam is the most effective technique of moist heat

sterilization. Steam may be employed in three ways.

Steam at 100°C

The apparatus used commonly is called Koch’s steamer. It

has a vertical metal cylinder with a conical lid. It is fitted

with a thermometer and has a small opening for escape

of steam.

Sterilization by free steam can be done in two ways.

Prolonged exposure: For 1½ hours, used for broth or

nutrient agar.

Intermittent heat or tyndallization: It involves exposure for

20 minutes on three successive days and is used to sterilize

sugars and gelatin which decompose on higher temperatures.

Principle: Spores would germinate after first steaming and

destroyed on the next, three steamings would eliminate all

spores and their vegetative forms.

Low Temperature Steam

This method is employed for sterilizing materials (blankets, polyethene tubing, etc.), which would be damaged at

higher temperatures.

Steam at Temperatures above 100°C (Autoclaving)

Autoclaves are made of strong metal jackets; strong enough

to withstand high pressures required (Figs 2.1A to C). The

autoclave door is hermetically sealed. It has a safety valve

set to blow off at a predetermined pressure. The principle

is that water boils when its vapor pressure is equal to the