INSTRUMENTATION

Objectives:

1. Identify various types of laboratory instruments and apparatus

2. Describe the principles of operation for the laboratory instruments
3. State the roles of essential functional units of the instruments
4. Observe the routine care and mantainace and decontamination of the instruments
5. Describe quality assurance of the instruments

MICROSCOPES:

An instrument for examining minute objects which cannot be seen with naked eye. There are 2
types of microscopes:

1. Monocular
2. Binocular

Approach to microscopy:

Concepts on principle of operation:

1. Refraction: change in direction of light passing obliquely (slanting or sloping direction)

from one medium to another of different optical density.
 2. Refractive index:
Sine of angle of incidence (AON) = Constant
Sine of angle of refraction (BON)

i.e. constant quantity for any two given media. It determines the refractive index of glass.

3. Spherical aberration: fuzzy appearance (indistinct) of the outer part of the field of view
of lens caused by non – convergence of rays to a common focus. It leads to distorted
image. Marginal rays intercepts axis closer than central rays.

4. Chromatic aberration: it is a non – convergence of the coloured components of white

light to a common focus. The images of colours are not clear and sharp as they are
surrounded by halo of unfocused light of other colours producing rain bow – like fringe.

NB: Bi – convex lens like prism splits white light into its components colours. Blue light being
refracted more than the red, so it comes to focus nearer to the lens.

5. Converging lens (bi – convex lens)

It has 2 spherical surfaces that curve outwards. It is named so because rays passing through
them converge to a focal point.

Components:

• Centres of curvature - centre of lens surface

• Principal focus – point of convergence of rays parallel to axis.
• Principal axis – straight line between these 2 centers of lens
• Principal plane – line at right angles to this principal axis
• Optical centre – centre of curvature.
• Focal length – distance between optical centre and principal focus
• Working distance – distance between the surface of the lens and the focal point.

NB: A bi – convex lens has 2 principal foci, one on either side.

6. Diverging lens:

Has 2 surfaces which curve inwards (bi – concave lens)

Rays passing through them are refracted away from principal axis as though originating from
principal focus.

Image formation:

Real image: converging lens can produce either a real image or virtual image. Real image is an
inverted image, which can be projected onto a screen. It is formed when object is placed
outside the focal length of lens or lens system. Size of image produced depends on distance
between the lens and the object.

Examples:

Let ‘u’ be a distance between object and lens.

‘2f’ be twice the focal length of the lens

‘f ‘ be focal length

Assignment:

Using ray diagrams (accurate) locate and describe the images formed when the object is placed
at the following points:

1. u >2f
2. u =2f
3. u<2f
4. u<f

7. Resolving power: power of lens to reveal details; also called resolution of the lens.
8. Numerical aperture: ratio of diameter of lens to its focal length or it is a measurement
and description of resolving of the objective or it is exact figure that has been worked
out mathematically from its equivalent focal length and lens diameter. Both NA and
magnification of objective are usually engraved on it. It can be the product of the
refractive index of medium outside the lens(n) and the sine of half the angle of the cone
of light absorbed by front lens of objective (v).

N.A = n x sine u

9. Useful magnification: it is where maximum magnification of the object is achieved with

clear definite images.
10. Empty magnification: it is the magnification without resolution. i.e. image looks hazy
and lacks definition
11. Primary magnification: magnification produced by the objective itself.
 12. Secondary magnification: magnification produced by eyepiece
13. Total magnification: total magnifying power of the compound microscope is the product
of the magnification of the objective and eye piece at normal optical tube length of 160
mm.

Example: 1.

Objective 40x

Eyepiece 10x

If tube length is 160 mm (normal optical tube length). Total magnification = 40x10 = 400x.

Example: 2.

If the tube length is varied, formula for final magnification =

(Magnification of objective) x (magnification of eye piece )x (working tube length)

(Normal optical tube length)

Objective = 40x, eye piece = 10x, working tube length = 180 mm

Total magnification = 40x10x180 = 450x

160
Example: 3

If objective are marked with focal length only.

Total magnification is:

(Magnification of eye piece) x working tube only

Focal length of objective

Eye piece = 10x, focal length= 4mm, working tube length = 160mm

Therefore: total magnification = 10x160 = 400x

4
MICROSCOPE COMPONENTS:

Microscope is an instrument for examining minute object which cannot be seen with naked
eye. We have 2 types of microscopes: monocular and binocular.

Laboratory microscope consists of 3 sets of parts:

1. The stand
2. Mechanical adjustments
3. The optics of the microscope
 1. The microscopic stand:
It consists of:

• The tube – holds objectives and eyepiece lens

• The body – it is a block of metal containing the focusing mechanisms. It is
attached to the tube.
• The arm – it provides a lifting handle for microscope and supports the body and
tube.
• The stage - it is a flat plate where specimen lies at its centre.
• Mechanical stage-it holds slide in place and move it systematically in straight
lines either across or along the stage by the rotation of two knobs.
• The substage – it is below the stage and holds a condenser lens with an iris
diaphragm and holder for light filters.
• The foot (base) – offers a resting base for the whole instrument.

Mechanical adjustments:
Includes:
• The course and fine adjusting mechanisms
• The condenser adjustments

NB:
o low power lenses are focused by course adjustment knobs.
o Condenser can be focused by rotating knob (i.e. up and down).
o Its aperture is adjusted by the iris diaphragm.

The microscopic optics:

Include:
o The objectives
o The eyepiece
o The condenser and iris and filters
o The mirror(or electrical source of illumination)

Microscope objectives:
Quality of the image depends on objectives. It is engraved with both magnification and
numerical aperture scale.
Upon N.A, depends on highest resolving power and useful magnification of objective.
Rule: total magnification of the microscope shouldnot exceed the N.A. of the objective being
used multiplied by 1000.
3 types of objectives:
o Achromatic – routine microscopic
o Fluorite – valuable for searching blood films
o Apochromatic – for special work.
Filters:
Uses:
o To reduce intensity of light when required
o To increase contrast and resolution
o To transmit light of selected wavelength
o To protect eye from injury caused by U.V. light.

Oil – immersion:
Uses:
o To prevent bending of light when leaving glass slide and allowing little light in the
objective
o Prevent bending of light which affects N.A. and resolving power of objective.

Eye piece:
Common form: Huygens eyepiece with 2 lenses. Range of magnification usually 4x,6x,
8x,10x,15x and 20x. Higher the power, greater the total magnification of the microscope; lower
the power of the eyepiece, the brighter and sharper is the image.

Condenser and iris:

It is a large lens with an iris diaphragm. It is mounted below the stage. Consists of 2-3 lenses
that direct light to objectives from illuminant.

Sources of illumination:
Day light, electric light, battery lamp and oil lamp.

HOW TO USE THE MICROSCOPE:

Procedure:
1. Place it on a firm bench so that it does not vibrate, user must sit at the correct height
2. Select the source of light; incase of inbuilt source, just switch on or reflect the light with
a mirror
3. Place the specimen on the stage, making sure the underside of the slide is dry
4. Select the objective to be used, always better to begin with lower objective before you
use higher power lens e.g. 10x
5. Focus the objective i.e. using course focusing knob, rack the objective downwards as
you observe at the side, then looking through the eyepiece rack upwards until focus is
reached.
6. Adjust the light to obtain good illumination
7. Focus the condenser; it is done by focusing the details of the light source and then
racking downwards until they are out of focus
8. Adjust aperture of the condenser iris , smaller the aperture , greater is the contrast
 9. Examine the specimen, moving it by the mechanical stage systematically
10. For higher magnification, swing the 40x objective into place
11. Focus the 40x objective using the fine adjustment
12. Adjust the iris for this objective
13. For the highest magnification, add a drop of oil – immersion to the specimen and swing
the 100 x oil- immersion objectives into place
14. Open the iris fully to fill the oil – immersion objective with light
15. Adjust the binocular head t fit user’s eyes; it is done by sliding two eye pieces after focus
is reached.

DOS AND DON’T’S OF MICROSCOPY

1. Do cover microscope when not in use

2. Do remove all oil-immersion from objective immediately after use
3. Do clean the optical parts with lens paper before use
4. Do support the microscope carefully, when moving it
5. Don’t rack the objective downwards to bring the object into focus whilst looking down
the microscope
6. Don’t attempt to dismantle objectives
7. Don’t use a high power objective when a low power one is sufficient
8. Don’t lubricate the microscope with oil other than that provided for the purpose
9. Don’t place wet preparations on the stage without wiping the undersurface of the slide.

TYPES OF MICROSCOPES
1. Phase contrast
2. Dark ground illumination
3. Fluorescence microscopy
4. Polarizing microscope
5. Interference microscope
6. MacArthur microscope
7. Electron microscope

Phase contrast microscope

Principle:
A ray of light is regarded as of consisting waves travelling in straight line. If 2 rays of similar
waves travel together in step with each other, they are said to be in phase; then they produce a
brighter illumination. If they are out of step with each other, then they are out of step phase,
then they hinder and interfere with each other and hence less bright illumination. This variation
in the intensity of illumination results in increased contrast in the microscopical image.

Areas of use:
To examine bacteria such as Vibrio cholerae, amoeba in wet preparation, trypanosomes in
blood, and trichomonas vaginalis in direct smears.
Dark ground microscope
Used when maximum contrast is needed.
Principle:
Light enters special condenser which has a central blacked – out areas that it cannot pass
directly through it to enter the objective; instead light is reflected to pass through outer edge of
the condenser lens at wide angle illuminating micro – organisms by a ring of light surrounding
them.
Areas of use:
To examine: treponema pallidum, borellia species in blood, leptospira in urine, microfilaria in
blood, Vibrio cholerae.

Fluorescence microscope/polarizing microscope/interference microscope:

Principle:

Ultraviolet light or just visible deep blue light may be used to illuminate particles or micro-
organisms which have been previously stained with fluorescent dyes. These dyes transform
invisible ultraviolet light into visible light or just visible deep blue light into much more visible
yellow or orange light by increasing their wave length. This enables the organism to be easily
seen, glowing against a black background.

Areas of use:
A rapid identification of acidfast organisms e.g. TB.
Applied in fluorescent antibody technique in parasitology and bacteriology.

Mc Arthur microscope:
Area of use:
Research in malarial parasites (malarial infection) and other tropical diseases.

Electron microscope:
Area of use:
Virus identification
COLORIMETER

It is an instrument used for measuring the amount of light which passes through a coloured
solution. It refers to a device that measures the absorbance of particular wavelengths of light
by specific solution. It determines the concentration of a known solute in a given solution by
the application of beer – Lambert law, which states that concentration of a solute is
proportional to absorbance. The unit of absorbed light is optical density (OD) which is also
called the extinction coefficient. The colorimeter was invented in the year 1870 by Louis J Duboscq.

Colorimetry is based on beer – Lambert’s law:

It states : when monochromatic light passes through a coloured solution , the amount of light
transmitted decreases exponentially with the increase in the concentration of the solution
and with the increase in the thickness of the layer of solution through which the light passes.
Using a standard, this law can be applied to measuring concentration of a substance in a test
solution by using a formula.

Absorbance of the test

Concentration of the test = ____________________ x concentration of the standard
Absorbance of standard

a) Beer’s law :

States: The amount of monochromatic light absorbed by a dilute coloured solution is

directly proportional to the concentration of the solution

b) Lambert’s law

States: The amount of monochromatic light absorbed by a dilute coloured solution is

directly proportional to the thickness of the solution.

The essential parts of a colorimeter are:

• a light source (often an ordinary low-voltage filament lamp)

• an adjustable aperture
• a set of colored filters
• a cuvette to hold the working solution
• a detector (usually a photoresistor) to measure the transmitted light
• a meter to display the output from the detector

NB: light should be rendered monochromatic i.e. light of one colour or having a narrow band in
the spectrum of light before through a solution.
(1) Wavelength selection, (2) Printer button, (3) Concentration factor adjustment, (4) UV
mode selector (Deuterium lamp), (5) Readout, (6) Sample compartment, (7) Zero
control (100% T), (8) Sensitivity switch, (9)ON/OFF switch

PRINCIPLE OF COLORIMETER
SYSTEMATIC SEQUENCE OF LIGHT PATH IN ABSORPTIOMETER (COLORIMETER)

• Light travels from source through monochromator

• The monochromatic allows through light of either chosen (filtered) colour or selected
wavelength and then through a dilute coloured solution and happens as follows:

✓ A part of it is reflected.
✓ A part of it is absorbed.
✓ Rest of the light is transmitted onto photocell.
• The photocell converts light energy into electrical energy which is recorded /registered
in OD units or percentage transmission (transmittance).
DIAGRAM OF COLORIMETER

IN PHOTOCELL:

• Transmitted light passes through lacquered later and barrier layer onto selenium layer
• The selenium layer on receiving the light, gets excited and emits electrons which are
thrown back to lacquered layer
• The selenium layer becomes positively charged and lacquered layer becomes negatively
charged
• On connecting the positive and negative poles, through an electrical device such as
galvanometer/digital meter, the passage of the current is indicated by the deflection of
the pointer or digital readings which registers in OD or % transmittance (T).
• The magnitude of the deflection of the pointer is directly proportional to the
transmitted light (light falling on the selenium of the photocell).

UNITS OF COLORIMETRY OR ABSORPTIOMETRY:

OD (optical density), absorbance

T (transmittance), percentage transmission (%T)

INTERCONVERSION OF OD AND T

OD = 2-Log T
Examples:

1. Calculate OD when transmittance reading of the colorimeter is 50%.

Formula = OD = 2-Log T

OD = 2- log 50
= 2-1.6990
= 0.3010
= 0.3

OD = 0.3

2. Calculate the transmittance when the OD reading of the colorimeter is 0.4.

Formula = OD = 2-Log T

OD = 2- log T
0.4 = 2- log T
2 - 0.4 = log T
1.6 = log T
=T

Antilog 1.6 = 39.8 (40%)

NB: In galvanometer, light is recorded in form of:
• Absorbance
• Extinction coefficient
• Optical density

It is a logarithmic scale (from zero to infinity)

Light transmitted is recorded as:

• Transmittance
• Percentage transmission
• % or T
• 0-100%

It is a linear scale (from 0 – 100%)

MANTAINANCE OF EEL PORTABLE ABSORPTIOMETER (COLORIMETER)

1. Keep the instrument clean and dry

2. Switch off after use to prolong life of the lamp
3. Keep it in vibration free bench
4. Weakness of photocell can be detected by fluctuations of the scale pointer and if there
is difficulty in setting zero
5. If the instrument is on , and does not show any light, probable causes:
o Blown out bulb, check and replace
o Damaged fuse, check and replace
o Wire disconnection, do the connection
o Deliberate disconnection of main electricity supply by electricians.

FLAME PHOTOMETER

It facilitates the accurate estimation of potassium and sodium in serum/plasma

PRINCIPLE:

When atoms of certain elements burn in a flame ,the atoms get excited and increase in
energy to a high level , and emit energy in form of characteristic light before attaining their
original state. The light energy emitted is specific in wavelength for each element and is
directly proportional to the amount of atoms being excited. The emitted light is then
quantitated in galvanometer scale connected with photocell.

Examples of elements:
Na+ ions - golden yellow flame
K+ ions - purple, violet flame
Ca2+ ions - brick red flame

Basis of technique:
A dilution of sample is converted to aerosol
Aerosol is mixed with gas used as fuel usually propane
Mixture is ignited in a burner chamber; the heat from the flame releases free atoms from
molecular vapour and increases their energy state. As atoms return to their ground state, they
emit energy in form of light. The emitted light is then quantitated in galvanometer scale
connected with photocell.

PARTS:

• Nebulizer – burner system that consists of a nebulizer (atomizer) , cloud chamber with
condensation vanes and burner.
• Detector system - consists of a monochromator in form of filter and photodetectors fed
to galvanometer.

Precautions:
1. Do not leave instrument running unattended while the flame is alight
2. Never attempt to look down the chimney whilst the flame is running
3. Nebulizer, mixing chamber and burner should be kept clean.
4. Take care when preparing standards
5. Always use recommended spares.

REFRIGERATORS:

Instruments used to remove heat from objects and transfer it outside the refrigerator. To
achieve this objective, three observations should be made:
1. Heat flows from hot area to colder but not vice versa
2. Temperature of boiling depends on the pressure exerted on the surface of the liquid
3. Liquid boiled and converted to vapour absorbs quantity of heat(latent heat of
evaporation)
Parts:
1. Evaporator (coiled metal tube built into the walls of freezing compartment . it is
situated inside the fridge)
2. Condenser (coiled metal tube outside the refrigerator)
3. Refrigerant - liquid easily convertible to vapour (high latent heat)
4. Compressor pump – to pump refrigerant

Uses:
Preserve drinks
Preserve biological specimens
Preserve chemicals

Principle:
Refrigerant is boiled in evaporator (metallic coiled tubes) under low pressure (temperature
below that existing in the refrigerator). It changes to vapour absorbing the heat from air inside
the refrigerator cabinet; leaving the cabinet cool. The vapour is pumped into another coiled
metal tube called the condenser by the use of compressor pump.(maintained at high
pressure)whereby the combined heat from the compressor pump and refrigerator is given off
into air at a temperature higher than that of air outside the refrigerator. When the condenser
loses heat to the surrounding, the refrigerant which is in vapour cools at room temperature
and gets converted into a liquid which is allowed to flow back into the evaporator , where it is
boiled again under low pressure.

Examples of refrigerants:
• Ammonia
• Carbon dioxide
• Sulphur dioxide

Precautions:
o Never place hot liquids inside the refrigerator, this burdens it
o De –frost the freezing compartment regularly 7-10 days
o Never overload the refrigerator, it hinders air circulation
o Do no tamper with the mechanism, if faults develop
o Keep the inside of the refrigerator clean and dry
o Keep the doors of the refrigerator closed unless in use

There are two types of the refrigerator:

o Absorption type – used where quite running is required
o Compression type – (common type) – they make noticeable noise.
AUTOCLAVE:
It is a devise to sterilize equipment and supplies by subjecting them to a high pressure
saturated steam at 1210C or more, typically for 15 – 20 minutes depending on the size of the
load and contents. The name comes from the Greek word “auto” meaning “self” and Latin word
“clavis” meaning self – locking devise. Typical load includes laboratory glassware, surgical
instruments, and medical waste, patient and care utensils. Medical autoclave is a device that
uses steam to sterilise equipment and other objects. This means that all bacteria, viruses, fungi
and spores are destroyed. There are physical, chemical, and biological indicators that can be
used to ensure an autoclave reaches the correct temperature for the correct amount of time.

Principle:
When water is boiled within a closed vessel at increased pressure , the temperature at which it
boils and that of the steam it forms ,will rise above 1000C i.e. temperature increases with
pressure.

Points to note:
1. Admixture of air and steam results in lower temperatures
2. Air hinders penetration of the steam into the interstices of porous materials e.g. surgical
dressing
3. Air being denser than steam tends to form a separate and cooler layer in the lower part
of the autoclave and so prevents adequate heating of the articles.

Parts:
o Vertical or horizontal cylinder of stainless steel
o Lid fastened by screw clamps
o Cylinder contains water to certain level and is heated by gas or electric heater
o Lid is fitted with discharge tap for air and steam pressure gauge and safety valve (blow
off at desired pressure)
o Perforated tray placed above water level
Operation:
1. Check whether water level is sufficient
2. Insert material
3. Place lid, open discharge tap (air outlet)
4. Adjust safety valve to required pressure e.g. 15psi (pounds per square ins) at 121 0C
5. Turn on the heater
6. Steam rises from boiling water, mixes with air and escapes through discharge tube, let it
continue until all air is eliminated
7. Close discharge tap
8. Steam pressure rises until desired level 15lb/ins for 1210C when safety valve opens and
allows excess steam to escape
9. At this point, begin timing (15 -20 minutes)
10. Turn off heater and allow autoclave to cool until pressure gauge reads 0 lbs/ins
11. Open discharge tap slowly to allow air to enter the autoclave

Precautions:
When discharge tap is opened at high pressure, liquid media will boil violently and spill from
the container
Two types of autoclaves:
o Non –jacketed autoclave
o Steam - jacketed

Maintenance:
1. Do not use autoclave if it defective
2. Use time, steam and temperature (TST) controls to check performance
3. Check regularly for signs of wear and damage
4. Clean inside after use and around the valve and stop cocks
5. When trouble shooting refers to operation manual
6. If using electricity mains, protect from power surge by using a voltage stabilizer
7. If the instrument is faulty, consult a qualified biomedical engineer.
8. Don’t overload the autoclave, make sure there is sufficient space inside the autoclave.
9. Never open the lid when the autoclave is working.

DE-IONIZERS

Devise for preparing de-mineralized water (de-ionized water).

There are two most common types of deionization
Two – bed deionization
Mixed – bed deionization

1. Two – bed deionization

Consists of two vessels , one containing a cation – exchange resin in the hydrogen (H+)
form and the other containing an anion resin in hydroxyl(OH-) form
2. Mixed bed deionization
In mixed - bed deionizers, the cation exchange and anion – exchange resins are
intimately mixed and contained in a single pressure vessel.

Principle:
In context of water purification, process of de-ionization or ion – exchange is a rapid and
reversible process in which impurity ions present in water are replaced by ions released
by ion exchange resin. There are two basic type of resin: cation - exchange and anion -
exchange resins. Cation exchange resins releases hydrogen (H+ ) ions or positively
charged ions in exchange for impurity cations present in water. Anion – exchange resins
will release hydroxyl (OH-) ions or other negatively charged ions in exchange for
impurity anions present in water.
Cation exchange resins(R-S03) - H+ which are insoluble acids. R – Represents: polystyrene
resin
Anion - exchange resin (R NH3)+ OH- which are insoluble bases.
 Process:
When impure water passes through a de-ionizer and possibly contains ( Na+Cl-), cations
in water are removed at cation - exchange resin, while the anions are removed at the
anion – exchange resin. The emergent water is called de-ionized water.

Impure water with (Na+Cl-) cation - exchange resin (R-SO3)- H+ +Na+ (R-SO3) Na++
+
H
Anion – exchange resin (R –NH3) + OH- + Cl- (R-NH3)+ Cl- + OH-

H+ + OH H20 (ion free)

NB:
• Na+ cations replaces H+ cations in cation exchange resin.
• In anion – exchange resin, Cl – anions replaces OH- anions of the resin.

Impure water Cation exchange Anion exchange resin De – ionized

resin water

Cations which may be present in water:

Calcium

Magnesium are exchanged by the anion resin which in turn release OH-

Sodium

Anion impurities:

Sulphate

Bicarbonate

Silica are exchanged by the anion resin which in turn release OH-

Nitrate

Chloride

Parts:

• Elgacan cartridge – for ion exchange resin

• Meter to check for conductivity
• Battery – power supply
NB:

• Ion exchange resin may be regenerated by passing HCL through the cation exchange resin and
NAOH through the anion exchange resin, followed by in both cases by thorough wash with
water.
• It is not guaranteed for biological purity i.e. it is not sterilized which means not free from micro –
organisms.
• Has low conductivity, it means amount of ions present is minimal or neglible hence low needle
deflection.
• It is used for chemical purposes

WATER DISTILLER.

A water distiller is a machine that purifies water by removing more than 99.9% of
contaminants, including chemicals, heavy metals, microorganisms and sediment. While
design may vary, a typical water distiller consists of a boiling chamber, a cooling
system and a separate storage tank.

PRINCIPLE:

Distillers work on the principle of evaporation and condensation. An electric heating

element boils water in a stainless steel tank. The resulting steam leaves the tank and
enters a stainless steel cooling coil. In the cooling coil, the steam condenses to form
distilled water.
 The Water Distillation Process

Distillation effectively removes contaminants such as suspended particles, infective

organisms, dissolved metals and solids, chemicals and industrial solvents and
VOCs to give you sparking clean, pure, crystal clear water.

1. Ordinary tap water is heated to 100 C to start the evaporisation

process.
2. The rising steam is condensed in a stainless steel coil by the cooling
fan to turn the steam into pure water.
 3. Volatile gases that may be present during condensation are released
into the atmosphere through the gas vent.
4. The distilled water then percolates through an activated carbon filter to
remove any remaining traces of dissolved gas and odours.
5. The purified water is collected in a reservoir tank.
6. At the end of each distillation process, all the impurities and
contaminants (also called precipitates) are left behind in the stainless
steel boiler tank.

What Contaminants Does a Water Distiller Remove?

A water distiller removes a broad range of contaminants, including organic compounds,

heavy metals like lead, chemicals like chlorine, and microorganisms like bacteria,
hardness minerals, dissolved salts, and almost every other impurity commonly found in
drinking water.

What makes the distillation process unique is that it can remove contaminants of all
sizes, from tiny viruses to large particles of suspended sediment.

There are a couple of contaminants that can convert to gas with water, namely benzene
and (volatile organic compounds) VOCs. Most distillers use a small activated
carbon filter at the spout, which removes these contaminants as water drips down into
the holding container.

What is Distilled Water Used For?

At home, distilled water can be used for topping up steam irons, adding to car batteries,
filling aquariums and cleaning.

Distilled water is also widely used industrially, medically and scientifically.

How to Maintain a Water Distiller

1. Clean the system regularly.

2. Replace the activated carbon filter when necessary to ensure it can work at its
optimum.
3. It is recommended to clean your distiller’s boiling chamber after every distillation
cycle for efficient distillation performance.
WATER BATH:
A water bath is a lab constant temperature equipment, providing heat source for varieties of devices
that need heating. The circulating water bath is used to keep water at a constant temperature.
It is a tool that is used to maintain a very stable temperature much like an incubator. They are made to
maintain water at a required temperature. They are used particularly for serological work (antibody
/antigen reactions). They have thermometers and others have rotors to mix the water. They perform
laboratory tests at 370C. They are electrically heated and have thermostat to control the temperature.
They do maintain uniform temperature.

Parts:

▪ A water bath generally consists of a heating unit,

▪ a stainless-steel chamber that holds the water and samples,
▪ Are equipped with thermostatic controls
▪ Lids are included to prevent excessive heat loss and evaporation.
TYPES OF WATER BATH:

1. Circulating water baths

2. A shaking water bath in action

Circulating water baths:

Also called stirrers, are ideal for applications when temperature uniformity and consistency are critical,
such as enzymatic and serologic experiments. Water is thoroughly circulated throughout the bath
resulting in a more uniform temperature.

Non-circulating water baths:

This type of water bath relies primarily on convection instead of water being uniformly heated.
Therefore, it is less accurate in terms of temperature control.

Shaking water baths:

This type of water bath has extra control for shaking, which moves liquids around. This shaking
feature can be turned on or off. In microbiological practices, constant shaking allows liquid-grown
cell cultures grown to constantly mix with the air.

WORKING PRINCIPLE:

: The sensor transfer water temperature to resistance value, amplified and compared by integrated
amplifier, then output the control signal, efficiently control the average heating power of electric
heating tube and maintain water in constant temperature. When you require balanced high
temperature heating that, water bath is a good choice.

Operating the Water Bath:

1. Connect the power supply.

2. Ensure the water level in water bath is sufficient to pour the heating element.
3. Switch “ON” the main power supply and instrument mains.
4. For temperature settings, Press SET key to set the required temperature. press ↑ to increase
the temperature and ↓ to reduce the temperature
5. The temp. Sensor will maintain the set temp. During use of water bath.
 6. Switch “OFF” the instrument mains & main power supply after use.

MAINTENANCE:

1. Before incubating a material, always check the temperature and recommended water level
should be maintained above the level of whatever is being incubated.
2. If empty don’t switch on the water bath
3. Clean the water bath regularly taking care not to damage the heating unit and the thermostat.
4. Unplug the water bath from the wall socket when not using it
5. Use de-ionized water or distilled water
6. Protect the instrument from the power surge by using a voltage stabilizer
7. Standard operating procedures of the system should always be displayed for efficient
management of the system.

BACTERIOLOGICAL INCUBATOR:
An incubator, in microbiology, is an insulated and enclosed device that provides an
optimal condition of temperature, humidity, and other environmental conditions
required for the growth of organisms.
An incubator can be used to cultivate both unicellular and multicellular organisms. It is a
device used to grow and maintain microbiological cultures or cell cultures. It maintains optimal
temperature, humidity and other conditions such as CO2 and O2 content of the atmosphere
inside.
The simplest incubators are insulated boxes with an adjustable heater typically going upto60 0C
– 650C(1400F 1500F).
The most commonly used temperature both for bacteria such as frequently used E. coli as well
as mammalian cells is approximately 370C.

Most incubators include timers. Some incubators are single – jacketed and others double
jacketed. Single jacketed contain dry air which is heated and transmits heat to the metal cover.
Double jacketed ones contain water which is heated to provide heat. Double ones are reliable.
Temperature
Components/Parts of Incubator
A microbial incubator is made up of various units, some of which are:
Cabinet
• The cabinet is the main body of the incubator consisting of a double-walled cuboidal
enclosure with a capacity ranging from 20 to 800L.
• The outer wall is made up of stainless steel sheets while the inner wall is made up of
aluminum.
• The space between the two walls is filled with glass wool to provide insulation to the
incubator.
• The insulation prevents heat loss and reduces electric consumption, thereby ensuring
the smooth working of the device.
• The inner wall of the incubator is provided with inward projections that support the
shelves present inside the incubator.
Door
• A door is present in all incubators to close the insulated cabinet.
• The door also has insulation of its own. It is also provided with a glass that enables the
visualization of the interior of the incubator during incubation without disturbing the
interior environment.
• A handle is present on the outside of the door to help with the maneuvering of the
door.
Thermostat
• A thermostat is used to set the desired temperature of the incubator.
• After the desired temperature is reached, the thermostat automatically maintains the
incubator at that temperature until the temperature is changed again.
Perforated shelves
• Bound to the inner wall are some perforated shelves onto which the plates with the
culture media are placed.
• The perforations on the shelves allow the movement of hot air throughout the inside
of the incubator.
• In some incubators, the shelves are removable, which allows the shelves to be cleaned
properly.

Asbestos door gasket

• The asbestos door gasket provides an almost airtight seal between the door and the
cabinet.
• This seal prevents the outside air from entering the cabinet and thus, creating an
isolated hot environment inside the cabinet without being interrupted by the external
environment.

Thermometer
• A thermometer is placed on the top part of the outer wall of the incubator.
• One end of the thermometer provided with gradations remains outside of the
incubator so that temperature can be read easily.
• The next end with the mercury bulb is protruded slightly into the chamber of the
incubator.
 Principle/ Working of Incubator
• An incubator is based on the principle that microorganisms require a particular set of
parameters for their growth and development.
• All incubators are based on the concept that when organisms are provided with the
optimal condition of temperature, humidity, oxygen, and carbon dioxide levels, they
grow and divide to form more organisms.
Procedure for running an incubator
Once the cultures of organisms are created, the culture plates are to be placed inside an
incubator at the desired temperature and required period of time. In most clinical
laboratories, the usual temperature to be maintained is 35–37°C for bacteria.
The following are the steps to be followed while running an incubator:
1. Before using the incubator, it should be made sure that no remaining items are
present in the incubator from the previous cycles.
2. The door of the incubator is then kept closed, and the incubator is switched on. The
incubator has to be heated up to the desired temperature of the growth of the
particular organism. The thermometer can be used to see if the temperature has
reached.
3. In the meantime, if the organism requires a particular concentration of CO 2 or a
specific humidity, those parameters should also be set in the incubator.
4. Once all the parameters are met, the petri dish cultures are placed on the perforated
shelves upside down, i.e., media uppermost.
5. Now, the door is locked, and the plates are kept inside for the required time before
taking them out.
Types of incubators
Incubators are divided into the following types:
Benchtop incubators
• This is the most common type of incubator used in most of the laboratories.
• These incubators are the basic types of incubators with temperature control and
insulation.
CO2 incubators
• CO2 incubators are the special kinds of incubators that are provided with automatic
control of CO2 and humidity.
• This type of incubator is used for the growth of the cultivation of different bacteria
requiring 5-10% of CO2 concentration.
Cooled incubators
• For incubation at temperatures below the ambient, incubators are fitted with modified
refrigeration systems with heating and cooling controls.
• This type of incubator is called the cooling incubator.
• In the cooling incubator, the heating and cooling controls should be appropriately
balanced.
Shaker incubator
• A thermostatically controlled shaker incubator is another piece of apparatus used to
cultivate microorganisms.
• Its advantage is that it provides a rapid and uniform transfer of heat to the culture
vessel, and its agitation provides increased aeration, resulting in acceleration of
growth.
• This incubator, however, can only be used for broth or liquid culture media.

Portable incubator
• Portable incubators are smaller in size and are used in fieldwork, e.g. environmental
microbiology and water examination.
Mantainance:
1. Keep it clean regularly and Service it regularly (3- 6 months)
2. As microorganisms are susceptible to temperature change, the fluctuations in
temperature of the cabinet by repeatedly opening the door should be avoided.
3. The required parameters growth of the organism should be met before the
culture plates are placed inside the cabinet.
4. The plates should be placed upside down with the lid at the bottom to prevent
the condensation of water on to the media.
5. The inside of the incubators should be cleaned regularly to prevent the
organisms from settling on the shelves or the corners of the incubator.
6. Protect from power surge by using a voltage stabilizer
7. If faulty, consult a qualified biomedical engineer
8. Standard operating procedures of the system should always be displayed for efficient
management of the system

HOT – AIR OVEN:

• A hot air oven is a laboratory instrument that uses dry heat to sterilize laboratory
equipment and other materials.
• Some examples of material which cannot be sterilized by employing a hot air oven such as
surgical dressings, rubber items, or plastic material.
 • We can sterilize Glassware (like petri dishes, flasks, pipettes, and test tubes), Powder (like
starch, zinc oxide, and sulfadiazine), and Materials that contain oils, Metal equipment
(like scalpels, scissors, and blades) by using hot air oven.
• To destroy microorganisms and bacterial spores, a hot air oven provides extremely high
temperatures over several hours.
• The widely used temperature-time relationship in hot air ovens to destroy microorganisms
are 170 degrees Celsius for 30 minutes, 160 degrees Celsius for 60 minutes, and 150
degrees Celsius for 150 minutes.
• They have thermostat to control the temperature
• Their double walled insulation keeps the heat in and conserves the energy, the inner
layer being a poor conductor and outer layer being metallic.
Advantages:
• Can be converted to incubators (bacteriological incubator)
• They have a wide range of temperature
• They do not require water

Working Principle:

Sterilization by dry heat is performed by conduction. The temperature is consumed by the surface
of the objects, then moves towards the core of the object, coating by coating. The whole object
will ultimately attain the temperature needed for sterilization to take place.

Dry heat causes most of the injury by oxidizing particles. The primary cell components are
damaged and the organism dies. The temperature is kept for about an hour to eliminate the most
ambitious of the resistant spores.

Hot air oven Parts and Functions

1. External cabinet: The External cabinet is made of stainless sheets. It covers the inner
chamber.
2. Glass wool insulation: The space between the inner chamber and external cabinet is
filled with Glass wool. It provides insulation to the hot air oven.
3. Inner chamber: The inner chamber of the hot air oven is made of Stainless steel.

4. Tubular air heaters: They help to generate heat within the inner chamber. Two Tubular
air heaters are located on both sides of the inner chamber.
5. Motor-driven blower: It helps in uniformly circulating the air within the chamber.

6. Temperature sensor: It measures the temperature within the hot air oven and displays
it on the controller screen.
7. Tray slots: The inner wall of the chamber contains several try slots that hold the trays.

8. PID temperature controller: It maintains the accurate temperature during the entire
cycle. It also controls the temperature and also displays the temperature values.
9. Load indicator: it indicates the hot air oven is overloaded.

10. Mains on/off switch: It helps to turn on/ turn off the hot air oven.
 11. Safety thermostat: It is also known as an over-temperature protection device. It keeps
your oven and specimen safe in case of controller malfunction.

Maintenance:
1. Clean the oven regularly
2. Service it regularly
3. Check the required temperature before use
4. Protect from power surge by using a voltage stabilizer
5. If faulty consult a qualified biomedical engineer
6. Standard operating procedures of the system should always be displayed for efficient
management of the system

Common functional parts for water bath, hot –air oven & incubator:
• Thermostat
• Heating element

LABORATORY CENTRIFUGE:

It is a piece of laboratory equipment, driven by a motor, which spins liquid samples at a high
speed. There are various types of centrifuges, depending on the size and the sample capacity.
Like all other centrifuges, laboratory centrifuge work by the sedimentation principle where the
centripetal acceleration is used to separate substances of greater and lesser density.

PRINCIPLE OF CENTRIFUGATION:

• In a solution, particles whose density is higher than that of the solvent sink (sediment),
and particles that are lighter than it floats to the top.
• The greater the difference in density, the faster they move. If there is no difference in
density (isopycnic conditions), the particles stay steady.
• To take advantage of even tiny differences in density to separate various particles in a
solution, gravity can be replaced with the much more powerful “centrifugal force”
provided by a centrifuge.
• A centrifuge is a piece of equipment that puts an object in rotation around a fixed axis
(spins it in a circle), applying a potentially strong force perpendicular to the axis of
spin (outward).
• The centrifuge works using the sedimentation principle, where the centripetal
acceleration causes denser substances and particles to move outward in the radial
direction.
• At the same time, objects that are less dense are displaced and move to the center.
• In a laboratory centrifuge that uses sample tubes, the radial acceleration causes denser
particles to settle to the bottom of the tube, while low- density substances rise to the
top

PARTS:
• Head of centrifuge, contains containers called buckets
• The head is connected to a rotor which holds it
• Rotor is controlled by a motor
• Centrifuge is covered from outside by metal casing to control atmospheric pressure thus
creating a partial vacuum and friction is lessened.

How to Operate a Centrifuge

Often found in chemistry, biochemistry, and hematology labs, centrifuges are expensive lab
instruments but are fairly easy to operate. All you have to do is the following:

1. Place the centrifuge in its operating place, preferably a very stable lab
workbench. Open the centrifuge lid. Keep the power off for now.
2. Perform basic cleanup as per the manufacturer’s instructions.
3. Put your liquids in centrifuge tubes (designated to fit snugly in the machine you
are using) and put on the caps properly.
4. Insert the tubes in the boreholes on the platform. If you have less number of
tubes than there are holes (the common situation), try to distribute them evenly
around the platform so that the spin can stay even.
 5. Close the lid and turn on the power to the device. Operate the controls to start
the spin. The controls may be different from model to model, you had better
consult the instruction manual first.
6. If the device supports it, you may want to set a timer beforehand.
7. When the spin stops (manually or automatically), let it stay still for a few minutes.
8. Now collect the tubes from the platform one by one, very carefully not to shake
them. Put them in a designated centrifuge tube rack .
9. At the end of the workday, make sure to perform regular cleaning and
maintenance for the lab centrifuge machine.

Centrifuges fall under two categories:

1. Angle – head
2. Swing – out

Angle – head type of centrifuge:

1. They deposit suspended material at an angle in the centrifuge tube
2. When rotated, the buckets remain at a fixed angle or stationary
3. Time taken by angle head to form a deposit is shorter than that taken by a swing out
type
4. Particles travel at a shorter distance in angle – head centrifuge
5. Pipetting a clear supernatant from an angle head is not easy because the deposit get
easily disturbed due to formation pattern.

Swing – out type of centrifuge:

1. They deposit suspended material at the tip of the tube
2. When rotated, the buckets are observed to swing and spread horizontally.
3. Time taken by swing out to form a deposit is longer than that taken by a angle – head
type
4. Particles travel at a longer distance in swing out type of centrifuge.
5. Pipetting a clear supernatant from swing out type of centrifuge is easier.
It should be noted that, the rate of centrifugation is specified by the acceleration applied to the
sample, typically measured in revolutions per minute (RPM) or relative centrifugal force (RCF).

Types of centrifuge:

Hand centrifuge
• Has 2-4 buckets and manually operated
• Used for simple elementary work where there is no electricity
• It is a swing out
• Does not give a complete separation
• Have no speed knob

Ordinary centrifuge
 • Can be either a swing out or angle – head
• Uses electricity
• Can be 2 – bucketed or multi -bucketed.
• They have speed knobs
• They may have timing knobs

Specialized centrifuges
• Used in research labs or advanced laboratories
• Very expensive
• Can be either a swing out or angle – head
• Maintains temperature and pressure
• Can be connected with alarms
• Are connected with timers and speed knobs

MANTAINACE:
1. Clean the centrifuge and tube holders weekly
2. Do not centrifuge at higher speed
3. Always ensure that centrifuge tubes are balanced
4. Avoid stopping the machine mechanically as this practice wears out the brushes and can
cause injury
5. Rubber cushions should be checked whether they are in place before spinning
6. In case of a mechanical problem consult a qualified biomedical engineer
7. Standard operating procedures of the system should always be displayed for efficient
management of the system

WHEIGHING BALANCES:

A weighing balance is an instrument which is used to determine the weight or mass of an

object. Available in a wide range of sizes with multiple weighing capacities they are essential
tools in laboratories, commercial kitchens and pharmacies.

Balances are instruments used to determine weights of objects and

substances.

PRINCIPLE:

A weighing balance operates through gravity comparing the torque of the weighed object with a
known mass.

Balance and Scale Terms

Accuracy The ability of a scale to provide a result that is as close as possible to the actual value.
Calibration The comparison between the output of a scale or balance against a standard value.
Usually done with a standard known weight and adjusted so the instrument gives a reading in
agreement.
Capacity The heaviest load that can be measured on the instrument.
Precision Amount of agreement between repeated measurements of the same quantity; also
known as repeatability.

Readability This is the smallest division at which the scale or balance can be read. It can vary as
much as 0.1g to 0.0000001g. Readability designates the number of places after the decimal
point that the scale can be read.
Tare The act of removing a known weight of an object, usually the weighing container, to zero a
scale. This means that the final reading will be of the material to be weighed and will not reflect
the weight of the container.

Calibration is another care issue when it comes to scales. A scale cannot be accurate
indefinitely; they must be rechecked for accuracy.

Types of Balances and Scales

1. Analytical balance:
These are most often found in a laboratory or places where extreme sensitivity is needed for
the weighing of items. Analytical balances measure mass. Chemical analysis is always based
upon mass so the results are not based on gravity at a specific location, which would affect the
weight.

Generally capacity for an analytical balance ranges from 1 g to a few kilograms with precision
and accuracy often exceeding one part in 106 at full capacity.

Parts of the Analytical Balance

Analytical balances on the market today are generally classified into two different types: digital
analytical balances and analog analytic balances. Discussions on the sections on a digital
analytical balance:

1. Balance plate (pan)

Used as a container to put the sample of material to be measured in mass. To clean this area, you
can use a special cleaning brush or a clean microfiber cloth.

2. Weights
Serves as a tool to calibrate analytical scales. The mass of the material being the weights is
precisely made so that the balance calibration remains accurate.

3. Water pass
A tool used to determine the position of the balance plate. Water pass is also used to re-position
the balance plate,

4. Power button (on / off button)

Used to turn the balance on or off. After pressing the power button to turn on the balance, the
instrument will usually be left idle for approximately 10-15 minutes before using it to produce a
more accurate measurement.

5. 'Re-zero' or 'Tare' button

Used to adjust the balance to return to a neutral position (zero). We recommend that you use
this button not too often so that the balance can still produce accurate measurements.

6. 'Mode' button
Used to set the conversion system used when measuring. By using the 'Mode' button, you can specify
the conversion system as needed.

Advantages of Analytical Balance

Analytical balances are generally used to measure the mass of objects with a high degree of
precision (sub-milligram range). This means that the analytical balance function is able to display
more detailed measurement results up to sub-milligram units.
Usually, analytical balances are used to measure the mass of materials or objects that are solid.
However, it is also possible if this instrument is used to measure materials or objects in the form
of liquids. Apart from having high accuracy, the analytical balance is also strong. Analytical
balance can be used in free space without having to fear the sample is exposed to air flow. This
is because the analytical balance has a special cover that protects the sample from airflow. Not
only that, the analytical balance is also equipped with an internal calibration system so that the
balance can be identified and adjusted easily.

Level of Accuracy Analytical Balance

Analytical balances, especially digital types, have a high degree of accuracy. What is the level of
accuracy of the analytical balance? This instrument is capable of displaying the measurement
results of up to four decimal places. Most digital analytical balances on the market are capable
of detecting the mass of objects in the range of 100 grams to approximately 0.0001 grams.

2. Equal Arm Balance/Trip Balance

This is the modern version of the ancient Egyptian scales. This type of
laboratory scale incorporates two pans on opposite sides of a lever. It can be used in two
different ways. The object to be weighed can be placed on one side and standard weights are
added to the other pan until the pans are balanced. The sum of the standard weights equals the
mass of the object.
Another application for the scale is to place two items on each scale and adjust one side until
both pans are leveled.

3. Platform Scale This type of scale uses a system of multiplying levers. It allows a heavy object
to be placed on a load bearing platform. The weight is then transmitted to a beam that can be
balanced by moving a counterpoise, which is an element of the scale that counterbalances the
weight on the platform. This form of scale is used for applications such as the weighing of
drums or even the weighing of animals in a veterinary office.

4. Spring Balance This type of balance utilizes Hooke's Law which states that the stress in the
spring is proportional to the strain. Spring balances consist of a highly elastic helical spring of
hard steel suspended from a fixed point. The weighing pan is attached at the lowest point of
the spring.
An example of this type of balance would be the scale used in a grocery store to weigh produce.

5. Top-Loading Balance: This is another balance used primarily in a laboratory setting. They
usually can measure objects weighing around 150–5000 g. They offer less readability than an
analytical balance, but allow measurements to be made quickly thus making it a more
convenient choice when exact measurements are not needed.
Top-loaders are also more economical than analytical balances. Modern top-loading balances
are electric and give a digital readout in seconds.
6. Torsion Balance Measurements are based on the amount of twisting of a wire or fiber. Many
microbalances and ultra-microbalances, that weigh fractional gram values, are torsion balances.
A common fiber type is quartz crystal.

7. Triple-Beam Balance This type of laboratory balance is less sensitive than a top-loading
balance. They are often used in a classroom situation because of ease of use, durability and
cost. They are called triple-beam balances because they have three decades of weights that
slide along individually calibrated scales. The three decades are usually in graduations of 100g,
10g and 1g.

DIFFERENCE BETWEEN MASS AND WEIGHT.

What Is Mass?

Mass is a constant unit of the amount of matter an object possesses. It stays the same no
matter where the measurement is taken.
The most common units for mass are the kilogram and gram.
What Is Weight?

Weight is the heaviness of an item. It is dependent on the gravity on the item multiplied by the
mass, which is constant.
Unit of measurement for weight is the newton. A newton takes into account the mass of an
object and the relative gravity and gives the total force, which is weight.
Although mass and weight are two different entities, the process of determining both weight
and mass is called weighing.

MAINTENANCE:

1. Weights and balances must be kept in places free of dust and dry.
2. You should never weigh chemicals in bare pans, can react with the metal in the pan
and corrode the surface. This will affect the accuracy of the scale
3. The balances should always in be in zero position if not in use.
4. Don’t handle weights in bare fingers, use forceps.
5. Always place balances on firm benches to avoid vibrations and should not share
benches with centrifuges.
6. Items to be measured should be at room temperature before weighing.
 7. A hot item will give a reading less than the actual weight due to convection currents
that make the item more buoyant.
8. Balances should always be kept clean.

HYDROMETERS:
It is an instrument used to measure the specific gravity (or relative density) of fluids. That is the
ratio of the density of the liquid to the density of water.

it operates on the basis of Archimedes principle of floatation, that solid suspended in a fluid
will be buoyed up by a force equal to the weight of the fluid displaced .the lower the S.G the
deeper the instrument floats; S.G readings are taken from the stem below the meniscus .

Hydrometers are calibrated to suit the variation limits of the S.G. of the fluids for which they
are designed to measure. e.g. Hydrometers for urine called urinometers; have ranges 1.000 –
1.060; milk – are called lactometers have range 1.029 – 1.033; saccharometer – measuring the
density of sugar in a liquid; alcoholmeter – measuring higher levels of alcohol in spirits.

Parts:
Hydrometer is made of glass and consists of cylindrical stem and a bulb weighted with mercury
or lead shot to make it float upright.

Operation:
Liquid to be measured is poured into a tall container of graduated cylinder, and hydrometer is
gently lowered into a liquid until it floats freely.

Hydrometers contain a scale inside the stem, so that the specific gravity can be read directly.
High floatations of hydrometers shows that the density of liquid is high, when it has low
floatation shows that the density of the liquid is low.

Maintenance:
• Handle the instrument carefully
• Before a test is made the hydrometer should be thoroughly washed, rinsed and dried by wiping
with a clean, lint free cloth.
• The hydrometer jar should be thoroughly washed and rinsed before the clean test liquid is
added.

PH METER:

A PH meter is an electric instrument to measure the PH (acidity or alkalinity) of a liquid. Have a

range of PH 0.1 – 14.0. it is designed to measure the concentration of hydrogen ions in a
solution.
Principle:
It is based on difference in electrical potential and could be measured between two
solutions of different PH separated by a thin glass membrane. The potential thus
produced varies with the hydrogen ion concentration of the two solutions. i.e., the
glass membrane is H+ ion sensitive. The potential difference is measured in millivolts,
which on the PH meter is calibrated both in millvolts and PH. Measurement of PH is
possible only when glass electrode (standard) is coupled with a calomel electrode
(reference). Glass electrode functions like a semi permeable membrane, permeable
only to hydrogen ions which enter the lattice of the glass (i.e. selective for H+ions
only). A calomel (mercurous chloride) reference electrode consists of mercury in
contact with a solution of potassium chloride saturated with calomel. it is
surrounded by an outer vessel containing saturated potassium chloride which acts
as salt bridge between the reference and test solution (this electrode is not
dependent upon PH).

MANTAINANCE:
1. PH meter should be kept in a cool place
2. Electrodes should be kept safely preferably in water constantly
3. Avoid touching the electrodes especially standard electrode
4. Protect electrodes especially standard against corrosive materials e.g.
concentrated acids
 5. Avoid highly proteinous materials, if possible de-proteinze before measuring.
6. Check PH meter with a buffer
7. Avoid taking PH of hot materials
8. Remember to rinse the electrodes thoroughly after being immersed in a
solution

Macintosh Felds jar

It is used for growing anaerobic bacteria it excludes free O2 to suit the growth of those
bacteria.

It has the following components:

• Hollow jar
• The lid
• The clamp
• Indicator
• Catalyst
• Rubber

Hollow jar: where Petri dishes with bacterial growth are kept

The lid: Have 2 outlets (valves), one to extract the air from the jar by connecting it to vacuum
pump.
Other outlet should be closed (second one)
Second one (outlet) is connected to H2 supply and due to vacuum created, H2 is drawn in and
then outlet is closed.

The clamp: it holds the lid tight.

The indicator: to indicate whether anaerobic condition is achieved in the jar, it changes its
colour; colourless in absence of O2 and blue in presence of O2.
Catalyst (alumina pellets coated with 0.5% palladium): a metal catalyst that accelerates
combination of O2 and H2 left to form H2O.

Rubber at the mouth of the jar: to make air tight.

Principle:
McIntosh and Fildes’ anaerobic jar works on the principle of evacuation and
replacement, where the air inside the chamber is evacuated and replaced with mixture
of gases (consisting of 5% CO2, 10% H2, and 85% N2).

Procedure:

1. Place the inoculated plates in anaerobic jar

2. Clamp the lid in position
3. Close the inlet valve
4. Connect the outlet valve to the vacuum pump and suck out the air until manometer
indicates that there is a negative pressure in jar about 30 – 40mm of mercury.
5. Close the valve and disconnect the pump.
6. Attach the inlet valve to a low pressure source of H 2
7. Open the inlet valve to allow the gas to enter for 3 minutes
8. Close the inlet valve, disconnect the jar and place it carefully in the incubator.
9. After incubation & before opening the jar, check that the chemical indicator is colourless
(methylene blue)
10. Open the jar by unscrewing one of the valves to allow the air to enter.
11. Then remove the lid & read your cultures.
MANTAINANCE:
1. Ensure the catalyst is dry and fully activated
2. Methylene blue indicator should be checked for any drying of the medium
3. Should be kept clean
4. Washer around the rim of the jar should be inspected regularly for wear.

BIOSAFETY CABINET:

A biosafety cabinet (BSC), biological safety cabinet, or microbiological safety cabinet is an

enclosed ventilated work space for safely working with materials contaminated with (or
potentially contaminated with) pathogens in the laboratory. The primary purpose of a BSC is to
serve as the primary means to protect the laboratory worker and the surrounding environment
from pathogens. All exhaust air is HEPA – filtered as it exits the biosafety cabinet, removing
harmful bacteria and viruses.

The US centres for disease control and prevention (CDC) classifies the BSCs into 3 classes:
Class I, Class II, and Class III

Class I and II cabinets are used for diagnostic and containment laboratories for the work with
risk group 3 organisms; they prevent high risk to laboratory worker but low risk to community.
E.g. TB
Class III cabinets are used almost exclusively for risk group 4 viruses (all viruses – offer high risk
to the laboratory worker and to the community).

CLASS I SAFETY CABINET:

Operator sits at the front of the cabinet, looks through the glass screen and works with hands
inside. Any aerosols released from cultures or other infectious material is retained because a
current of air passes in at the front of the cabinet and sweeps the aerosols up through filters
which remove all or most of the organisms. Clear air then passes through the fan, which
maintains the air flow and is discharged to atmosphere where any particles or organisms not
retained on the filter are so diluted that they are no longer likely to cause infection if inhaled.
CLASS II SAFETY CABINET:

Sometimes called laminar flow cabinet. The term is used for clean air cabinets which do not
protect the worker and should be avoided. 70% of the air is re- circulated through filters so that
the working area is bathed in clean air. The air flow caries along any aerosols produced in the
course of the work and these are removed by the filters. Some of the air about 30%is
discharged to the atmosphere and is replaced by a “curtain “of room air which enters the
working face. This prevents the escape of any particles or aerosols released in the cabinet.

CLASS III SAFETY CABINET:

It is for risk group 4 viruses. It is totally enclosed and is tested under pressure to ensure that no
particles can leak from it into the room. The operator works with gloves which form part of the
cabinet. Air enters through a filter and is exhausted to atmosphere through one or two more
filters.
MANTAINANCE:
1. Manufacturer’s instructions should be studied carefully
2. Safety cabinets should be swabbed pout with a suitable disinfectant after use e.g.
glutaradehyde
3. Should be regularly decontaminated with formaldehyde
4. Should not be loaded with unnecessary equipment or it will not carry its job properly
5. The operator should avoid bringing the hands and arms out of the cabinet while working
6. Wash hands immediately after finishing work in safety cabinets.