Nephelometry and Turbidimetry
Introduction:
Nephelometry is the measurement of scattered light. This technique
requires a special measuring instrument, where the detector is set at an angle
to the incident light beam.
Turbidimetry is the measurement of unscattered light, i.e. light transmitted
through a turbid solution. This method of measurement is carried out using
an absorptiometer (photometer)
Nephelometry:
The intensity of scattered light can be approximately related to the
concentration of suspended particles.
In practice, light scattered at right angles to the incident beam is plotted
against concentration. Light scattering intensity units have been used for
some nephelometric methods in clinical chemistry laboratories. These units
relate to standardised suspensions of polystyrene latex particles which are used
to calibrate the instrument.
In certain nephelometers, low power lasers are sometimes used as light sources.
Light is passed through a lens system and a filter into a turbid solution
contained in a cuvette and the resultant scattered light is measured using a
photo detector. Commonly an angle of 90 between the incident and reflected
light is used for detection purposes, but the optimal angle depends on the ratio
between the particle size and the wavelength.
Turbidimetry:
In a solution containing particles in suspension, the incident light beam is
diminished due to scattering. Turbidity is based on measuring the
transmitted light which is related to the concentration.
The proportionality constant depends on the particle size and shape, the
wavelength of the incident light and the refractive index of both the suspending
medium alone and the suspending medium containing particles. This
expression is only valid for small particles.
In turbidimetry, consideration must be given to the question of colour of the
final solution. If the solvent and dispersed particles are both free from
colour then a wavelength in the blue or near ultraviolet should be selected for
maximum sensitivity. If colour is present, a coloured filter must be used.
If a blue filter is used with a red solution a certain amount of the light
will be absorbed resulting in false measurement.
Precautions:
To make measurements reproducible it is important that the particle size is
maintained constant. therefore the following items should be considered:
1. The relative concentration of ingredients must be controlled.
2. The relative ratios of reactants must be constant.
3. The manner, order of addition and rate of mixing must be constant.
4. The nature of other solutions present must be considered and, if
necessary, protective colloids added to avoid protein precipitation.
5. the temperature must be maintained constant.
6. Constant time lapse before reading.