Colorimeter and Fully Auto Analyzer - Group H

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RINCIPLE OF COLORIMETER

AND SPECTOPHOTOMETER AND


VARIOUS TYPE OF ANALYSER
USED IN CLINICAL
BIOCHEMISTRY
COLORIMETER
• What is colorimeter ?
• Colorimetry.
• Principle of colorimeter.
• Beer's and Lambert's law.
• Components of colorimeter.
• Functions of components.
• Advantages and Disadvantages of single cell
photometry.
What is colorimeter ?

meter is a instrument used for the measurement of


ed substance in solution.
nstrument is operative in the visible range of the
magnetic spectrum.
COLORIMETRY
• It is a most common analytical technique used in
biochemical estimation in clinical laboratory.

• It involves the quantitative estimation of colour.

• A substrate must be estimated colorimetrically,


must be coloured or it should be capable of
forming chromogens (coloured complexes)
through the addition of reagents.
• Coloured substance absorb light in relation to
their colour density.

• The colour density will be proportional to the


concentration of coloured substance.

• The instruments used in this method are called


colorimeter or photometer.
PRINCIPLE OF COLORIMETER

• When a monochromatic light passes through a


coloured solution, some specific wavelength of light is
absorbed which is related to colour density.

• The amount of light absorbed or transmitted by a


colour solution is accordance with two law, i.e. Beer’s
law and Lambert’s law.
COMPONENT OF COLORIMETER
Light source
Slit
Monochromator(filter)
Cuvette
Photocell
Galvanometer
FUNCTION OF EACH COMPONANT

Light source

Two kinds of lamp:-


1.Halogen Deuterium :- for measurement in the
ultraviolet range 200 – 900 nm.
2.Tungsten lamp:- for measurement in the visible 400
– 760 nm and near-infrared ranges.
MONOCHROMATOR(FILTER) :

FILTER:

•Used for selecting the monochromatic light.


•Filters will absorb light of unwanted
wavelength and allow only monochromatic light
to pass through.

Three Types:
1.Prism
2.Grating
PRISM
•When light travels from one medium to another
medium , it is refracted and enters in the new medium
at a different angle.
Prism wavelength spectrum
GLASS FILTER:-

• Glass filters are selectively transmit light in


particular range of wavelength.
GRATINGS :
• GRAPHITE

• Light (Tungsten light) is


reflected on graphite. This
graft separate light in
different wave length . By
rotation of slit, desirable
wave length of light come out
from slit. And Beam of that
wave length is generated.
• Desired wavelength selected
by the adjustment of an exit
slit.
CUVETTE (Sample cell ):

As per lambert – beer's


law path length is fixed
to 1 cm.
Sample cell has 1 cm
diameter.
A container that
contains a sample is
usually called cell.
THREE TYPES OF CELL:-

1.Glass

• 340nm wavelength of light absorbed in


glass cell.
• cheap
2. Quartz
• It allows passage both type of light,
ultraviolet & visible ranges.
• So used for measurement of both ranges.
costly.

3.Plastic cuvette
• Shorter Life Span
•Easily get Scratches
•Low Cost
PHOTOCELL (PHOTODETECTOR)
•These are the devices to measure the
intensity of light by converting light energy in to
electric energy.
•They are made up of light sensitive material
such as selenium.

GALVANOMETER
• Readout device.
• A galvanometer is used to detect and measure
electrical current produced by the
photodetector.
ADVANTAGE:-

•It is very easy to operate.

DISADVANTAGES:-

• Less sensitive.
• Limited range of filters are available.
• If the light source is not stable ,there is a
possibility of errors due to a change from the
initial light intensity during a measurement.
• The manual operation are limited.
Spectrophotometer
Principle
The working of colorimeter & Spectrometers is
based on Beer's & Lambert's law.

Beer's Law:-It states that the optical density


of a solution is directly proportional to the
concentration of the solution .

Lambert's law:-It states that the optical density


of a coloured solution is directly proportional to
the path of light.
According to Beer's & Lambert's law where,
T=10-kcL,
T=transmittance
K=Constant characteristic of the solution
C=concentration of the coloured solution
L=Path of light through the coloured solution

O.D.=2 – log T%
Differences: Colorimeter &
Spectrophotometer
Colorimeter Spectrophotometer

Limited for the visible 
Ultra violet & infrared
portion of spectrum region also visible
(visible light) e.g.340nm

Cheap 
Very costly

Two digit reading after 
Four digits reading after
desimal point. desimal point .

Less sensitive 
More sensitive

Glass are used. 
Prism are used.

Glass cuvette or test 
Quartz cuvette is used
tube is used for reading which does not absorb
which absorb 340nm 340nm light.
light. 
Halogen lamps are used.

Tungsten lamps are 
Can use specific filter.
used. • Can do kinetic method.

Can’t use specific filter.

Can’t do kinetic method.
Auto analyzers are mainly two types:-
• Semi Auto Analyzer
• Fully Auto Analyzer

Semi Auto Analyzer

Example : ERBA CHEM 5- PLUS


Advantage :
 Displaying the test results
 Printing & memorizing these results
 Graphs of all linear & nonlinear reactions.


Disadvantage :

initial stage of Analysis are performed manually
 Pipetting of reagent
 Pipetting of specimen
 Mixing & incubation.

This instrument require minimum 500 microliters of reagent for test.

Manual L-J chart to draw
Fully Auto Analyzer
The auto analyzer perform all the function of semi
auto analyzer.

1.Automatic dispensing of reagent (by reagent


probe).

2.Automatic dispensing of samples .

3.Automatic mixing of reaction mixtures.

4.Incubating of reacting mixture .


Advantages:-
• Many samples with different parameter can analyzed at time.
• Good precision
• Less reagent required.
• Less sample required.
• Less man power required .
• Maintain the temperature
• For Sample & For Reagent
• For incubation period.
• Can stored result in memory.
• It have facility to accommodate various samples, standards, calibrations &
Q.C. Sera.
• Automated L-J Chart is visible
• Programmable wash cycles between samples & tests for minimum carry over.
• Auto dilution is also possible
Two types of fully auto analyzer:-
Batch analyzer
Random analyzer

Random Access analyzer
- Perform Any number of Parameter from any number of sample.
- More sample in the short period of time .
- Facility of continuous loading of sample
- Facility of “stat” analysis - Urgent sample.
- Facility of autodilution.
- Plotting of daily & monthly Q.C. Chart (L-Jchart).
- Capability to perform a test with 2 to 3 reagents.
- Some of the analyzer has separate assembly to
wash cuvette so very less chance of
contamination.

Example : ERBA XL 640


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