pharos university faculty of allied medical science clinical laboratory instrumentation (meli-201)
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Pharos university Faculty of Allied Medical SCIENCE Clinical Laboratory Instrumentation (MELI-201). Dr. Tarek El Sewedy. Lecture 3. Incubators , Microscopes and Thermo cyclers. Intended Learning Outcomes. By the end of this lecture the student should learn the basics of the following: - PowerPoint PPT PresentationTRANSCRIPT
Pharos university Faculty of Allied Medical SCIENCE
Clinical Laboratory Instrumentation
(MELI-201)
Dr. Tarek El Sewedy
Lecture 3
Incubators , Microscopes and
Thermo cyclers
Intended Learning Outcomes
By the end of this lecture the student should
learn the basics of the following:
1. Cell Incubators used in Clinical Labs
2. Microscopes
3. Thermo cyclers.
Lecture content
Laboratory Incubators
1. Microscopes.
2. Thermo cyclers
Cell IncubatorsA cell incubator is an apparatus used to grow and maintain
mammalian cell cultures, microbes,or plant cells
The concept of incubation refers to the maintenance of
controlled environmental conditions needed to sustain the
development or growth of cells, eggs, tissues, or whole
organisms.
The incubator maintains optimal temperature, humidity and
carbon dioxide (CO2) levels .
Incubator Alarm System
A controlled atmosphere is achieved by using a CO2-monitoring
device, which draws air from the incubator into a sample
chamber, determines the concentration of CO2, and injects pure
CO2 into the incubator to make up any deficiency.
Air is circulated around the incubator by using a fan to keep
both the CO2 level and the temperature uniform.
Mammalian Cell incubator
Bacteriological Incubator
Cell incubators According to design
Cell incubators usually come in one of three types
of designs:
1. Air draft.
2. Dry wall.
3. Water jacket.
1. Air draft incubators circulate air throughout the interior of the incubator to maintain
constant temperatures.
This style of incubator is used for growing large amounts of cells in a
big growth chamber.
These incubators are designed to respond quickly to environmental
needs of the cells.
However, these incubators lose the temperature very quickly and
must consistently adjust the air to control the temperature.
These are the most commonly used type of incubators
2. Dry wall incubatorsPass air within the walls of the incubator.
The walls then radiate the temperature to the growing
chamber. For this reason these incubators are sometimes
called radiant incubators.
They are better at maintaining a constant temperature
because the jacket does not let heat escape through the
walls of the incubator.
These incubators respond slowly to temperature
adjustments compared to Air draft incubators
Water jacketed incubatorsWater-jacket incubators are surrounded by water within the
walls making up the three sides, the top, and the bottom.
Advantages include stable temperature control and
increased security in the event of power failure (due to
water’s natural insulation abilities).
These are usually smaller incubators and work by the
same principles as dry wall incubators.
These incubators also respond more slowly to temperature
changes.
Use of copper in incubators Copper fights contamination: Some CO2 incubators incorporate copper in
their cabinet design to resist contamination. As the copper breaks down, it
releases copper oxide, which destroys microbes present in the chamber.
Copper is also used inside the water jacket to eliminate contamination
growth over time inside the jacket.
No chemicals are recommended or needed in the water jacket.
Copper is also used around the sample port and the gas injection tube.
Some incubators have a decontamination cycle incorporated where the
temperature can be increased up to 200C
Cleaning of incubators
1. Remove the humidity pan weekly and autoclave
2. Remove all shelves. Autoclave or wash and disinfect as described.
3. Clean and disinfect all access ports, air bleeds, shaft holes, electrical
pass-through. and any other passages into the incubator.
4. Remove the door gasket and gasket guards. Clean and disinfect.
5. Replace all air and CO2 filters as needed, approximately every six
months or when noticeably dirty.
Microscopes A microscope is an instrument used to see objects that are too small for
the naked eye. The science of investigating small objects using such an
instrument is called microscopy.
In Clinical labs the microscope is an essential instrument for the diagnosis
of disease.
There are many types of microscopes, the most commonly used is the
optical microscope which uses light to image the sample.
Other major types of microscopes are the electron microscope (both the
transmission electron microscope and the scanning electron microscope)
Components of a microscope
The various components of the microscope can be classified into four
systems:
Support system
Magnification system
Illumination system
Adjustment system.
Magnification System• Consists of a system of lenses.
The lenses of the microscope are
mounted in two groups:
• The first group of lenses is at the
bottom of the tube, just above the
preparation under examination (the
object), and is called the objective.
• The second group of lenses is at the
top of the tube and is called the
eyepiece.
Objectives of the microscopes The magnifying power of each objective is shown by a
figure engraved on the sleeve of the lens:
— the x10 objective magnifies 10 times;
— the x40 objective magnifies 40 times;
— the x100 objective magnifies 100 times.
(The X100 objective is usually marked with a red ring to show that it must be used with immersion
oil.)
Immersion oil increases the resolving power by focusing light rays on the sample
Eyepiece Magnification
a x5 eyepiece magnifies the image produced by the objective five times;
a x10 eyepiece magnifies the image 10 times.
If the object is magnified 40 times by the x40 objective, then by five times
by the x5 eyepiece, the total magnification is: 5 x 40 = 200 times
Illumination system Light source It is provided by a lamp built into the microscope beneath the stage, MirrorThe mirror reflects rays from the light source onto the object CondenserThe condenser brings the rays of light to a common focus on the object DiaphragmIs used to reduce or increase the amount of light that passes into the condenser.
Resolving power of a microscope
The resolving power of an objective is its ability to reveal closely adjacent
details as separate and distinct.
The greater the resolving power of the objective, the clearer the image.
A good resolving power of a good medical laboratory microscope is
about 0.25 micrometer (the resolving power of the normal human eye is
about 0.25mm).
Care and maintenance Heavy contamination can be removed with mild soapy solutions.
Grease and oil can be removed with the special cleaning solution with a
50 : 50 mixture of distilled water and 95% ethanol.
The mechanical parts should be periodically cleaned and lubricated with
machine oil to make them run freely.
In hot, humid climates fungi may grow on the microscope, particularly on
the surface of the lenses. This can be prevented by keeping the
microscope under an airtight plastic cover when not in use, together with
a dish filled with blue silica.
Electron microscopes
Is a type of microscope that uses a beam of electrons to illuminate the
specimen and produce a magnified image.
Electron microscopes have a greater resolving power than a light-powered
optical microscope, because electrons have wavelengths about 100,000
times shorter than visible light, and can achieve better than 50 pm
resolution and magnifications of up to about 10,000,000x
The electron microscope uses electrostatic and electromagnetic "lenses" to
control the electron beam and focus it to form an image. These lenses are
analogous to, but different from the glass lenses of an optical microscope that
form a magnified image by focusing light on or through the specimen.
Electron Microscopes
Electron Microscopes
Scanning Electron Microscopes
Thermocyclers A laboratory instrument that repeatedly cycles through a series of
temperature changes required for chemical reactions such as the
polymerase chain reaction or PCR.
PCR is used to make multiple copies of DNA. This process is called
amplification because it can generate million copies of DNA.
PCR is used to produce ample of quantities of DNA when only a small
amount is available. It is a valuable tool for DNA analysis, disease diagnosis,
and genetic engineering.
The technology is regularly used in crime scene analysis to collect DNA from
traces of blood, hair, saliva, or skin.
Thermo cyclers
PCR
Stage one is a hot stage called
denaturation At 90◦C temperature
opens up the DNA for copying.
Stage two is a cooling or annealing that
permits the DNA to attach to chemicals
needed to copy it.
Stage three is a warm temperature
cycle called the extension. It
encourages the growth of the DNA
strand.
The effectiveness of a thermo cycler is its ability to
change temperature rapidly with precision.
Part of the heating and cooling efficiency is due to the
small thin reaction tubes used in the thermo cycler.
Samples of DNA to be copied are placed in minuscule
containers called micro tubes.
The thermo cycler is set to run a particular number of
these cycles depending on the amount of DNA a person
wants to collect.
Applications of PCR
Medicine: Detection of mutations in genes causing tumor.
Infectious disease: early diagnosis of viruses such as AIDS.
Forensic: Human DNA fingerprinting.
Research: isolation of certain DNA or RNA regions.
Thermo cycler Specification
Sample block should have 96x 0.2 ml and have option of dual block of 2 X
48 X .2 ml wells in addition to the traditional micro tubes.
Independent control for all blocks.
High ramp rate of 3-5 °C/ sec.
Temperature range should be 0-100°C with high accuracy.
High Temperature uniformity.
Programmable (up to 1000 programs)
Security features should be password protected.
Option of using the instrument through a PC
USB peripheral compatibility
Assignment
Goseph Adel is selected to make the assignment Different applications of
PCR
The Assignment should be delivered before next lecture
Study questions
Mentions 3 different applications of PCR
Mention the main difference between different types of incubators
Suggesting reading
Encyclopedia of Medical Devices and Instrumentation, 2nd ed. New York: Wiley, 2006