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A nucleic acid is made up of various chains of monomeric nucleotides, known as a macromolecule. It contains the genetic information of the cell. They commonly manifest themselves as RNA (ribonucleic acids) or DNA(deoxyribonucleic acid).
Various staining methods such as Haemotoxylin are used to differentiate between a red blood cell and white blood cell. Stain (Methyl green pyronin) is used because of its preferential affinity for DNA. It stains DNA, green and RNA, rose-red. The intensity of the colours reflected would represent the concentration of DNA and RNA present.
RNase and DNase is an enzyme that catalyzes the degradation of RNA and DNA respectively.
A Granulocyte is a cell with multi nucleus, while Agranular has a single large nucleus. The stain enables the nucleus to be visible.
-Colour of stain observed
Firstly, the control slide (MGP slide) was stained with both rose red and green because both DNA and RNA nucleic acids where still present in the cell. Therefore the control slide had both colours present.
Next, the treated slide (MGP+RNase slide) was stained green. This was because RNase is an enzyme which degrades RNA. Therefore leaving behind only DNA, which, when treated with the MGP stain leaves a green colour.
The control simple(MGP) was stained green and red in the nucleus and red in the cytoplasm. This shows us that DNA was located mainly in the nucleus while RNA was located throughout the cell in the nucleus and cytoplasm.
This is because DNA molecules are too large to be able to move through the nuclear membrane, however RNA molecules are smaller hence able to diffuse through the nuclear membrane and in to various parts of the cell.
Both granular and agranular cells were observed in the blood smear. This is because white blood cells have both granulocytes
IN 1902 two scientist named Unna and Pappenheim, treated RNA with MGP stain which obtained a magenta colour. The result obtained from the source has the same colours however in different shades as compared to the results obtained from the experiment. From the source RNA(Refer to Fig 1) is stained magenta while RNA from experiment was stained rose red.
The two scientist mention above also recorded where RNA was located in the cell. They concluded that RNA was mostly found in the cytoplasm of the cell. This is consistent with the results obtained from the experiment.
During the air drying process of blood smear, a hair dryer was used to increase the rate of evaporation. However, if the amount of heat was not carefully monitored it might cause the blood smear to crack and hence the denaturing of the nucleic acids.
In order to prevent this, alternative measures could be used such as placing slides in heating thingy to increase the rate of evaporation
The experimental process of dehydrating was to submerge the slides instantly into 95% ethanol for a few seconds. However this method might not have removed all the water. To ensure proper dehydration has been done, the slide should be placed in increasing concentrations of alcohol.
e.g 30%->60%->90% .
Xylene is used as a solvent, thus using this solvent, the background will appear transparent and the cells can be more clearly seen. However it is carcinogenic (Any substance that is radionuclide or radiation) therefore great care must be taken when handling this substance.
-Blood Smearing process
During blood smearing process, ensure that the drop of blood is smeared in one swift and smooth motion. This is to prevent cell breakage due to abrasion.
Methyl green pyronin is light sensitive, hence ensure that the coplin jar that contains MGP is wrapped with a foil to protect it from the light
As body fluids was used in the experiment, it is important we use the correct safety measures. Firstly, ensure that lab coat and covered shoes are wore at all times in the lab.
Next, when handling samples, ensure that gloves are wore. This is to ensure that no infectious disease can be transmitted as well as prevent contamination of sample.
Lastly, discard materials that have come into contact with body fluids into the sharps container. E.g gloves, slides
Before the lancing of the finger, ensure that the finger was cleaned with a alcohol swab to minimize the risk of infection.
Next, lancet must be discarded in the sharp immediately after use.
Finally, use a plaster to cover the wound hence preventing foreign particles and bacteria from entering
The aims of the experiment were met.
Effects of MGP on nucleic acids were recorded for both RNA and DNA.
Observation of the location of nucleic acid and identification of granulocytes were also possible after staining.
The experiment was also carried out with outmost care and results were consistent with external source hence it was a success.
The aims of the experiment is to prepare a blood smear. Classify white blood cells into Granular and Agranular. Identify, differentiate and locate DNA and RNA nucleic acids using methyl green pyronin stain as well as observe effects of DNase and RNase respectively on nucleic acids.
The control slide which contains nucleic acid stained with MGP (Methyl green pyronin) stain , should result in both rose red and green being expressed.
While the RNase slide which contains nucleic acid treated with RNase and then stained with MGP stain, should result in only green being expressed