Observing blood cells under the microscope is often part of the medical analysis to find any abnormalities in the structure of the blood. The process is called blood smear or hematology analysis. Often, doctors would request for complete blood count to check the disparity of the red blood cell, white blood cells and get the total blood volume.
Humans have many different components in their blood, including white blood cells, platelets, and various proteins.
Red blood cells, commonly known as erythrocytes, are the primary oxygen-carrying cells in humans. They account for 70% of a person’s cells by count and play an important function: transporting oxygen from the lungs to all other body areas and returning carbon dioxide to the lungs. They have a few unique characteristics that help them do their job. But there are other cells that you can observe when you take a sample of your blood and observe them under the microscope.
We will look into the blood cells under the microscope to learn unique techniques and processes for a fun and exciting experiment.
What will you learn?
You’ll learn how to take cell samples for high-powered magnification using a lancet and how to properly smear them in slides. To inspect blood cells using a brightfield microscope, you’ll need to draw blood from your finger with a lancet. After reviewing this information and beginning materials, you will have a step-by-step guide for viewing blood cells under a microscope.
Background about Blood Cells
Blood is responsible for keeping almost all of our body’s cells alive and growing. The human blood consists of four major components: white blood cells, red blood cells, platelets, and plasma. Before getting into the ins and outs of circulatory systems, though, it might help to get familiar with the terms used when discussing blood.
Humans have a closed circulatory system. It delivers oxygen, nutrients, hormones, and other essentials throughout our bodies. A “closed” system implies that all of our blood flows through arteries, veins, and capillaries. It means that rather than just sloshing around inside us, our blood is being vigorously circulated by our hearts.
A hemorrhage is a condition in which blood leaks from broken blood vessels, such as a cut or scrape. The ability of our body’s blood to clot or form a blockage that prevents bleeding, allowing the body time to generate cells and heal the damage, is an essential function.
- White blood cells (WBC) – are part of the body’s immune system and can be found in low numbers in the blood. White blood cells help defend against diseases, infections, parasites, etc. The larger white blood cells that can be seen without magnification are called lymphocytes. White blood cells are essential in an inflammatory response and in producing antibodies that the body needs when ill.
- Red blood cells (RBC) – The most common type of cell in human blood, red blood cells carry oxygen throughout the body. They’re shaped like a disk and have a characteristic biconcave shape that helps them carry more oxygen. Compared to white blood cells, the red blood cells are smaller.
- Platelets – These are tiny cells found in the blood that participate in the process called “blood-clotting” to stop bleeding. They are not immune system cells, but they play an essential role in protecting us from severe bleeding.
- Plasma – This is the fluid component of blood in which the other cells are suspended. It contains water, minerals, nutrients, proteins, and dissolved gases.
- Oxygen-carrying capacity is measured in two ways: hemoglobin concentration and red cell volume. The average red blood cell has a lifespan of 120 days. As they die, they’re replaced by new red blood cells. As a side note, the total volume of the blood in your body is about 10 liters or one gallon! That’s a lot of blood! So you can be sure that it’s doing a lot of work every day to keep you alive and healthy.”Blood smear” or “Hematocrit” – A blood smear is a thin film of blood that’s spread out on a microscope slide. This film is stained and then examined under a microscope. Hematocrit is the percentage of the volume of whole blood that’s made up of red blood cells.
Observing Blood Cells Under the Microscope
Magnification: 1000x Objective: 4/0 (largest-sized)
The microscope we will use is a brightfield one. Brightfield microscopy is an optical illumination system that enhances the contrast of translucent objects or thin tissue samples being examined under a microscope.
Using brightfield microscopy to view blood cells, you can see both the red blood cells and platelets and other entirely opaque cells. This type of microscope helps observe things like human embryos or insects where you need to distinguish between tissues without your perception being obscured by the color of the object.
Using a microscope with only one light source, we can see much more to blood than just red cells and white cells.
OUR RECOMMENDED BRIGHTFIELD MICROSCOPES
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The requirements are the following:
- a brightfield microscope
- blood sample
- Begin by preparing your brightfield microscope and set it at 10x ocular. You may increase magnification at 1000x as you progress.
- Clean your glass slide and cover with a glass cleaning solution and a special glass cleaning cloth. If you have a camera or wear glasses, the manufacturer’s cleaning cloth will work just fine. You may also use general-purpose cleaning paper, although it is unnecessary.
- Using a cotton swab and rubbing alcohol, clean one finger’s skin thoroughly.
- Remove the stopper. Open a lancet and expose the sharp point (about 3 mm long). Quickly puncture the fingertip, lay down the lancet, and gently squeeze the finger until a small drop of blood appears on the tip.
- You may now place the droplet of blood from your finger onto the center of the slide and then clean any extra blood from your fingertip. (Bleeding should not be an issue, but if it continues, apply pressure with a cotton ball or paper towel until it subsides).
- Place a coverslip on one edge of the slide before the droplet begins to dry, allowing it to come into contact with the blood drop at an acute angle (blood drop inside the sharp angle of the coverslip to slide). Moving slowly and smoothly across the surface of the slide, push the edge of the coverslip away from the blood droplet. Be careful not to smear the coverslip area adjacent to the blood.
- Place a clean slide on top of the one with your sample, and then press down firmly to ensure that both sides are level. Tamp down the surface gently with a finger if necessary to remove bubbles. If the smear is bright red (rather than light pink), there has been too much blood, and you’ll have to begin again with a clean slide. Only a thin line of blood is needed.
- Place the covered blood smear on the microscope stage with the coverslip facing the objective lens and focus until blood cells appear.
Red blood cells are the most common, measuring approximately 0.007mm in diameter. White blood cells are somewhat larger and more difficult to detect because they are white.
There is typically about one white blood cell for every 1,000 red blood cells. The cells should be free-floating in plasma under the coverslip if your smear is freshly prepared, and all of the blood cells will still be viable. Take a toothpick and gently press down on the edge of the coverslip while you’re still viewing the slide under magnification to observe how the cells move through the plasma. The pressure of your toothpick tap should propel the cells around.
Artifacts or Refractiles
When viewing your dried blood smear under a microscope, you may notice refractile if they are present. There are certain things to consider and factors to consider when detecting refractile when looking at your dried blood smear with a microscope.
First, what stain and method are you using? There might be a problem with the optics or magnifications. What are these criteria? Finally, perhaps your issue is with the specimen or specimen preparation, such as poor spreading technique.
If the sample isn’t being replaced, you may notice a drying-related artifact or a stain-related fault that becomes more apparent during humid summer months. Internal reflections or contamination in the sample and/or optics can be responsible for refractile. It’s also conceivable that the components or mechanisms on your microscope are out of alignment.
Some common causes of refractile in blood cell samples are:
- Presence of water in alcohol
- Under-fixation or late fixation
- Thick smear
- Excess buffer to stain
After the plasma has been appropriately sprayed and dried, water is washed away from thin regions first, and then cells are flushed. The surface of the cortex is thicker, and it dries from the outside to the inside. Loose water can no longer spread from the cell if there is an inactive cell membrane, so cells with loose water will not survive long. Trapped water in a fixed cell membrane can be a cause of refractile due to insufficient drying.
Water-based stains and procedures offer better chances of success, even when dealing with refractile. With faster heating and drying before staining, you may avoid them. Perhaps merely swishing the slide about immediately after placing the smear will be enough to dry it quickly. On the other hand, when it is caused by staining, it is more difficult to resolve.
Live Blood Analysis Refractiles
Much of the same questions and variables may be considered in live blood analysis. A new piece of optical equipment/dye may alter your signal image. The autofluorescence in hemoglobin, for example, might interfere with your emission signal from your fluorescent dye.
If halos are present and causing refractile due to autofluorescence, you should consider using methylene blue, Toluidine blue, or trypan blue in your stain.
A blood smear is easily obtained and has visible characteristics of blood, which is especially advantageous when the disease is present. It’s critical to remember sterility, quality control, and precise technique in collecting your blood smear.
Observing blood cells under a microscope can be an exciting and rewarding experience. In fact, when you see your blood cells for the first time, it’s a feeling that is hard to describe.
However, if you do notice refractile in your blood smears, specific requirements or criteria may need to be addressed. Check with stains and procedures and their connection to humidity.
Maintain proper staining procedures, including methylene blue or other stains for autofluorescence issues. Ensure proper fixation and mounting on slides so that no water is trapped within the smear.
Optical microscopy offers the ability to gather information and aid in continued research. With careful observation, one can diagnose many medical conditions. Properly preparing blood smears is essential in yielding accurate and repeatable results.
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