Buffy Coat

What is the buffy coat?

It is a concentration of leukocyte suspension. It is called buffy coat because of a buff; which is a kind of yellowish to brownish in color (buff in hue). Basically, a buffy coat is a combination of platelets and white blood cells from the whole blood sample.

The blood is concentrated using a centrifugation process collected in EDTA anticoagulant. The fraction of the anti-coagulated blood sample consisting of white blood cells and platelets is the buffy coat.

Buffy coat is less than 1% of the total blood volume. Buffy coat is situated in between the plasma and erythrocytes. Although the color is a bit yellow to brown, there are some deviations. The color is dependent on a lot of factors such as the presence of neutrophils. If there is a huge amount of neutrophils, then the color is more on the greenish side. It is because of neutrophil’s green-colored myeloperoxidase. ^{(1, 2, 3, and 4)}

Image 1: A test tube outlining the different components of the blood wherein the buffy coat sits in between the plasma and red cells.
Picture Source: wikimedia.org

 

Image 2: A closer look at the buffy coat component of the blood.
Picture Source: researchgate.net

 

Image 3: Separation of plasma from buffy coat.
Picture Source: researchgate.net

Buffy coat : Uses

• It is primarily used to extract DNA from the mammal’s blood sample.
• It helps detect infection with malaria and blood parasites. Examples of blood parasites are:
  • Leishmania donovani
  • Microfilariae
  • Trypanosomes
  • Fungal pathogens Histoplasma capsulatum
• It helps in performing manual differential white blood cell. A buffy coat is obtained and the blood is smeared. The smeared blood contains a higher number of white blood cells as opposed to using the whole blood.
• It improves platelet yield.
• It is the ideal method in meeting the emergency requirement for platelet.
• It greatly reduces the possibility of leukocyte contamination in platelet.
• It reduces the cost to poor patients. ^{(4, 5, and 6)}

How to collect a buffy coat and test for the presence of parasite?

1. Using a Pasteur pipette, fill the small narrow bore plastic with EDTA anti-coagulated blood.
2. Centrifuge the blood for a total of 15minutes at RCF 1000g. Individual laboratories centrifuge at different times, usually ranging between 10 and 25 minutes.
3. After the centrifuge, there will be a supernatant plasma above the buffy coat layer. You should remove and discard that upper layer.
4. The buffy coat layer and the red cells should be transferred immediately to the end of the slide. The end of the pipette is used to mix the blood component on the slide. Make a thin preparation using a spreader. Usually, a small amount of plasma is withdrawn together with the buffy coat as some parasites are concentrated in the plasma, which is just right above the buffy coat layer. Examples are trypanosomes and microfilariae.
5. Air dry the preparation.
6. Once dry, fix with ethanol or methanol for about two minutes.
7. Staining is done using Giemsa or Field’s thin filming staining methods.
8. Examine under 40x and 100 x objectives. ^{(6, 7, 8, and 9)}

Why some laboratories prefer buffy coat method?

A platelet-rich buffy coat is an alternative source for the platelet concentration method, especially in Europe primarily because buffy coat causes less platelet activation and damage. Buffy coat method is used worldwide such as in Latin America, the United States, and Asian countries.  ^{(5, 7)}

Image 4: A comparison image between buffy coat and leuko pak.
Picture Source: cloudfront.net

Alternative to the buffy coat

Buffy coat is a traditional method of obtaining human donor leukocytes. There is a better alternative and it is in the form of Leuko Paks. It is preferred by many because it enables you to perform more experiments using a single product. It is also time-efficient as you don’t need to collect, isolate, and perform a culture of the cells. ^{(9, 10)}

Which method to use primarily depends on the preference of the laboratory. Of course, there are other things that need to be considered such as the budget, urgency, and purpose of the procedure.

References

1. https://en.wikipedia.org/wiki/Buffy_coat
2. https://www.stemcell.com/how-to-generate-a-buffy-coat.html
3. https://www.sciencedirect.com/topics/immunology-and-microbiology/buffy-coat
4. https://www.bioivt.com/human-buffy-coat/
5. https://microbeonline.com/buffy-coat-definition-composition-preparation-uses/
6. https://professionaleducation.blood.ca/en/transfusion/publications/blood-bag-spiking-procedure
7. https://www.researchgate.net/topic/Blood-Buffy-Coat
8. https://www.ncbi.nlm.nih.gov/pubmed/3252919
9. https://www.transfusionguidelines.org/red-book/chapter-7-specifications-for-blood-components/7-9-platelets-pooled-buffy-coat-derived-leucocyte-depleted
10. https://www.allcells.com/allcells-leuko-pak-vs-buffy-coat/