How to identify normal leukocytes in a blood smear


Q. I’m not sure if I can identify leukocytes correctly. Could you give me some tips? Thanks very much.

A. Sure! When you are just starting out in hematopathology, it can be a bit overwhelming. It’s really not as difficult as it seems. There are just 5 kinds of cells that you see in normal peripheral blood, and with a few guidelines, you can tell them apart pretty easily.

Neutrophils
These are the most numerous white cells in normal blood. There are two in the above image (from WebPath), one at about 3 o’clock and one at about 10 o’clock. They are part of a category of white cells called “granulocytes,” which refers to the cytoplasmic granules you see in these cells. In neutrophils, the cytoplasmic granules are mostly small, pale peachy-pink granules. These granules (called “specific granules”) are what give the neutrophil cytoplasm its pinkish color. There are also scattered larger, dark purple (or “azurophilic”) granules. These are called “primary granules” because they are the granules that appear first as the neutrophil matures. If you forget the thing about neutrophil maturation, you can remember which is which by remembering that the p words go together (primary=purple).

The nucleus of a normal neutrophil is also unique-looking. It’s segmented – pinched off into different sections, like sausage links – rather than round, like most other cells. Neutrophils are sometimes called polymorphonuclear leukocytes because there are several (poly) bodies (morpho) in the nucleus (nuclear). Rarely, you might see a “band” cell (which is the neutrophil at 10 o’clock), which is the stage of neutrophil right before the nucleus becomes segmented. Neutrophil chromatin in general is clumpy, and you can’t see any nucleoli.

Lymphocytes
These are the second most numerous type of white cell in normal blood. There’s one lymphocyte at 8 o’clock in the above image. Lymphocytes are generally a bit smaller than neutrophils, and the thing that sets them apart is their chromatin, which is both clumpy and smudgy at the same time. It looks like someone took a finger and rubbed the nucleus before the ink fully dried. Although there are clumps in lymphocyte chromatin, there aren’t discrete white spaces between the clumps, like you see in neutrophil chromatin. Check out the above image and you’ll see what I mean. Sometimes you’ll see lymphocytes that are a bit larger, with more cytoplasm and maybe a few coarse granules. T cells often have this appearance (though you really can’t tell for sure without doing some special studies).

Monocytes
Monocytes are big cells (there’s one at about 8 o’clock above) with lots of cytoplasm. The cytoplasm often has a “dishwater” appearance, meaning it is sort of cloudy and grayish. Sometimes, as in the cell above, it’s more of a pale purple color. You can see some fine purple granules scattered about as well. The nucleus is big and it’s usually indented, or horseshoe shaped. The chromatin is pretty fine (finer than neutrophil or lymphocyte chromatin), and it has a weird “raked” appearance on high power (it looks like someone messed up the chromatin by dragging a rake across it).

Eosinophils
These cells, along with basophils, are probably the easiest to spot (there’s an eosinophil at 2 o’clock above). Both eosinophils and basophils are granulocytes. The granules in eosinophils are beautiful – they are large, luminous, and reddish-orange. The word eosin comes from the Greek word eos, which means “flush of the dawn sky.” Very cool name for these gorgeous, sunrise-colored granules. The nucleus is nothing to write home about, really – it’s segmented into a few different parts, and it looks kind of like a neutrophil nucleus.

Basophils
You can tell a basophil from a mile away: it’s the cell with the big, super-dark-purple-blue granules (there’s one at 4 o’clock above). The granules are so numerous and dark that they often obscure the nucleus (which is a rather boring nucleus, usually divided into two segments). Basophils are the least numerous of all white blood cells – you may have to look several fields to find one.

And that’s it! When you start looking at different diseases (like infection, or leukemia), it gets a bit more complicated, because you often see immature cells out in the blood. But for now, you can just focus on the normal, mature white cells. Once you get familiar with these, they start looking like little friends that you happily recognize from across the street.

Which direction does blood flow through the ductus arteriosus?

Q. I just had a quick question for you. Our notes say that a ductus arteriosus allows flow from the pulmonary artery to the aorta, which I knew.
However, they also say that it’s a left to right shunt, and that it can become right to left. This confuses me, since from what I know, flow would be going from right (pulmonary artery) to left (aorta).

A. When we talk about the ductus allowing flow from the pulmonary artery (right) to aorta (left), we’re talking about intrauterine flow through the ductus. Before birth, the pressure on the right side of the heart is greater than the pressure on the left – so blood flows from pulmonary artery to aorta (through the ductus).

After birth, though, the pressure on the left becomes greater than the pressure on the right. In most babies, the ductus closes (probably in response to the new levels of oxygen in the blood). In some babies it remains patent, in which case flow would now be from the aorta (left; higher pressure) to the pulmonary artery (right; lower pressure).

If the ductus is widely patent, then after a while, that left to right shunt can put enough pressure on the lungs that they react by closing down vessels, effectively making it more difficult to push blood through. Now the right heart has to work really hard to push blood through the lungs – and it can get to the point where the right heart is actually bigger and stronger than the left, making the shunt reverse and go from pulmonary artery (right; higher pressure) to aorta (left; lower pressure).

Why would you wash red cells for patients with PNH?

Q. I’m currently doing my rotations at Children’s Memorial Hospital’s blood bank and I was reading the standard operating procedure for washing red cells. One of the conditions in which they need to be washed is paroxysmal nocturnal hemoglobinuria. Do you know why this is?

A. The reason for washing red cells for people with PNH is to get rid of any ABO incompatible plasma.

In a person without PNH, ABO incompatible plasma doesn’t cause any perceptible hemolysis. The antibodies in the donor unit probably just get diluted out enough that they don’t have much of an effect. Or perhaps they get sopped up by other ABO antigens on other cells (did you know that you have A and B antigens on cells besides red cells?! Weird.).

But patients with PNH are super susceptible to complement-induced red cell destruction. They lack the ability to anchor certain proteins (including proteins that protect the red cell against complement) to the red cell membrane. Patients with PNH have a hard time down-regulating even a small amount of complement activation – so theoretically, a transfusion of even a small amount of non-ABO compatible plasma could lead to hemolysis. There have been a few cases of hemolytic transfusion reactions in patients with PNH that have been attributed to this phenomenon…so in 1948, blood banks began washing red cells before giving them to patients with PNH.

However, this practice has been called into question. The Mayo Clinic reviewed 38 years of experience with transfusing patients with PNH, and only found one documented episode of post-transfusion hemolysis). Their conclusion was that the important thing is to use group-specific blood products for patients with PNH; washing seems to be an unnecessary precaution.