Answering your questions

Feb. 1, 2011

Sometimes we get patients at our hospital who generate an H&H (hemoglobin and hematocrit) check from our Coulter LH 750. When we perform a spun hematocrit (Hct), we sometimes get a difference of more than 3% between the automated and spun Hct, except in the case of a cold agglutinin, which result is accurate?


The Hct is simply the ratio of the volume of the red blood cells (RBCs) to the volume of whole blood. It could be measured by two different methods. Traditionally, hematocrit is directly determined by centrifugation (spun hematocrits). Now, most hematocrits (instrument hematocrit) are calculated from RBC count and mean cell volume (MCV) using automated hematology analyzers, such as the LH 750. What are the pros and cons with manual and automatic hematocrit?

Spun hematocrit is 1% to 3% higher than the hematocrit from automated instrument due to plasma that is trapped in the erythrocytes in the normal situation. Spun hematocrit, however, may give spuriously higher results (up to 6%) in a number of disorders — including polycythemia, macrocytosis, spherocytosis, hypochromic anemias, sickle-cell anemia, burn patients — due to increased trapped plasma compared to the normal condition. Of course, insufficient centrifugation can also introduce high-spun hematocrit.

In the past few decades, automated hematology analyzers have replaced manual hematocytometer blood-cell counts with increased reliability, precision, and accuracy. For automated hematologic analyzers (LH 750), the RBC and MCV are directly measured by the instrument depending on the number of pulses for RBC and the amplitude of each pulse for MCV using electronic impedance; then the hematocrit is calculated by using following equation:

Hct = RBC x MCV

The interferences that may cause erroneous results of RBC and MCV by automated hematology analyzer could introduce abnormal Hct, which include very high white blood cell (WBC) count, high concentration of large platelets, or agglutinated RBCs.

Using both spun hematocrit and automated hematocrit on same patient samples is not recommended. Switching hematocrit methods on the same patient creates discrepancies, causing confusion for the clinician.

Spun hematocrit is not recommended due to the low accuracy. Automated analyzers provide more accurate and stable results if hematocrit is measured in the linear range. If WBC and RBC are not in the linear range, proper dilution is required. In the situation that a large number of giant platelets are suspected, blood-smear review is recommended, and hemoglobin concentration would be more accurate than hematocrit for testing anemia.

— Guang Fan, MD, PhD
Medical Director, Hematology Service
Oregon Health and Science University
Portland, OR

WBC correction needed with nRBC?

Basic hematology has the lab correct the WBC when there are six or more nucleated red blood cells (nRBCs) when doing a manual differential. With new instruments, sometimes it corrects the WBC and sometimes it does not. If the size of the nRBC does not fall within the WBC parameter, the instrument does not correct for it. When a manual differential is performed and gets greater than six nRBCs, should we correct or not? The instrument says it is not part of the white count; but since the beginning of time, we have been correcting them.


While readers may know the answer to this question instinctively, some may have a hard time giving up on an established practice of correcting for nRBC. Conventional practice in performing manual differentials is that the lab corrects for the presence of nRBCs. This was due to limited ability of previous cell-counting technologies to separate nRBCs from WBCs in the automated counting of WBCs. Newer technologies for cell counting rely upon both better technology (flow-cytometry cells, predominantly) and much improved algorithms for identifying both the expected size and nuclear/cytoplasmic characteristics of each cell population counted. In the vast majority of cases, when an automated cell counter indicates that there is no nRBC interference, even when nRBCs are present on the slide being viewed, the WBC will be accurate and not influenced by the presence of nRBCs. When nRBC interference is noted, newer technologies automatically subtract that and report an accurate count for WBCs. Thus, manual correction for nRBCs should not be necessary if the automated cell counter is working as designed.

In the presence of nRBCs or under any other condition that causes/requires the lab to perform a manual cell count or manual slide review, should the automated WBC number from the analyzer be replaced with a count from manual slide review? Generally, no. Several studies have shown that the precision of manual slide counts is significantly inferior to that of an automated cell counter. This is due to both variation in experience of technologists and subjectivity in defining cell populations; and to the sheer number of cells that can be counted routinely by a technologist compared to an automated instrument. In general, the WBC produced by the automated cell counter should be used. There are no infallible automated cell-counting technologies, however, which is why labs have criteria for both manual review of slides and manual differentials. In the case where the WBC produced by the instrument just does not seem right — given what you see on the slide — hold that result and perform further investigation. This could include repeating the automated cell count on the same instrument after additional sample mixing, repeating the automated count on another instrument (preferably another technology), or making additional slides (or sharing the slide) to consult with other persons with hematology expertise.

— Brad S. Karon, MD, PhD

Transfusions, tests, and timing


We were asked to draw two sets of blood cultures from a patient who was receiving a unit of blood. My thought was that it is best to draw her one hour after completion/discontinuation of the transfusion. The attending physician vehemently disagreed and called my pathologist who consented to having the phlebotomy done while the patient was getting the unit of blood. If this were a transfusion reaction, the blood cultures would be ordered soon after receiving the pint, along with a clot to test ABO/Rh incompatibility, culturing the unit, and collecting urine. If the doctor thought the patient was experiencing signs of sepsis, he should have stopped the transfusion. Can you make sense of this request?


Transfusion-reaction protocols vary from site to site; but in any suspected transfusion-reaction investigation, the transfusion would be stopped and no further transfusions would be given until a cause identified. It is unlikely that the blood culture was part of a transfusion-reaction workup. For most other indications for blood culture (e.g., sepsis, pneumonia) the timing of transfusion to blood culture is probably of little importance. From a pure perspective of blood-culture yield, there is probably little data to guide us on the appropriate timing of blood transfusion to blood-culture collection.

For many laboratory tests, there are guidelines and recommendations regarding the timing of blood transfusion to blood collection. For post-transfusion cell counts, it is recommended to wait 15 minutes to 60 minutes after the transfusion to accurately assess the impact of the transfusion on raising cell counts. For most common chemical tests, the same recommendation exists to allow for determination of steady state electrolyte and chemical analysis. Thus, in the past, many readers have asked for advice regarding timing of blood collection following blood (red cell, predominantly) transfusion and the answer is usually 15 minutes to 60 minutes after the completion of the red-cell transfusion. For blood culture, one would generally follow these same recommendations and wait some period of time after the transfusion to collect. Data linking blood-culture yield to transfusion is lacking, however; and there are certain situations that would call for a blood culture to be completed while a transfusion was being administered.

The primary issue would be that of wanting to collect the first set of cultures before antibiotics are administered in a patient with suspected bacteremia or sepsis. For this reason, in most instances, blood cultures would be collected first, followed by the transfusion. If the patient's condition were considered so critical that the transfusion could not be delayed, or if the blood culture were missed or omitted in a patient suspected of sepsis and the transfusion started, then it would be appropriate to collect the culture while a transfusion was being performed in order to start antibiotics as soon as possible. In theory, transfusion of “good” blood could dilute the bacterial load of a septic patient making recovery of the causative bacteria more difficult, again providing a rationale for collection of blood cultures before transfusion or soon after a transfusion begins in the case that the culture was missed initially. Though this situation would probably not arise often, I also do not have data to support a policy against collection of blood cultures while a transfusion is running.

— Brad S. Karon, MD, PhD

Brad S. Karon, MD, PhD, is associate professor of laboratory medicine and pathology, and director of the Hospital Clinical Laboratories, point-of-care testing, and phlebotomy services at Mayo Clinic in Rochester, MN.

MLO's “Tips from the Clinical Experts” provides practical, up-to-date solutions to readers' technical and clinical issues from a panel of experts in various fields. Readers may send questions to Brad S. Karon, MD, PhD, by e-mail at [email protected].