nRBNC and giant-platelet interference on a cell counter.

Q Our lab recently purchased a cell counter that will enumerate nucleated red blood cells (nRBCs) and report a corrected white blood cell count (WBC) if the size of the nRBCs is at the threshold for triggering a cellular interference flag. It is our protocol to first estimate the WBC under hi-dry to verify the counter's corrected WBC. We then go to 100x to see if nRBCs or other interferences are present, such as clumped platelets, giant platelets, and so forth.

If just nRBCs are seen, we perform a diff and enumerate the number of nRBCs. We report the counter's nRBC result if it agrees with our manual count; we report the counter's WBC if it agrees with our estimate.

The scenario gets tricky if both nRBCs and giant platelets are seen on smear. How would the panel verify/determine the corrected WBC if both interferences are seen?

This is what we do:

1. Perform separate counts to enumerate giant platelets and nRBCs/100 cells counted as WBCs on the counter by a) enumerating nRBCs by counting the number of nRBC/100 cells (when counting 100 cells include both giant platelets and WBCs), and b) enumerating giant platelets by counting the number of giant platelets/100 cells (when counting 100 cells, include both nRBCs and WBCs);

2. Add the number of giant platelets and nRBCs from the first step together; and

3. Calculate WBC correction as follows:

* Number of nRBCs plus giant platelets x Uncorrected WBC = Total Cellular Interference 100 + (number of nRBCs plus giant platelets)

* Corrected WBC = Uncorrected WBC minus Total Cellular Interference

If this corrected WBC result is close to the counter's corrected WBC, we report out the counter results. If this does not agree with the counter result but agrees with our estimate from the smear, we report out our calculated result. Does that panel agree with this? If not, can you recommend a detailed procedure?

A You are correct in recognizing that the counter will enumerate nRBCs and can generate an nRBC and cellular interference flag. It is our laboratory's experience that there are situations where a cellular interference flag is generated and no nRBCs are enumerated, as well as times where the nRBC flag will be present and no cellular interference flag is present. Our laboratory has taken the approach that if either of these flags are present, we review the smear, and if there are nRBCs present we will report the number of nRBCs seen on the smear and the corrected WBC.

The issue arose regarding what to do if neither the cellular interference flag nor the nRBC flags are present; but on manual review of the blood smear, nRBCs are noted. We initially wanted to utilize the uncorrected WBC count as part of our correction formula. When we reviewed this approach with the manufacturer, its representatives indicated that the uncorrected WBC count is simply a number to indicate how may particles greater than 35 fL are present and that it could not be utilized as an accurate WBC count or used in any calculation. We decided that we would run such a sample on a different complete blood count instrument that did not provide/calculate a "corrected" WBC count and then perform the manual correction utilizing the formula of: WBC x 100/100 + nRBCs noted on smear review.

While our laboratory has never corrected for giant platelets, we do utilize a similar correction for the presence of circulating megakaryocyte nuclei. We take the number of nRBCs plus the number of megakaryocytic nuclei and apply the same formula.

Utilizing the same approach for giant platelets, you would recalculate the WBC for giant platelets in those situations where you did not have an nRBC flag or a cellular interference flag, and on smear review where you saw giant platelets and your platelet smear estimate did not match the platelet count. I do have some concerns related to which platelets (i.e., how giant) will end up being counted by the instrument and how you would be able to ensure that the platelets you are counting on smear review are or are not being incorporated into the corrected WBC count. A giant platelet is defined as a platelet larger than a red cell (typically > 100 fL) but we know that particles >35 fL may impact the WBC count. How would you be able to reliably ensure that the platelets that are interfering with the white cell count (those between 36 fL and 100 fL) are appropriately identified by the technologist when they review the smear? Unless the giant platelets were numerous concurrent with a very low white count, the impact to the white count and then subsequent clinically significant change in WBC and associated values, such as the absolute neutrophil count, are minor. While the issue you raise has technical merit, the inability to reliably compensate for a technical issue with minimal clinical impact may end up having a negative impact on the overall quality and consistency of your result.

Finally, I would add that WBC counts can be estimated by peripheral smears, and this will aid in the verification of the corrected WBC count. The use of a disposable diluting pipette system and a hemacytometer for a manual white blood cell count, however, can be useful in the event that the corrected count disagrees with your peripheral estimate due to interference of any sort.

--Louis Rose, MT (ASCP)

Ohio State University Medical Center

Columbus, Ohio