1 Department of Haematology 2 Department of Nephrology St. James's, Hospital Dublin Ireland
Sir,
Recently, the optimal haematocrit (Hct) level that minimizes cardiovascular risk in renal patients has been a matter of some debate [13], with studies giving apparently contradictory results. Accurate and reproducible monitoring of the degree of anaemia by the laboratory is essential if valid comparisons are to be made between data generated in different renal centres.
Traditionally the anaemia of renal disease has been monitored and managed exclusively using the Hct, but haemoglobin (Hb) is considered to be equally useful and these tests have been used interchangeably. We have been concerned over reports regarding the relative accuracy of Hct for anaemia screening [3,4] and we have completed some comparisons of Hct values analysed using different methods. All blood samples were collected into dipotassiumethylenediaminetetraacetic acid (K2EDTA) and were analysed within 5 h of venipuncture.
We initially compared the Hb and Hct values of 46 fresh diagnostic blood samples using our in house Coulter STKS cytometer (Coulter Electronics, Hialeah, FL) and also the H2 cytometer (Technicon Instruments Corp). All cytometers calculate Hct indirectly by first estimating the mean cell volume (MCV) electronically, but the STKS and the H2 use different principles of operation: the popular impedance principle of cell sizing and the more recently available two-angle light scatter technology on iso-volumetrically sphered red cells, respectively. Both cytometers were calibrated as recommended by the manufacturers.
Full blood count Hct levels measured on sequential diagnostic requests using the STKS (0.396±0.081, mean±SD) ranged from 0.255 to 0.567 and were significantly different to those obtained using the Technicon H2 instrument (0.416±0.080) when analysed by paired t-test (P<0.0001). The concurrent Hb concentrations of these same samples, (13.9±3.0 and 13.6±3.1, respectively) were not significantly different (P=>0.17).
We then compared automated Hct values (STKS Coulter) with a manual microhaematocrit centrifuge method [5] and a radio-isotopic dilution method [6] where we used iodinated (125I) human serum albumin. The microhaematocrit method was a National Committee for Clinical Laboratory Standards (NCCLS) approved standard. Microcapillary red cell column lengths for each blood sample were measured precisely (n=5, CV% <2) by microscopy. The blood samples (n=121) were selected to have a wide range of Hct values (0.2150.664) by STKS and 60 of these were microcytic (MCV <80 fl). The Hct values obtained were 0.438±0.105 (mean±SD) for centrifuged, 0.403±0.107 for the automated cytometric and 0.421±0.113 for the isotopic dilution method of analysis. The results of all three methods were significantly different by paired t test (P<0.0001) and the differences are expressed in Table 1.
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The technical reasons for the method specificity of Hct values are complex [7]. Briefly, cytometers are not direct measurement devices but comparators which are calibrated using normal cells. Errors can occur in the electronic sizing of iron deficient red cells because cell membranes are less flexible and the mean cell haemoglobin (MCH) is reduced compared to normal [7]. Considerable Hct variations have been noted previously in renal blood samples using cytometers with different operating principles [4]. The sizes of such abnormal cells are always overestimated using the traditional centrifugation methods due to an increase in the volume of plasma trapped in the red cell column (Table 1), and need adjustment. We considered that the isotopic approach, although tedious, would give the most accurate (100%) measure of red cell volume (Table 1
).
On the other hand Hb concentrations are measured directly using simple colorimetric methods which are common to all analysis, both manual and automated. Haemoglobin measurement is calibrated accurately and precisely using the cyanmethaemoglobin reference method [8] for which international reference preparations are available. We consider Hb to be superior to Hct for monitoring anaemia because of the availability of these international reference standard preparations and due to the directness and simplicity of the assay methodology. No such reference standards exist for Hct measurements and Hct results are method specific. This point is emphasized by the fact that commercial quality control blood preparations for the calibration of cytometers such as CBC-Tech (R and D Systems, Inc, Minneapolis, MN) provide tables of Hct (and MCV) target values that are method (or instrument) specific and significantly different; the corresponding Hb target values are virtually identical.
An additional advantage of Hb for anaemia monitoring is its relative stability post venipuncture. Hct values will increase with blood sample age due to extracellular fluid uptake whereas Hb levels will not. This point was made in the recently published multi-centre normal haematocrit trial in North America [1] where blood samples were processed in one central laboratory and Hct values for incoming specimens (some aged) were derived from Hb values by multiplying by 3 so as to eliminate Hct over-estimations.
The important message for nephrologists is that Hb is always superior to Hct for monitoring the anaemia of renal disease because it can be measured with greater accuracy both within and between laboratories. Haemoglobin and Hct are both excellent correlates of anaemia and correlate well with one another. However, Hct results are method specific, whereas Hb results are not. Furthermore, iron deficiency can exaggerate the discrepancies between the results of different techniques for the measurement of Hct. Haemoglobin measurements are as little influenced by variables such as the hydration status of renal patients [4] as they are by red cell shape. Consequently, Hb monitoring allows a finer control in the clinical management of individual renal patients and should facilitate valid comparisons to be made between clinical data generated in different renal centres.
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