haematology reference intervals for established and …...05 diagnostic perspectives | volume 1 |...
TRANSCRIPT
Both clinical evaluation and monitoring critically depend on knowledge of laboratory reference ranges. With the continuing development of the automated blood count, studies often restrict themselves to standard, time-honoured parameters, limiting the spread of innovation. Here, we present reference interval for a state-of-the-art haema-tology analyser, the Sysmex XE-5000, including a variety of parameters introduced only recently and some for which clinical application is currently under investigation. Blood samples were taken from 176 female and 133 male apparently healthy hospital employees and a broad spectrum of parameters available with the software installed assessed.
1) Reinier de Graaf Zieken- huis, Delft, Netherlands2) Sysmex Europe, Norderstedt, Germany3) Erasmus Medisch Centrum, Rotterdam, Netherlands
Haematology reference intervals for established and novel parameters in healthy adults
J. M. Pekelharing 1, O. Hauss 2, R. de Jonge 3, J. Lokhoff 1, J. Sodikromo 1, M. Spaans 1, R. Brouwer 3, S. de Lathouder 3, R. Hinzmann 2
Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Introduction
Since the first automated cell counters have been introduced in the haematology laboratory some decades ago, the flow cytometry and aperture technology have steadily been further developed and refined. The first generation of analysers counted only the number of red cells pre-sent in blood, while subsequent models were also able to quantify the white blood cells and the platelets. During the seventies and eighties the ‘three-part WBC differential’ was introduced, later followed by the ‘five-part differential’ which provides the absolute and relative number of leukocyte subsets: the neu-trophilic granulocytes, lymphocytes, monocytes, eosinophils and basophils, next to the number of platelets and erythrocytes 1, 2.
The recently introduced Sysmex XE-5000 analyser has the latest analytical inno-vations on board and is able to quantify 78 parameters in a blood sample of 130 µL in manual mode (or in 40 µL when in capillary mode) in only one minute (manufacturer specifications). It also measures 12 parameters of cells in other body fluids than blood, such as cere-brospinal fluid, peritoneal fluid, ascites and synovial fluid, but that was not a part of this study.
The Sysmex XE-5000 uses laser light to measure on a cell-by-cell basis the side scatter, forward scatter and side fluores-cence light. Additionally, impedance-based counts are performed for RBC and platelet counts. Further counts and measurements use radio frequency and direct current methods and haemoglo-bin colourimetry. Compared to previous generations, the XE-5000 software is able to quantify a number of new para-meters.
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02Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Blood parameters included in this study
1 WBC – white blood cell concentration
2 RBC – red blood cell concentration
3 HgB – haemoglobin concentration
4 HCt – haematocrit
5 MCV – mean corpuscular volume
6 MCH – mean corpuscular haemoglobin
7 MCHC – mean corpuscular haemoglobin concentration
8 PLt – platelet concentration
9 RDW-SD – RBC distribution width, standard deviation
10 RDW-CV – RBC distribution width, coefficient of variation
11 PDW – platelet distribution width
12 MPV – mean platelet volume
13 P-LCR – platelet-large cell ratio
14 PCt – platelet crit
15 Neut % – percentage of neutrophils
16 Neut # – neutrophil concentration
17 LyMPH % – percentage of lymphocytes
18 LyMPH # – lymphocyte concentration
19 MoNo % – percentage of monocytes
20 MoNo # – monocyte concentration
21 eo % – percentage of eosinophils
22 eo # – eosinophil concentration
23 BaSo % – percentage of basophils
24 BaSo # – basophil concentration
25 Ig % – percentage of immature granulocytes
26 Ig # – immature granulocyte concentration
27 HFLC % – percentage of highly fluorescent lymphocytic cells
28 HFLC # – highly fluorescent lymphocytic cell concentration
29 NRBC % – percentage of nucleated red blood cells
30 NRBC # – nucleated red blood cell concentration
31 Ret% – percentage of reticulocytes
32 Ret# – reticulocyte concentration
33 HFR – percentage of high-fluorescence reticulocytes
34 MFR – percentage of medium-fluorescence reticulocytes
35 LFR – percentage of low fluorescence reticulocytes
36 IRF – immature reticulocyte fraction
37 Ret-He – reticulocyte haemoglobin content
38 WBC-B – WBC from WBC/BASO channel
39 WBC-D – WBC from DIFF channel
40 PLt-o – fluorescence optical platelet concentration
41 PLt-I – impedance platelet concentration
42 IPF % – immature platelet fraction
43 MicroR – percentage of microcytic red blood cells
44 MacroR – percentage of macrocytic red blood cells
45 Neut-X – average side scatter of neutrophils (granularity)
46 RBC-He – mean RBC haemoglobin content from optical RBC count
47 Delta-He – RET-He minus RBC-He
48 FRC % – percentage of fragmented red cells (schistocytes)
49 FRC # – fragmented red cell concentration
50 RPI – reticulocyte production index
51 H-IPF – highly fluorescent IPF
52 IPF # – immature platelet concentration
53 PLt-X – average platelet fluorescence
54 % HyPo-He – percentage of hypo-haemoglobinised red cells
55 % HyPeR-He – percentage of hyper-haemoglobinised red cells
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03Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
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Before conducting clinical studies to determine the value of each of the new parameters in medical practice, it is im-perative to know in detail what values can be found in the healthy general po-pulation. It was the purpose of this study to determine the exact reference inter-vals for all parameters that are measured by the XE-5000 analyser, including novel parameters not included in the initial release.
The fact that different institutes have established different reference intervals finds its origin in a number of variables, including preanalytical conditions. Most importantly, however, there are diffe-rences in the selection of the reference populations that are used by each insti-tute. For instance, some calculate the reference intervals from a large number of patients who visit the clinics.
Others use a double selection by using for instance, only samples from the eye disease clinics, reasoning that few patients with haematological diseases will be found in this category.
Others use volunteers from the hospital or laboratory staff, while some refine this strategy by paying attention to the age of the volunteer. In short, all these selec-tions will create different reference inter-vals, although the differences are some-times quite small 3–5.
Blood parameters not included in this study
1 NRBC + W – sum of NRBC and WBC
2 RBC-o – optical RBC count
3 Neut-y – average neutrophil fluorescence
4 area % – percentage of haematopoietic progenitor cells
5 area # – haematopoietic progenitor cell concentration
6 IMI # – immature WBC concentration
7 Ret-y – average reticulocyte forward scatter
8 RBC-y – average RBC forward scatter
9 IRF-y – average IRF forward scatter
10 HPC # – haematopoietic progenitor cell concentration
11 Neut % & – NEuT% minus IG%
12 Neut # & – NEuT# minus IG
13 LyMPH % & – LymPH% minus HFLC%
14 LyMPH # & – LymPH# minus HFLC#
Table 1 List of parameters on XE-5000 with brief explanation
We decided to recruit our reference population from the hospital employees after the launch of a publicity campaign, and to exclude some samples on the basis of clearly aberrant results.
Materials and methods
Blood samples for this reference study were collected from employees of the hospital. On three separate days blood was obtained from a total of 176 female and 133 male subjects, 16–63 years of age, apparently healthy employees. Employ-ees were encouraged to donate blood by means of a publicity campaign on the hospital intranet and by information leaflets distributed throughout the hos-pital. Donation was anonymous; however, a small questionnaire had to be filled in. Age, gender, use of multivitamins and the use of drugs, including hormonal contra-ceptives etc. had to be reported. A veni-puncture was performed by qualified personnel. EDTA (7.2 mg K2-EDTA 4 mL, BD Vacutainer, BD, Plymouth, UK) tubes were used for the measurement of hae-mocytometry, serum gel tubes (SST II gel 8.5 mL, BD Vacutainer) were used for additional tests. Tubes were sent to the laboratory for further processing and measurement throughout the day. All measurements were performed the same day, within 2h of venesection.
04Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
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All haemocytometry parameters available on XE-5000 (Sysmex, Kobe, Japan) with the software version installed at the time (version 00-04) were measured in closed mode in EDTA tubes. Ferritin, LDH and total-bilirubin were measured in serum on an Architect 8000 (Abbott, North Chicago IL, USA). These were measured to exclude possible pathological samples.
After exclusion of subjects considered non-healthy, samples from 133 male sub-jects and 176 female subjects remained. Reasons for exclusions were abnormal ferritin (< 15 or > 300 µg for males, < 15 or > 150 µg for females), an abnormal WBC DIFF, declaration of having hay fever (1 subject), folic acid deficiency (1 sub-ject), abnormal lactate dehydrogenase (2 subjects), declared use of non-stero-idal anti-inflammatory drugs (NSAIDs) (1 subject), declared use of antibiotics (2 subjects), abnormal bilirubin (1 sub-ject), declared recent infectious disease (1 subject), an abnormal ESR (3 subjects) and declaration of having a hereditary coagulation disorder (1 subject).
The reference intervals were determined as 95% confidence intervals of the popu-lation, by striking the top and bottom 2.5% of samples for each parameter, that is the top and bottom 3 samples for men (2.5% of 133), 4 samples for women (2.5% of 176) and 8 samples overall (2.5% of 309). The intervals for male and female subjects were compared using the Mann-Whitney-U-test and the Analyse-It soft-ware (Analyse-It software Ltd, Leeds, UK) for Microsoft Excel. p-values of 0.05 or lower were considered significant.
For the distribution curves, the intervals were divided into 10 equal classes, with the exception of HFLC, due to an interval too narrow for a meaningful division into 10 classes. The results were then distributed over these classes and the results plotted for men, women, and where no statistically significant differ-ence was found, for the overall population.
Results
The reference intervals determined are compiled in table 2. Reference intervals are listed separately for men and women where a statistically significant (p ≤ 0.05) difference between the intervals was found, otherwise, the interval was calcu-lated over all samples. Table 2 lists these reference intervals and demonstrates the statistically significance – or lack there-of – of differences in reference intervals for male and female samples. Figure 1, 2 and 3 illustrate the distribution of several new WBC, RBC and platelet parameters.
Discussion
While previous studies on reference intervals in haematology do, of course, exist, including for Sysmex X-Class ana-lysers 6, most focus on the standard set of parameters available from most suppliers. Here, for the first time, we report reference intervals for most of the full range of diagnostic parameters on the new XE-5000, including para-meters unique to Sysmex X-Class instru-ments 7, 8. Additionally, we provide inter-vals for some research parameters for which literature exists which suggests clinical usefulness 9.
05Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
0
10
20
30
40
50
0.090.080.070.060.050.040.030.020.010
IG#% of reference population
IG# [10 ]
malefemaleoverall
0
10
20
30
40
50
60
70
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
0-
0.1
IG%% of reference population
IG% [%]
malefemaleoverall
0
10
20
30
40
50
60
0.050.040.030.020.010
HFLC#% of reference population
HFLC# [10 /l]
malefemale
0
5
10
15
20
25
30
35
0.80.70.60.50.40.30.20.10
HFLC%% of reference population
HFLC% [%]
malefemaleoverall
0
5
10
15
20
25
30
35
40
498-
540
455-
497
412-
454
369-
411
326-
368
283-
325
240-
282
197-
239
154-
196
111-
153
PLT-O% of reference population
PLT-O [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
45
2.625-
2.708
2.541-
2.624
2.457-
2.54
2.373-
2.456
2.289-
2.372
2.205-
2.288
2.121-
2.204
2.037-
2.120
1.953-
2.036
1.869-
1.952
RET-He% of reference population
RET-He [fmol]
malefemaleoverall
Figure 1 Distribution curves for selected WBC parameters. Individual distribution curves are given for male and female subjects as well as the overall distribution in the absence of a significant difference. (a) The absolute immature granulocyte (IG) count; (b) the IG percentage of all WBCs; (c) the percentage of highly fluorescent lymphocytic cells (HFLC); (d) NEUT-X, the average side scatter of the neutrophils, i.e. the x-axis projection of the centre of the neutrophil ‘cloud’ in the scattergram, given in channel units of the instrument (ch: channel number).
0
10
20
30
40
50
0.090.080.070.060.050.040.030.020.010
IG#% of reference population
IG# [10 ]
malefemaleoverall
0
10
20
30
40
50
60
70
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
0-
0.1
IG%% of reference population
IG% [%]
malefemaleoverall
0
10
20
30
40
50
60
0.050.040.030.020.010
HFLC#% of reference population
HFLC# [10 /l]
malefemale
0
5
10
15
20
25
30
35
0.80.70.60.50.40.30.20.10
HFLC%% of reference population
HFLC% [%]
malefemaleoverall
0
5
10
15
20
25
30
35
40
498-
540
455-
497
412-
454
369-
411
326-
368
283-
325
240-
282
197-
239
154-
196
111-
153
PLT-O% of reference population
PLT-O [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
45
2.625-
2.708
2.541-
2.624
2.457-
2.54
2.373-
2.456
2.289-
2.372
2.205-
2.288
2.121-
2.204
2.037-
2.120
1.953-
2.036
1.869-
1.952
RET-He% of reference population
RET-He [fmol]
malefemaleoverall
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0
10
20
30
40
50
0.090.080.070.060.050.040.030.020.010
IG#% of reference population
IG# [10 ]
malefemaleoverall
0
10
20
30
40
50
60
70
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
0-
0.1
IG%% of reference population
IG% [%]
malefemaleoverall
0
10
20
30
40
50
60
0.050.040.030.020.010
HFLC#% of reference population
HFLC# [10 /l]
malefemale
0
5
10
15
20
25
30
35
0.80.70.60.50.40.30.20.10
HFLC%% of reference population
HFLC% [%]
malefemaleoverall
0
5
10
15
20
25
30
35
40
498-
540
455-
497
412-
454
369-
411
326-
368
283-
325
240-
282
197-
239
154-
196
111-
153
PLT-O% of reference population
PLT-O [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
45
2.625-
2.708
2.541-
2.624
2.457-
2.54
2.373-
2.456
2.289-
2.372
2.205-
2.288
2.121-
2.204
2.037-
2.120
1.953-
2.036
1.869-
1.952
RET-He% of reference population
RET-He [fmol]
malefemaleoverall
0
5
10
15
20
25
138.5-
139.8
137.1-
138.4
135.7-
137.0
134.3-
135.6
132.9-
134.2
131.5-
132.8
130.1-
131.4
128.7-
130.0
127.3-
128.6
125.9-
127.2
NEUT-X% of reference population
NEUT-X [ch]
malefemaleoverall
0
10
20
30
40
50
60
1.31.21.11.00.90.80.70.60.50.40.30.20.1
%HYPO-He% of reference population
%HYPO-He [%]
malefemale
0
5
10
15
20
25
30
35
40
1.61.51.41.31.21.11.00.90.80.70.6
%HYPER-He% of reference population
%HYPER-He [%]
malefemale
0
10
20
30
40
50
4.7-
5.1
4.2-
4.6
3.7-
4.1
3.2-
3.6
2.7-
3.1
2.2-
2.6
1.7-
2.1
1.2-
1.6
0.7-
1.1
0.2-
0.6
MicroR% of reference population
MicroR [%]
malefemale
0
5
10
15
20
25
30
35
40
45
14.9-
15.9
13.8-
14.8
12.7-
13.7
11.6-
12.6
10.5-
11.5
9.4-
10.4
8.3-
9.3
7.2-
8.2
6.1-
7.1
5.0-
6.0
MacroR% of reference population
MacroR [%]
malefemaleoverall
0
10
20
30
40
50
0.090.080.070.060.050.040.030.020.010
IG#% of reference population
IG# [10 ]
malefemaleoverall
0
10
20
30
40
50
60
70
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
0-
0.1
IG%% of reference population
IG% [%]
malefemaleoverall
0
10
20
30
40
50
60
0.050.040.030.020.010
HFLC#% of reference population
HFLC# [10 /l]
malefemale
0
5
10
15
20
25
30
35
0.80.70.60.50.40.30.20.10
HFLC%% of reference population
HFLC% [%]
malefemaleoverall
0
5
10
15
20
25
30
35
40
498-
540
455-
497
412-
454
369-
411
326-
368
283-
325
240-
282
197-
239
154-
196
111-
153
PLT-O% of reference population
PLT-O [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
45
2.625-
2.708
2.541-
2.624
2.457-
2.54
2.373-
2.456
2.289-
2.372
2.205-
2.288
2.121-
2.204
2.037-
2.120
1.953-
2.036
1.869-
1.952
RET-He% of reference population
RET-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
521-
554
487-
520
453-
486
419-
452
385-
418
351-
384
317-
350
283-
316
249-
282
215-
248
Delta-He% of reference population
Delta-He [amol]
malefemaleoverall
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He% of reference population
RBC-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
1.6-
1.7
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
RPI% of reference population
RPI
malefemale
0
10
20
30
40
50
60
70
80
90
15.3-
16.9
13.6-
15.2
11.9-
13.5
10.2-
11.8
8.5-
10.1
6.8-
8.4
5.1-
6.7
3.4-
5.0
1.7-
3.3
0-
1.6
FRC#% of reference population
FRC# [10 /l]
malefemale
0
10
20
30
40
50
60
70
80
90
0.37-
0.40
0.33-
0.36
0.29-
0.32
0.25-
0.28
0.21-
0.24
0.17-
0.20
0.13-
0.16
0.09-
0.12
0.05-
0.08
0-
0.04
FRC%% of reference population
FRC% [%]
malefemale
0
5
10
15
20
25
30
35
6.9-
7.5
6.2-
6.8
5.5-
6.1
4.8-
5.4
4.1-
4.7
3.4-
4.0
2.7-
3.3
2.0-
2.6
1.3-
1.9
0.5-
1.2
IPF%% of reference population
IPF% [%]
malefemale
0
5
10
15
20
25
30
35
40
521-
554
487-
520
453-
486
419-
452
385-
418
351-
384
317-
350
283-
316
249-
282
215-
248
Delta-He% of reference population
Delta-He [amol]
malefemaleoverall
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He% of reference population
RBC-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
1.6-
1.7
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
RPI% of reference population
RPI
malefemale
0
10
20
30
40
50
60
70
80
90
15.3-
16.9
13.6-
15.2
11.9-
13.5
10.2-
11.8
8.5-
10.1
6.8-
8.4
5.1-
6.7
3.4-
5.0
1.7-
3.3
0-
1.6
FRC#% of reference population
FRC# [10 /l]
malefemale
0
10
20
30
40
50
60
70
80
90
0.37-
0.40
0.33-
0.36
0.29-
0.32
0.25-
0.28
0.21-
0.24
0.17-
0.20
0.13-
0.16
0.09-
0.12
0.05-
0.08
0-
0.04
FRC%% of reference population
FRC% [%]
malefemale
0
5
10
15
20
25
30
35
6.9-
7.5
6.2-
6.8
5.5-
6.1
4.8-
5.4
4.1-
4.7
3.4-
4.0
2.7-
3.3
2.0-
2.6
1.3-
1.9
0.5-
1.2
IPF%% of reference population
IPF% [%]
malefemale
0
5
10
15
20
25
30
35
40
521-
554
487-
520
453-
486
419-
452
385-
418
351-
384
317-
350
283-
316
249-
282
215-
248
Delta-He% of reference population
Delta-He [amol]
malefemaleoverall
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He% of reference population
RBC-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
1.6-
1.7
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
RPI% of reference population
RPI
malefemale
0
10
20
30
40
50
60
70
80
90
15.3-
16.9
13.6-
15.2
11.9-
13.5
10.2-
11.8
8.5-
10.1
6.8-
8.4
5.1-
6.7
3.4-
5.0
1.7-
3.3
0-
1.6
FRC#% of reference population
FRC# [10 /l]
malefemale
0
10
20
30
40
50
60
70
80
90
0.37-
0.40
0.33-
0.36
0.29-
0.32
0.25-
0.28
0.21-
0.24
0.17-
0.20
0.13-
0.16
0.09-
0.12
0.05-
0.08
0-
0.04
FRC%% of reference population
FRC% [%]
malefemale
0
5
10
15
20
25
30
35
6.9-
7.5
6.2-
6.8
5.5-
6.1
4.8-
5.4
4.1-
4.7
3.4-
4.0
2.7-
3.3
2.0-
2.6
1.3-
1.9
0.5-
1.2
IPF%% of reference population
IPF% [%]
malefemale
a) b)
c) d)
a) b)
c) d)
06Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Di
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Figure 2 Distribution curves for selected RBC parameters. Curves given as described for figure 1. (a) The reticulocyte haemoglobin content (RET-He); (b) the difference between the reticulocyte and mature erythrocyte haemoglobin content (Delta-He); (c) the absolute count of fragmented red cells (FRC); (d) the FRC percentage of all erythrocytes; (e) the percentage of hypo-haemoglobinised red cells (% Hypo-He); (f) the percentage of hyper-haemoglobinised red cells (% HypER-He); (g) the percentage of microcytic cells; (h) the percentage of macrocytic cells; (i) the haemoglobin content of mature erythrocytes from the fluorescence optical count (RBC-He); (j) a comparison of RBC-He and the mean cellular haemoglobin calculated from impedance counting and haemoglobin photo-metry. RBC-He is calibrated to MCH during instrument setup.
0
5
10
15
20
25
138.5-
139.8
137.1-
138.4
135.7-
137.0
134.3-
135.6
132.9-
134.2
131.5-
132.8
130.1-
131.4
128.7-
130.0
127.3-
128.6
125.9-
127.2
NEUT-X% of reference population
NEUT-X [ch]
malefemaleoverall
0
10
20
30
40
50
60
1.31.21.11.00.90.80.70.60.50.40.30.20.1
%HYPO-He% of reference population
%HYPO-He [%]
malefemale
0
5
10
15
20
25
30
35
40
1.61.51.41.31.21.11.00.90.80.70.6
%HYPER-He% of reference population
%HYPER-He [%]
malefemale
0
10
20
30
40
50
4.7-
5.1
4.2-
4.6
3.7-
4.1
3.2-
3.6
2.7-
3.1
2.2-
2.6
1.7-
2.1
1.2-
1.6
0.7-
1.1
0.2-
0.6
MicroR% of reference population
MicroR [%]
malefemale
0
5
10
15
20
25
30
35
40
45
14.9-
15.9
13.8-
14.8
12.7-
13.7
11.6-
12.6
10.5-
11.5
9.4-
10.4
8.3-
9.3
7.2-
8.2
6.1-
7.1
5.0-
6.0
MacroR% of reference population
MacroR [%]
malefemaleoverall
0
5
10
15
20
25
138.5-
139.8
137.1-
138.4
135.7-
137.0
134.3-
135.6
132.9-
134.2
131.5-
132.8
130.1-
131.4
128.7-
130.0
127.3-
128.6
125.9-
127.2
NEUT-X% of reference population
NEUT-X [ch]
malefemaleoverall
0
10
20
30
40
50
60
1.31.21.11.00.90.80.70.60.50.40.30.20.1
%HYPO-He% of reference population
%HYPO-He [%]
malefemale
0
5
10
15
20
25
30
35
40
1.61.51.41.31.21.11.00.90.80.70.6
%HYPER-He% of reference population
%HYPER-He [%]
malefemale
0
10
20
30
40
50
4.7-
5.1
4.2-
4.6
3.7-
4.1
3.2-
3.6
2.7-
3.1
2.2-
2.6
1.7-
2.1
1.2-
1.6
0.7-
1.1
0.2-
0.6
MicroR% of reference population
MicroR [%]
malefemale
0
5
10
15
20
25
30
35
40
45
14.9-
15.9
13.8-
14.8
12.7-
13.7
11.6-
12.6
10.5-
11.5
9.4-
10.4
8.3-
9.3
7.2-
8.2
6.1-
7.1
5.0-
6.0
MacroR% of reference population
MacroR [%]
malefemaleoverall
0
5
10
15
20
25
138.5-
139.8
137.1-
138.4
135.7-
137.0
134.3-
135.6
132.9-
134.2
131.5-
132.8
130.1-
131.4
128.7-
130.0
127.3-
128.6
125.9-
127.2
NEUT-X% of reference population
NEUT-X [ch]
malefemaleoverall
0
10
20
30
40
50
60
1.31.21.11.00.90.80.70.60.50.40.30.20.1
%HYPO-He% of reference population
%HYPO-He [%]
malefemale
0
5
10
15
20
25
30
35
40
1.61.51.41.31.21.11.00.90.80.70.6
%HYPER-He% of reference population
%HYPER-He [%]
malefemale
0
10
20
30
40
50
4.7-
5.1
4.2-
4.6
3.7-
4.1
3.2-
3.6
2.7-
3.1
2.2-
2.6
1.7-
2.1
1.2-
1.6
0.7-
1.1
0.2-
0.6
MicroR% of reference population
MicroR [%]
malefemale
0
5
10
15
20
25
30
35
40
45
14.9-
15.9
13.8-
14.8
12.7-
13.7
11.6-
12.6
10.5-
11.5
9.4-
10.4
8.3-
9.3
7.2-
8.2
6.1-
7.1
5.0-
6.0
MacroR% of reference population
MacroR [%]
malefemaleoverall
0
5
10
15
20
25
138.5-
139.8
137.1-
138.4
135.7-
137.0
134.3-
135.6
132.9-
134.2
131.5-
132.8
130.1-
131.4
128.7-
130.0
127.3-
128.6
125.9-
127.2
NEUT-X% of reference population
NEUT-X [ch]
malefemaleoverall
0
10
20
30
40
50
60
1.31.21.11.00.90.80.70.60.50.40.30.20.1
%HYPO-He% of reference population
%HYPO-He [%]
malefemale
0
5
10
15
20
25
30
35
40
1.61.51.41.31.21.11.00.90.80.70.6
%HYPER-He% of reference population
%HYPER-He [%]
malefemale
0
10
20
30
40
50
4.7-
5.1
4.2-
4.6
3.7-
4.1
3.2-
3.6
2.7-
3.1
2.2-
2.6
1.7-
2.1
1.2-
1.6
0.7-
1.1
0.2-
0.6
MicroR% of reference population
MicroR [%]
malefemale
0
5
10
15
20
25
30
35
40
45
14.9-
15.9
13.8-
14.8
12.7-
13.7
11.6-
12.6
10.5-
11.5
9.4-
10.4
8.3-
9.3
7.2-
8.2
6.1-
7.1
5.0-
6.0
MacroR% of reference population
MacroR [%]
malefemaleoverall
e) f)
g) h)
0
5
10
15
20
25
30
35
40
521-
554
487-
520
453-
486
419-
452
385-
418
351-
384
317-
350
283-
316
249-
282
215-
248
Delta-He% of reference population
Delta-He [amol]
malefemaleoverall
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He% of reference population
RBC-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
1.6-
1.7
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
RPI% of reference population
RPI
malefemale
0
10
20
30
40
50
60
70
80
90
15.3-
16.9
13.6-
15.2
11.9-
13.5
10.2-
11.8
8.5-
10.1
6.8-
8.4
5.1-
6.7
3.4-
5.0
1.7-
3.3
0-
1.6
FRC#% of reference population
FRC# [10 /l]
malefemale
0
10
20
30
40
50
60
70
80
90
0.37-
0.40
0.33-
0.36
0.29-
0.32
0.25-
0.28
0.21-
0.24
0.17-
0.20
0.13-
0.16
0.09-
0.12
0.05-
0.08
0-
0.04
FRC%% of reference population
FRC% [%]
malefemale
0
5
10
15
20
25
30
35
6.9-
7.5
6.2-
6.8
5.5-
6.1
4.8-
5.4
4.1-
4.7
3.4-
4.0
2.7-
3.3
2.0-
2.6
1.3-
1.9
0.5-
1.2
IPF%% of reference population
IPF% [%]
malefemale
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He versus MCH% of reference population
[fmol]
overall RBC-He
overall MCH
(j)i) j)
07Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Figure 3 Distribution curves for selected pLT parameters. Curves given as described for figure 1. (a) The absolute count of immature platelets; (b) the immature platelet percentage of all platelets; (c) the most highly fluorescent, i. e. most immature platelet percentage.
Di
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We confirmed the statistically significant differences of reference intervals for men vs. women for HGB, RBC, HCT, MCHC and the number of platelets reported by others e.g. 6, 10, 11. However, we observed such differences for a far wider range of parameters, including once more the neutrophil count, as already observed by Bain and England in 1975 12. Nonetheless, the differences in these additional cases, while statistically significant, were often such as not being clinically relevant. The reference intervals we report here for those parameters previously published match quite well those found by others. For some parameters, we found inter-vals that are slightly higher than in some publications, but in line with others. Such probably population-based shifts
stress the importance of verifying the applicability of a published reference interval for the population a given hae-matology analyser is used on before use as suggested by IFCC or CLSI 13, 14 – or determining intra-laboratory refer-ence limits directly 3.
0
5
10
15
20
25
30
35
40
2.5-
2.7
2.2-
2.4
1.9-
2.1
1.6-
1.8
1.3-
1.5
1.0-
1.2
0.7-
0.9
0.4-
0.6
0.1-
0.3
H-IPF% of reference population
H-IPF [%]
malefemaleoverall
0
5
10
15
20
25
30
35
15.1-
17.5
14.4-
15.0
12.8-
14.3
11.2-
12.7
9.6-
11.1
7.8-
9.5
6.2-
7.9
4.6-
6.3
3.0-
4.7
1.6-
3.1
IPF#% of reference population
IPF# [10 /L]
malefemaleoverall
0
5
10
15
20
25
30
22.1-
22.9
21.2-
22.0
20.3-
21.1
19.4-
20.2
18.5-
19.3
17.6-
18.4
16.7-
17.5
15.8-
16.6
14.9-
15.7
14.0-
14.8
PLT-X% of reference population
PLT-X [ch]
malefemaleoverall
0
5
10
15
20
25
30
35
40
45
479-
529
438-
478
397-
437
356-
396
315-
355
274-
314
233-
273
192-
232
151-
191
110-
150
PLT-I% of reference population
PLT-I [10 /l]
malefemale
0
5
10
15
20
25
30
35
16.18-
17.57
14.78-
16.17
13.38-
14.77
11.98-
13.37
10.58-
11.97
9.18-
10.57
7.78-
9.17
6.38-
7.77
4.98-
6.37
3.58-
4.97
WBC-B% of reference population
WBC-B [10 /l]
malefemale
0
5
10
15
20
25
30
35
15.34-
16.64
14.03-
15.33
12.72-
14.02
11.41-
12.71
10.10-
11.4
8.79-
10.09
7.48-
8.78
6.17-
7.47
4.86-
6.16
3.55-
4.85
WBC-D% of reference population
WBC-D [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
2.5-
2.7
2.2-
2.4
1.9-
2.1
1.6-
1.8
1.3-
1.5
1.0-
1.2
0.7-
0.9
0.4-
0.6
0.1-
0.3
H-IPF% of reference population
H-IPF [%]
malefemaleoverall
0
5
10
15
20
25
30
35
15.1-
17.5
14.4-
15.0
12.8-
14.3
11.2-
12.7
9.6-
11.1
7.8-
9.5
6.2-
7.9
4.6-
6.3
3.0-
4.7
1.6-
3.1
IPF#% of reference population
IPF# [10 /L]
malefemaleoverall
0
5
10
15
20
25
30
22.1-
22.9
21.2-
22.0
20.3-
21.1
19.4-
20.2
18.5-
19.3
17.6-
18.4
16.7-
17.5
15.8-
16.6
14.9-
15.7
14.0-
14.8
PLT-X% of reference population
PLT-X [ch]
malefemaleoverall
0
5
10
15
20
25
30
35
40
45
479-
529
438-
478
397-
437
356-
396
315-
355
274-
314
233-
273
192-
232
151-
191
110-
150
PLT-I% of reference population
PLT-I [10 /l]
malefemale
0
5
10
15
20
25
30
35
16.18-
17.57
14.78-
16.17
13.38-
14.77
11.98-
13.37
10.58-
11.97
9.18-
10.57
7.78-
9.17
6.38-
7.77
4.98-
6.37
3.58-
4.97
WBC-B% of reference population
WBC-B [10 /l]
malefemale
0
5
10
15
20
25
30
35
15.34-
16.64
14.03-
15.33
12.72-
14.02
11.41-
12.71
10.10-
11.4
8.79-
10.09
7.48-
8.78
6.17-
7.47
4.86-
6.16
3.55-
4.85
WBC-D% of reference population
WBC-D [10 /l]
malefemale
0
5
10
15
20
25
30
35
40
521-
554
487-
520
453-
486
419-
452
385-
418
351-
384
317-
350
283-
316
249-
282
215-
248
Delta-He% of reference population
Delta-He [amol]
malefemaleoverall
0
5
10
15
20
25
30
35
2.099-
2.158
2.039-
2.098
1.979-
2.038
1.919-
1.978
1.859-
1.918
1.799-
1.858
1.739-
1.798
1.679-
1.738
1.619-
1.678
1.559-
1.618
RBC-He% of reference population
RBC-He [fmol]
malefemaleoverall
0
5
10
15
20
25
30
35
40
1.6-
1.7
1.4-
1.5
1.2-
1.3
1.0-
1.1
0.8-
0.9
0.6-
0.7
0.4-
0.5
0.2-
0.3
RPI% of reference population
RPI
malefemale
0
10
20
30
40
50
60
70
80
90
15.3-
16.9
13.6-
15.2
11.9-
13.5
10.2-
11.8
8.5-
10.1
6.8-
8.4
5.1-
6.7
3.4-
5.0
1.7-
3.3
0-
1.6
FRC#% of reference population
FRC# [10 /l]
malefemale
0
10
20
30
40
50
60
70
80
90
0.37-
0.40
0.33-
0.36
0.29-
0.32
0.25-
0.28
0.21-
0.24
0.17-
0.20
0.13-
0.16
0.09-
0.12
0.05-
0.08
0-
0.04
FRC%% of reference population
FRC% [%]
malefemale
0
5
10
15
20
25
30
35
6.9-
7.5
6.2-
6.8
5.5-
6.1
4.8-
5.4
4.1-
4.7
3.4-
4.0
2.7-
3.3
2.0-
2.6
1.3-
1.9
0.5-
1.2
IPF%% of reference population
IPF% [%]
malefemale
a) b)
c)
08Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
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While we report reference intervals for both PLT-I, the impedance platelet count, and PLT-O, the fluorescence-optical count, it should be noted that the ana-lyser uses a switching algorithm deciding which count is more reliable on a case-by-case basis and reports the more reli-able one as PLT. The same holds true for WBC-B, the WBC count from the WBC-BASO channel, which together with WBC-D, the WBC count from the DIFF channel is interchangeably reported as the actual WBC count, depending on which the system assesses as the more reliable one for the individual case.
For the use of the reference interval for RPI some limitations do apply: interest-ingly, whereas it is usually assumed that a reticulocyte production index (RPI) of ‘ 1’ is normal, our data show that the refer-ence interval can actually reach as low as 0.2 and women, especially, despite the regular blood loss during menstruation, tend to have an RPI significantly below 1. This suggests that the assumptions made in calculating RPI are somewhat questio-nable. They assume a rather monocausal regulation of reticulocyte maturation time in the bone marrow tied directly to the haematocrit. It needs to be em-phasised that RPI should only be used for adult anaemic patients. While an RPI higher than 2 can indeed suggest a sig-nificantly increased haematopoiesis, our data suggests that the clinical value of a low RPI should not be overestimated. Erythropoietic bone marrow activity can alternatively be assessed via the immature reticulocyte function and the reticulocyte haemoglobin content, in addition to, of course, the reticulocyte count itself.
09Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Parameter unit Sex Sysmex Xe-5000
Mann-Whitney p-value
WBC 109/L m 3.91–10.90 0.0013
f 4.49–12.68
RBC 1012/L m 4.44–5.61 <0.0001
f 3.92–5.08
HGB mmoL/L m 8.44–10.49 <0.0001
f 7.39–9.06
g/L m 135–169
f 119–146
HCT % m 40.0–49.4 <0.0001
f 36.6–44.0
mCV fL m 81.8–95.5 <0.0001
f 82.9–98.0
mCH fmol m/f 1.676–2.005 0.9950
pg m/f 27.0–32.3
mCHC mmoL/L m 20.1–21.7 <0.0001
f 19.7–21.5
g/L m 324–350
f 318–347
PLT 109/L m 166–308 <0.0001
f 173–390
RDW-SD fL m 37.1–45.7 <0.0001
f 38.2–49.2
RDW-CV % m 12.0–13.6 0.0002
f 12.1–14.3
PDW fL m 10.1–16.1 0.0003
f 9.9–15.4
mPV fL m 9.3–12.1 0.0006
f 9.1–11.9
P-LCR % m 18.5–42.3 0.0004
f 17.5–42.3
PCT % m 0.17–0.32 <0.0001
f 0.18–0.39
NEuT# 109/L m 1.8–6.98 <0.0001
f 2.1–8.89
LymPH# 109/L m/f 1.26–3.35 0.1170
mONO# 109/L m 0.29–0.95 0.0011
f 0.25–0.84
EO# 109/L m 0.03–0.59 0.0068
f 0.01–0.40
BASO# 109/L m/f 0.01–0.07 0.0780
NEuT% % m 41.0–70.7 <0.0001
f 42.9–74.3
LymPH% % m 19.1–47.9 0.0285
f 18.3–45.7
mONO% % m 5.2–15.2 <0.0001
f 4.2–11.8
EO% % m 0.6–7.6 <0.0001
f 0.2–5.3
BASO% % m 0.1–1.2 0.0009
f 0.1–1.0
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10Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Parameter unit Sex Sysmex Xe-5000
Mann-Whitney p-value
NRBC# 109/L m/f 0–0
NRBC% % m/f 0–0
RET# 109/L m 23.0–70.1 0.0005
f 17.0–63.8
RET% % m/f 0.43–1.36 0.7483
LFR % m/f 89.9–98.4 0.0539
mFR % m/f 1.6–9.5 0.0668
HFR % m/f 0–1.7 0.0649
IRF % m/f 1.6–10.5 0.0539
IG# 109/L m/f 0–0.06 0.5170
IG% % m/f 0–0.6 0.0764
HFLC# 109/L m 0–0.03 0.0120
f 0–0.03
HFLC% % m/f 0–0.4 0.0827
PLT-O 109/L m 173–329 <0.0001
f 174–407
RET-He fmol m/f 1.996–2.407 0.9066
pg m/f 32.1–38.8
Delta-He fmol m/f 0.268–0.451 0.2904
pg m/f 4.31–7.26
RBC-He fmol m/f 1.676–2.023 0.9950
pg m/f 27.0–32.6
RPI m 0.4–1.5 <0.0001
f 0.2–1.1
FRC# 109/L m 0–8.3 0.0011
f 0–11.6
FRC% % m 0–0.17 0.0012
f 0–0.25
IPF% % m 0.8–6.3 0.0348
f 0.8–6.2
H-IPF % m/f 0.2–1.9 0.0559
IPF# 109/L m/f 2.3–12.7 0.3568
PLT-X ch m/f 14.3–20.1 0.3461
PLT-I 109/L m 166–308 <0.0001
f 173–390
WBC-B 109/L m 3.91–10.9 0.0013
f 4.49–12.68
WBC-D 109/L m 3.94–10.76 0.0133
f 4.23–12.71
NEuT-X ch m/f 127.9–138.0 0.3822
%HyPO-He % m 0.1–0.5 0.0468
f 0.1–1.1
%HyPER-He % m 0.9–1.3 <0.0001
f 0.7–1.2
microR % m 0.5–3.0 <0.0001
f 0.3–2.8
macroR % m/f 5.6–11.5 0.1746
Table 2 Reference intervals for the parameters included in the study. one reference interval is listed where no significant difference was found between samples for male and female subjects. p-value as per Mann-Whitney U test is given.
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11Diagnostic Perspectives | Volume 1 | page 01 – 11
Haematology reference intervals for established and novel parameters in healthy adults
Sysmex Corporation 1-5-1, Wakinohama-Kaigandori, Chuo-ku, Kobe 651-0073, Japan, Phone +81 (78) 265-0500 · Fax +81 (78) 265-0524 www.sysmex.co.jp
Sysmex europe gmbH Bornbarch 1, 22848 Norderstedt, Germany, Phone +49 (40) 52726-0 · Fax +49 (40) 52726-100 · [email protected] www.sysmex-europe.com
Diagnostic Perspectives – The Sysmex Europe eJournal for Clinical Laboratory Medicine · Published 2010 by Sysmex
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9. Linssen J, Aderhold S, Nierhaus A et al. (2008): Automation and validation of a rapid method to assess neutrophil and monocyte activation by routine fluorescence flow cyto-metry in vitro. Cytometry. part B, Clinical Cytometry, 74: 295 – 309. 10. Heil W and Ehrhardt V (2008): Reference ranges for adults and children. pre-analyticalconsiderations. Mannheim: Roche Diagnostics. 11. Thomas L (1998): Hematology. In: Thomas L (Hg.): Clinical Laboratory Diagnostics. Use and Assessment of Clinical Laboratory Results. 1st English Edition. Frankfurt/Main: THBooks, 463 – 547. 12. Bain BJ and England JM (1975): Normal haematological values: sex difference in neutrophil count. British Medical Journal,5953: 306 –309. 13. Solberg HE and Stamm D (1991): IFCC rec-ommendation: the theory of reference values. part 4. Control of analytical variation in the production, transfer and application of reference values. The Journal of Automatic Chemistry, 13: 231 – 234. 14. Solberg HE (2004): The IFCC recommen-dation on estimation of reference intervals. The RefVal program. Clinical Chemistry and Laboratory Medicine: 42: 710 – 714.
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