ARTICLE
Auteur(s) : Silvia Izquierdo
Álvarez1, Alicia Boudet García1, Santiago
Otero Martínez2, María Dolores Fernández
González3, Joaquín González Revaldería4,
Ángel García De Jalón Comet1, Jesús Fernando Escanero
Marcén5
1Servicio de Bioquímica Clínica, Hospital
Universitario Miguel Servet, Zaragoza;
2Laboratorio Central, Hospital Clinico Universitario de
Santiago, Santiago de Compostela, A Coruña;
3Laboratorio de Bioquímica, Hospital Universitario
Marqués de Valdecilla, Santander, Cantabria;
4Servicio de Bioquímica, Hospital Universitario de
Getafe, Madrid;
5Departamento de Farmacología y Fisiología, Facultad de
Medicina, Uuniversidad de Zaragoza, Spain
Magnesium (Mg) is the most abundant intracellular divalent
cation present in human organism and participates in numerous
physiological processes [1, 2]. The serum Mg determination is an
important biochemical parameter in the context of clinical medicine
for monitoring patients and for helping to diagnose some
pathologies. Magnesium deficiency is known to be linked with
cardiovascular alterations and several renal, gastrointestinal,
neurological and muscular disorders [2, 3]. Each clinical and/or
biochemical laboratory has responsibility for demonstrating its
competence and therefore must obtain results of good (required)
quality.
The ISO (International Standard Organization) 15189 standard
specifies the quality management system requirements, in particular
to medical laboratories and states:
- – “the laboratory shall use only validated procedures
for confirming that the examination procedures are suitable for the
intended use”;
- – “the validations shall be as extensive as are
necessary to meet the needs in the given application or field of
application”;
- – “procedures need to be periodically revalidated in
view of changing conditions and technical advances” [4, 5].
Precision and inaccuracy are two of the key validation
parameters for any instrumental method in the determination of
trace elements and Mg. These parameters can be estimated on the
basis of interlaboratory comparison programmes, or the external
quality control programmes in which participation is required for
ISO 15189 accreditation [1, 6, 7].
The magnesium research field is more complex (more variety of
specimens and different methodologies) than clinical laboratories
(where automatized — colorimetric — methods to determine
Mg in serum are usually used). The standards of quality control in
Mg research are determined by the researchers themselves and by the
magazines where they publish their work.
According to these facts, the aims of the present study were: 1)
to estimate the precision and inaccuracy of different colorimetric
methods for Mg determination in serum, used in different hospitals,
on the basis of the results of participation in interlaboratory
comparison programmes and 2) to establish a common methodology in
the estimation of precision and inaccuracy for the Mg determination
methods used in scientific laboratories.
Material and methods
The results of participation in the external quality control
(Quality Control Interlaboratory Programmes: Bio-Rad EQAS and
external quality control SEQC) for serum Mg determination with
colorimetric methods were analysed during one year (2006). The
colorimetric methods used were:
- – the Mg method based on the modified xylidyl blue
reaction, described by Mann and Yoe [8]. Analytical parameters:
analytical range: 1.6 to 5.0 mg/dL for serum, C.V. < 5%;
- – another colorimetric method with chlorophosphonazo III
[9]. Analytical parameters: precision and accuracy < 5%;
- – the last colorimetric method with calmagite, LX 20
apparatus, auto-analyser with photometric module. Analytical
parameters have not been indicated by manufacturers.
All records of participation in these interlaboratory programmes
were studied in four different clinical laboratories of hospitals
(Santiago, Santander, Getafe and Zaragoza). From these data a
series of paired values were obtained: VL, the
concentration given by the laboratory, and VR, the
target value of the interlaboratory comparison programme. It was
necessary to obtain at least ten pairs of values by the same
analytical method within the range of concentration considered as
usual.
The steps for estimation of precision and accuracy (expressed as
inaccuracy: 100 – accuracy), are indicated in a flow chart (figure 1). Inaccuracy
is expressed in quantitative terms while accuracy is a qualitative
expression. This is the reason that justifies the election of
inaccuracy.
The data were grouped in a table 1
and the calculations were performed using Microsoft Excel.
With respect to second aim of this study, a review of all
original papers published in Magnesium Research during 2005 and
2006 was carried out. The type of sample, the methodology used for
Mg determination and some quality requirements were studied.
Table 1 Summary of data values obtained from records of
an intercomparison programme for estimation of precision and
inaccuracy.
|
VR (interlaboratory comparison value)
|
VL. (laboratory value)
|
Differences: dn = VLn -
VRn
|
Relative differences (accuracies): An = Dn
/ VRn
|
|
VR1
|
VL1
|
d1 = VL1 - VR1
|
d1 / VR1
|
|
VR2
|
VL2
|
d2 = VL2 - VR2
|
d2 / VR2
|
|
-
|
-
|
-
|
-
|
|
-
|
-
|
-
|
-
|
|
-
|
-
|
-
|
-
|
|
-
|
-
|
-
|
-
|
|
VRn
|
VLn
|
dn = VLn - VRn
|
dn / VRn
|
Results
1. Precision and inaccuracy of colorimetric methods used in
serum Mg determination in four different hospitals of Spain.
Precision and inaccuracy values obtained in all laboratories
analysed (four) were less than 10%, except in one hospital in which
the precision was less than 15% (table
2).
For all ranges of concentration established, the higher value of
precision was 12.52% (Zaragoza-Aragón, Spain) and the laboratory of
Santiago de Compostela (A Coruña, Spain) obtained the best value
(1.41%). Although all colorimetric methods analyzed gave acceptable
values of precision (< 15%), the chlorophosphorazo III one
used in this last laboratory is best adjusted to this
methodology.
In general, these results should be considered as acceptable
according to the validation requirement, because the precision and
inaccuracy were less than 10%.
2. Methodology for the estimation of some basic parameters of
validation in laboratories dedicated to Mg research.
The results found are presented in table
3.
The precedent table shows the Mg determination methods and
samples used in papers published in Magnesium Research during 2005
and 2006. With respect to the methods, 21 groups [15-35] used
spectrophotometric methods versus only five groups [10-14] that
reported colorimetric ones. Moreover, one group worked with the
method of Thuvasethakul & Wajjwalku [36], another with the ICAP
one [3] and, finally, one of them quotes the Mg2+
selective electrode [37]. In relation to the samples, the different
specimens studied can be observed. Compared with the clinical
laboratories, the researchers prefered to use spectrophotometric
methods for Mg determination in a great variety of samples.
Table 2 Summary of values of precision and inaccuracy
by the four laboratories analysed in four hospitals in Spain.
|
Santander (Cantabria)
|
Getafe (Madrid)
|
Zaragoza (Aragón)
|
Santiago de Compostela (A Coruña)
|
|
Precision
|
6.91%
|
6.66%
|
12.52%
|
1.41%
|
|
Inaccuracy
|
-3.49%
|
-3.87%
|
-3.46%
|
-1.27%
|
|
Method
|
- Colorimetric
- Xylidyl Blue/ADIVA
- 1650 Siemens Chemistry systems
|
Colorimetric
|
- Colorimetric
- Calmagite
- Beckman/LX-20 apparatus
|
- Colorimetric
- Chlorophosphonazo III/COBAS INTEGRA 400
|
|
Microrange
|
1.80-4.50 mg/100 mL
|
0.50-2.00 nmol/L
|
1.10-3.15 mg/100 mL
|
1.10-4.50 mg/100 mL
|
|
Intercomparison programme
|
BIO RAD Laboratories
|
CONTROL SEQC
|
BIO RAD Laboratories
|
BIO RAD Laboratories
|
Table 3 Magnesium determination methods and samples
used in papers published in Magnesium Research during 2005 and
2006.
|
Reference
|
Type of analytical method
|
Sample
|
|
Günther [15]
|
Null point method AAS
|
Erythrocytes
|
|
Dabrowski [10]
|
Colorimetric test xylidine blue
|
Serum
|
|
Chaudhary, et al. [36]
|
Method of Thuvasethakul &Wajjwalku
|
Blood
|
|
Nielsen, et al. [3]
|
ICAP-Ash Atom Com 1140
|
Serum-urine-feces
|
|
Dunicz-Sokolowska, et al. [16]
|
AAS method
|
Hair
|
|
Karakiewicz, et al. [17]
|
Flame AAS
|
Serum
|
|
Billyard, et al. [18]
|
Flame AAS
|
Plasma
|
|
Dunicz-Sokolowska, et al. [19]
|
In flame
|
Hair
|
|
Van Orden, et al. [20]
|
Flame Atomic Perkin Elmer 306.
|
Plasma
|
|
Grases, et al. [21]
|
Atomic emission spectrometry
|
Urine 24 hours
|
|
Pasternak, et al. [22]
|
Spectrophotometric methods
|
Blood
|
|
Pasternak, et al. [23]
|
Spectrophotometric methods
|
Blood
|
|
Ebel and Günther [24]
|
AAS
|
Erythrocytes
|
|
Katsumata, et al. [25]
|
AAS
|
Serum
|
|
Nemoto, et al. [26]
|
AAS
|
Feces and urine
|
|
Dunicz–Sokolowska, et al. [27]
|
Flame
|
Hair
|
|
Mousain-Bosc, et al. [11]
|
Colorimetric (chlorophosphonazo III)
|
Serum and intra-erythrocyte
|
|
Mousain-Bosc, et al. [12]
|
Colorimetric (chlorophosphonazo III)
|
Serum and intra-erythrocyte
|
|
Coudray, et al. [28]
|
Perkin Elmer
|
Blood
|
|
Sabbagh, et al. [13]
|
Xylidyl blue complexometric
|
Plasma
|
|
Sabbagh, et al. [14]
|
Xylidyl blue complexometric
|
Plasma
|
|
Kozielec, et al. [37]
|
Ion selective magnesium electrode
|
Blood
|
|
Pasternak, et al. [29]
|
Spectrophotometric methods
|
Blood
|
|
Korycinska, et al. [30]
|
Spectrophotometric methods
|
Blood
|
|
Iskra, et al. [31]
|
AAS
|
Serum
|
|
Katsumata, et al. [32]
|
AAS
|
Feces
|
|
Matsuzaki, et al. [33]
|
AAS
|
Feces
|
|
Feillet-Coudray, et al. [34]
|
AAS
|
|
|
Chiu, et al. [35]
|
Spectrophotometric methods
|
Drinking water
|
Discussion
The results of precision and inaccuracy of colorimetric methods
used in theclinical laboratories studied was obtained using
information from interlaboratory comparison programmes. This
methodology was implemented for the first time by our group and,
consequently, the results reported can not be compared with
previous ones. However, the precision and inaccuracy values
obtained for all laboratories analysed could be considered in the
same range as those reached by a classical method (experimental
estimation of precision: reproducibility, repetitively and accuracy
of the method studied) of validation. This agreement shows the
effectiveness of this new revalidation methodology for clinical
methods, much used for diagnostics in medicine.
At the moment, ISO 15189 is one recommendation that brings
progress in the obligations of clinical and/or biochemical
laboratories, specially the requirement of validation/revalidation
of analytical methods. Now, consequently, these laboratories can
see ISO 15189 as a guide to achieve accreditation and to begin to
practice some kind of methodology in order to validate/revalidate
their analytical methods.
In relation to Mg research laboratories, assuming the use of an
important variety of methodologies (spectrophotometry being the
main one) in different samples, they could participate in external
quality programmes or generate a specific methodology, as reported
in this paper. None of the papers referred in table 3 [3, 10, 11, 13-37] reported quality aspects
and values of parameters of validation with respect to the methods
used for Mg determination. In the near future, the values of
precision and inaccuracy estimated in the validation/revalidation
of any one method of Mg determination could be reported in all
publications, specifically in scientific papers, not only in those
concerning clinical laboratories.
Conclusion
The methodology that uses information on interlaboratory comparison
programmes to validate some colorimetric methods for Mg
determination in serum is useful, economical, fast and
recommendable. It avoids the need to disrupt routine work to carry
out validation trials, it does not require previous validation
trials, and it establishes the same validation parameters as those
reached by classical methodology.
This methodology could be applied to laboratories dedicated to
Mg research. Parameters of quality control such as precision and
inaccuracy could be reported in scientific papers independently of
the method used for Mg determination.
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