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Vol.
25 No. 4
July - August 2003
Physiological
Reference Values: A Shared Business?
by
Anders Kallner
The
theme of the Second European Symposium of the Clinical
Laboratory and in vitro Diagnostic Industry —held
6-7 February 2003 in Barcelona, Spain— was Physiological
Reference Values: A Shared Business? The symposium, chaired
by X. Fuentes-Arderiu, addressed several of the problems encountered
in establishing, teaching, and using physiological reference
values.
The
format of the symposium was unique: questions were posed to
a panel of experts who were then expected to address every
question. The audience was also invited to participate in
the discussion. The audience was comprised of scientists from
Spain and Europe and representatives of the major manufacturers
of reagents and instruments. The symposium was arranged by
the Catalan Association for Clinical Laboratory Sciences and
sponsored by IUPAC, IFCC, and major industries. Detailed proceedings
of the symposium will be published by Hyltoft-Petersen.1
Better
understanding of reference intervals is vitally important
because diagnosis of disease is frequently based on measurement
of biochemical quantities (components) of blood, serum, urine,
etc. Conclusions from obtained result are usually drawn by
comparison with results obtained in healthy individuals or,
less common, with individuals with specified diseases or conditions.
The acceptance of a reference point or reference interval
is as important for the physicians as a fix point was for
Archimedes: ‘Dòs moí poü stö
kaì kinö tën gen’ (‘give me a
fixed point and I will move the earth’). Originally
the profession used the term ‘normal value’ but
about 25 years ago it was agreed that normal was an improper
term since it was very hard to find any ‘normal’
individuals. The term was changed to ‘reference interval’
or ‘reference value’, depending on its use.2
Also the concept was changed and broadened by introducing
the concept of a ‘reference population’ and even
‘reference individuals’. We can now refer a reference
interval to a defined group of individuals who need not be
healthy but may suffer from a specific condition. The concept
also allows for relating the reference interval to age, gender
and ethnicity, as appropriate. The age and gender variation
is known for many commonly used quantities. 3
All
approaches to establishing reference intervals require large
groups of individuals (e.g., a minimum of 120 individuals
in the IFCC recommendation). It is an insurmountable task
for any individual laboratory to establish its own reference
values. Many published reference values are taken from textbooks
and lack an acceptable "audit trail" to the original
source. Thus, a reference interval is important for diagnosis
and screening of populations, but as soon as the individual
becomes a patient and undergoes treatment, the changes in
the values of certain quantities might be more important than
the absolute values. In these cases the patient becomes his
or her own reference.
Considering
the nature of the problem and the impact on health, diagnosis,
and economics, it is important that reference intervals are
scientifically based and appropriate for the actual situation
of the patient and the physician. Up until quite recently
establishment of reference intervals has been very loosely
regulated. However, the "EU IVD Directive 98/794
on in vitro diagnostic devices" can be interpreted as
placing the responsibility for establishing reference intervals
on the manufacturer of reagent kits and/or instruments:
"Where
appropriate, the instructions for use must contain [...]
the reference limits for the quantities being determined,
including a description of the appropriate reference population:
[…]."
Nonetheless,
the newly published international standard that is used for
the accreditation of medical laboratories EN/ISO 151895
states,
"Biological
reference intervals shall be periodically reviewed. If the
laboratory has reason to believe that a particular interval
is no longer appropriate for the reference population, then
an investigation shall be undertaken, followed, if necessary,
by corrective actions. A review of biological reference
intervals shall also take place when the laboratory changes
an examination procedure or pre-examination procedure, if
appropriate."
The
USA has a similar set of rules, whereby manufactures are obliged
to provide reference intervals and the laboratories are obliged
to prove that they are applicable in their environment.
The
symposium made clear that although the EU IVD Directive 98/794
puts the responsibility on industry to disseminate information
about physiological reference intervals for their measurement
systems, the industry is not well equipped or in a position
to collect the necessary raw data without collaboration from
the profession and medical laboratories. Panellists also argued
that reference intervals need to be locally modified by the
clinical laboratories. There are biological reasons for this,
such as ethnicity or dietary habits, and also methodological
reasons. Thus, there are many methods or method modifications
available that do not give the same results although they
set out to measure the same quantity (i.e., a deficient transferability
of results between laboratories).
From
a metrological point of view, transferability of results would
be achieved if the same calibrator were used. Globally that
requires that each calibrator is traceable to the same primary
standard, preferentially the realization of the SI unit. In
biological systems this hypothesis does not work very well.
A major reason is the complex nature of the sample and the
influence of the "matrix" defined as "all components
of a material system, except the analyte.6"
The matrix may not be consistent and may therefore have different
influences on the results from patient to patient. For years
the profession has tried to cope with the situation by external
quality assessment schemes (EQA, Europe) and proficiency testing
(PT, US). The effect has largely been to identify that there
are different measurement procedures on the market and that
their results differ. Most of these schemes use artificial
samples based on bovine or equine serum that may introduce
bias due to the changed matrix. Furthermore the EQA or PT
schemes are primarily designed to monitor the trueness of
results and measure the bias between laboratories and it serves
this purpose from a general point of view (e.g., CLIA 88 in
the USA). However, its usefulness for the individual laboratory
is limited.
An
alternate solution to approach the trueness, reduce the bias,
and meet the transferability demand would be to establish
a network of laboratory comparisons using patient material.
A NCCLS recommendation7 describes how such
comparisons can be made and a simplified and practical, yet
powerful procedure has been developed.8 A
very extensive experiment to align results of laboratories
and to create identical reference values in a region has been
carried out in the Nordic countries.9 Other
multicenter projects have been carried out.10-13
During the symposium, it was emphasized that establishment
or appropriate reviewing of reference intervals are very cumbersome
and expensive tasks and much would be gained if the reference
intervals could be established in a sustainable way. The costs
for establishing reference intervals can be exceedingly high
and it was argued that it could be an obligation of the heath
authorities to finance this procedure.
The
information technology is rapidly developing and most health
care systems either have or are developing laboratory information
systems (LIS) and hospital information systems (HIS). This
allows the laboratory to give more detailed information about
the patient and already many LIS give differentiated reference
intervals depending on the age and gender of the patient.
Most LIS provide cumulative reports and mark values outside
the reference intervals to the benefit of the physician and
patient.
It
might be useful to develop reference intervals for other than
healthy populations, e.g specific disease groups that can
be characterised by one or more biological properties. Typical
diseases are liver and kidney diseases, metabolic disturbances
like diabetes or screening procedures e.g. for Downs syndrome
(trisomy 21). Such reference intervals are usually based on
multivariate procedures, resulting in algorithms that require
special software, usually available in the LIS. The interpretation
of the results for a diseased individual is, however, much
more complex because most diseases develop – in a positive
or negative direction – and the values of the quantity
varies accordingly in a fashion that we presently do not have
sufficient structured information about. A solution might
be to report the deviation from a value or distribution of
values.
Conclusions
from the Symposium
The
rulings by the EU Directive 98/79/EC (Annex I 8.7) and the
EN/ISO 15189 (5.5.5) on the responsibilities of industry and
laboratories to create reference intervals are not clear and—as
written—difficult to interpret and follow. Symposium
attendees proposed that a request is sent to CEN TC 140 and
ISO TC 212 to clarify the demands of the directive and the
standard.
It was concluded that a document resolving this question should
address at least the following items:
-
selection of reference populations and definition of its
size
-
criteria for homogeneity/partitioning of the reference population
-
definition of pre-analytical conditions
-
definition of quantity
-
selection of method of measurement including calibrators,
reference materials, control procedures, and analytical
quality specifications
-
transferability and alignment of results
-
sustainability of reference intervals
-
calculation and presentation of reference intervals
In
addition, it was concluded that collaboration should be established
between professional organizations, industry, and health authorities
to achieve common reference intervals for homogeneous groups,
including multicenter reference intervals. Also, the concept
and interpretation of reference intervals should be taught
to pre- and post-graduate students of health sciences and
physicians.
References
1. Hyltoft-Petersen P., Report from the Second
European Symposium Clinical Laboratory and in vitro Diagnostic
Industry "Physiological Reference Values: A Shared Business?"
Clin Chem Lab Med 2003; 41(6):825-8
2. Solberg HE, PetitClerc C. International
Federation of Clinical Chemistry (IFCC), Approved recommendation
(1988) on the theory of reference values. Part 3. Preparation
of individuals and collection of specimens for the production
of reference values. J Clin Chem Clin Biochem. 1988;26(9):593-8.
3. Kallner A, Gustavsson E, Hendig E. Can
Age and Sex Related Reference Intervals be Derived for Non-Healthy
and Non-Diseased Individuals from Results of Measurements
in Primary Health Care? Clin Chem Lab Med 2000;38(7):633-654.
4. EU IVD Directive 98/79 on in vitro diagnostic
medical devices. Official J European Communities L331/1 98-12-07.
5. EN/ISO 15189 Medical laboratories –
Particular requirements for quality and competence. ISO, Geneva
2003.
6. Dybkaer R.Vocabulary for use in measurement
procedures and description of reference materials in laboratory
medicine. Eur J Clin Chem Clin Biochem 1977;35:141-73.
7. NCCLS EP9-A. Method comparison and bias
estimation Using Patient Samples; Approved Guideline. 1995
ISBN 1-56238.283-7.
8. Kallner A, Khorovskaya LA, Groth T. Can
the method comparison procedure in NCCLS recommendation EP9-A
be simplified? Clin Chem 2002;48:A174.
9. Nordic Reference Interval Project.
10. Andrew CE, Hanning I, McBain AM, Moody
D, Price A. A model for a multicentre approach to the derivation
of reference intervals for thyroid hormones and testosterone
for laboratories using indentical analysers. Clin Chem
Lab Med 2000;38:1013-9.
11. Ferre-Masferrer M, Fuentes-Arderiu X, Goma-Llongueras
M, Aluma-Trullas A, Aramendi-Ramos M, Castano-Vidriales JL,
Esteban-Salan M, Graells-Ferrer M, Jimenez-Gonzalez A, Jimenez-Lobo
C, Lopez-Esparza M, Pares-Pollan L, Regulez-Uranga M, Riesco-Prieto
M, Rodriguez-Camba A, Vidal-Martinez J. Regional reference
values for some quantities measured with the ADVIA Centaur
analyser. A model of co-operation between the in vitro diagnostic
industry and clinical laboratories. Clin Chem Lab Med.
2001;39:166-9.
12. Fuentes-Arderiu X, Ferre-Masferrer, M, Gonzalez-Alba JM,
Escola-Aliberas J, Balsells-Rosello D, Blanco-Cristobal C,
Gonzalez-Cruz E, Ibarz-Escuer M, Latorre-Marcellan P, Lugo-Arocena
J, Mar-Medina C, Muros-de-Fuentes M, Vicens-Manero M. Related
Articles, Links Multicentric reference values for some quantities
measured with Tina-Quant reagents systems and RD/Hitachi analysers.
Scand J Clin Lab Invest. 2001;61:273-6.
13. Fuentes-Arderiu X, Ferre-Masferrer, González-Alba
JM, Villarino-González MI, Arrimadas-Esteban E, Cabrero-Olivé
D, Cándenas-Arroyo M, García-García D,
García-Lario JV, Idoate-Cervantes I, León-López
C, López-Lazareno N, Mar-Medina C, Martí-Marcet
I, Mauri-Dot M, Pérez-Valero V, Reta-Manterola A, Sánchez-Eixeres
MR. Multicentric reference values for some quantities measured
with the Elecsys 2010 analyser. Clin Chim Acta 2001;304:143-6.
Anders
Kallner <[email protected]>
is from the Department of Clinical Chemistry of the Karolinska
Hospital in Stockholm, Sweden. He is the current president
of the IUPAC Division of Human and Human Health.
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last modified 30 June 2003.
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