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Approved recommendation (1987) on the theory of reference values. Part 2. Selection of individuals for the production of reference values
THE INTERNATIONAL FEDERATION CLINICAL CHEMISTRY
OF
IFCC Section (1987) Recent IFCC Publications
Published by SCIENCE PUBLISHERS on behalf of FEDERATION OF CLINICAL
ELSEVIER THE INTERNATIONAL
CHEMISTRY
The exclusive ‘I:; for all languages Federation of Clinical Chemistry
and
countries
is vested
in The
International
All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means. electronic, mechanical, photocopying. recording, or otherwise, without prior permission of the copyright owner.
IFCC Sections (1978) No. I No. 2
Clin. Chim. Acta (1978) Vol. 83, IWF-202F Vol. 87,&9F-46SF
1FCC Sections (1979) No. 1 No. 2 No. 3
Clin. Chim. Acta (1979) Vol. 95, 153F-188F Vol. 96,155F-204F Vol. 98. 127F-186F
IFCC Sections (1980) No. 1 No. 2 No. 3 No. 4
Clin. Chim. Acta (1980) Vol. 103, 239F-258F Vol. lO5,147F-172F Vol. 106.107F-l2OF Vol. 10&499F-539F
IFCC Section (1981)
Clin. Chim. Acta (1981) Vol. lO9,103F-124F
IFCC Sections (1982) No. 1 No. 2
Clin. Chim. Acta (1982) Vol. 119,35lF-362F Vol. l22,109F-123F
IFCC Sections (1983) No. 1 No. 2 No. 3 No. 4 No. 5
Clin. Chim. Acta (1983) Vol. l27,423F-448F Vol. 129, 245F-249F Vol. 131.349F-3591” Vol. 134.369F-386F Vol. 135, 313F-367F
IFCC Sections (1984) No. 1 No. 2 No. 3 Recent IFCC publications
Clin. Chim. Acta (1984) Vol. 137,95F-114F Vol. l37,369F-386F Vol. l39,203F-230F Vol. 143,69F-72F
IFCC Sections (1987) No. 1 No. 2 No. 3 No. 4
Clin. Chim. Acta (1987) Vol. 165.95-118 170, s3 170, s13 170. s33
Clinica Chimica Acta, 170 (1987) S3-S12 fi) 1987, Elsevier Science Publishers B.V. (Biomedical Division)
INTERNATIONAL
FEDERATION
s3
OF CLINICAL CHEMISTRY
’
SCIENTIFIC COMMITTEE, CLINICAL SECTION EXPERT PANEL ON THEORY OF REFERENCE VALUES (EPTRV) 2
Approved recommendation ( 1987) on the theory of reference values. Part 2. Selection of individuals for the production of reference values. Prepared
forpubiisation
by: C. PetitClerc, Serrwe de Biochimie, Hate/-Dieu de Montreal, Quebec, Canada and H. E. Solberg, Department of Clinical Chemistv, Rlkshospitalet, Oslo, Norwqv
Contents Preface ...................................... 1. introduction ................................ 2. Selection of reference individuals .................. 3. Exclusion of indi~du~s from the reference sample group 4. Partitioning of the reference sample groups ........... 5. Conclusion ................................. 6. References ..................................
. ..
s3 S4 S4 S6 S8 SlO SlO
Preface
This paper is the second in a series of Recommendations on the Theory of Reference Values. Other parts deal with: Part 1. The Concept of Reference Values [2]. Part 3. Preparation of Individuals and Collection of Specimens for the Production of Reference Values.
’ The exclusive 0 for all languages and countries is vested in The International Federation of Clinical Chemistry. ’ Members of the Expert Panel during the preparation of this R~o~endation were as follows: R. Grlsbeck (FI) 1970-1978 (Chairman 19751978), N. Montalbetti (IT) 1982-1986, C. PetitClerc (CA) 1977-1986, G. Siest (FR) 1973-1979, H.E. Solberg (NO) 1977-1986 (Chairman since 1979). D. Stamm (DE) 1979-1986, P. Wilding (US) 1976-1984. This paper is based on a Provisional Recommendation [I ] which has been revised to take account of the comments received, and it has been accepted by the Council of the lntemational Federation of Clinical Chemistry by mail ballot in March 1987. Correspondence and requests for reprints to: Claude PetitClerc, M.D., Ph.D.. Service de Biochimie, Hotel-Dieu de Montreal,~3840 St. Urbain Street, Montreal, Quebec, Canada H2W lT8.
s4
Part 4. Control of Analytical Variation in the Production, Transfer and Application of Reference Values. Part 5. Statistical Treatment of Collected Reference Values. Determination of Reference Limits [3]. Part 6. Presentation of Observed Values Related to Reference Values [4]. A guide to the Documents is also in preparation. The Expert Panel on Theory of Reference Values (EPTRV) was created in 1970 by the Committee on Standards (at present: Scientific Committee) of the Intemational Federation of CIinical Chemistry (IFCC). Its task was to develop a nomenclature and recommend procedures for the production or reference values and their treatment, and presentation of observed values in relation to reference data. The first document in the above-mentioned series describes the subject of reference values and defines various terms. It should be consulted prior to the reading of the present document for a thorough understanding. 1. Introduction 1.1. Traditionally, reference values from supposedly healthy subjects have been widely used in the clinical laboratory sciences. However, health is not a well-defined condition and has come to be regarded as relative. Often, the border between health and disease is diffuse, especially in relation to aging [5,6]. Reference values may be used to evaluate the state of health of individuals and populations, to identify people at risk for disease, to help in decision making in clinical medicine, and to be used for various scientific purposes. It must be realized that in selecting reference individuals, the criteria of health applied are dictated by the aim of the laboratory investigation. Thus, reference individuals may not always be healthy subjects.
The conditions under which the reference values are collected should be completely described and standardized, according to the intended use. The description should include: _ the characteristics of the reference individuals and the reference sample group, such as age, sex, body mass, genetic, ethnic, and socio-economic factors; _ the physiological and environmental conditions of the reference individuals and the time of specimen collection (see Part 3 of the EPTRV recommendations); _ the procedure for the collection and treatment of the specimen (see Part 3 of the EPTRV recommendations); _ the analytical methods and the quality control procedure (see Part 4 of the EPTRV recommendations). 2. Selection of reference individuals The selection of individuals approached from many angles, resources available.
for the production of reference values has been according to the different philosophies, needs, and
S5
2.1. Two types of selection are used (see Fig. 1): _ The ‘a posterior? (retrospective) selection of individuals from a large sample of population obtained randomly [7-111 or non-randomly [12-161 and followed by partitioning and exclusion, according to the characteristics of the reference sample group. The partitioning and exclusion criteria will differ depending on the type of quantity to be studied. - The ‘a priori’ (prospective) selection from a general population using established exclusion and partitioning criteria as determined by previous studies on the same population, or obtained from the literature.
2.2. A posteriori selection, ideally from large random-sample groups, is ideal for the study of exclusion and partitioning criteria; as such sample groups represent the
A posteriori selection
A
priori
F Exclusion
PartSAl
e statistical
treatment
Biological factors
Exclusion
Reference values
I?3tbOlOgy
f
PaltitiOll
I
StatiStiUl treatment
I ’ Reference values Fig. 1. The two types of selection
of reference
individuals
a posteriori
and a priori
selection.
S6
main elements of the general population (e.g., rural or urban environment, socioeconomic classes, ethnic groups, etc.). Few laboratories have access to the large population samples required and the resources to analyse them. 2.3. A priori selection is far more convenient but requires known, or arbitrarily partitioning and exclusion criteria. As more data become available from literature. this type of selection will be largerly favoured. 2.4. For both types of selection, the nature of the exclusion criteria and The a posteriori selection is more healthy individuals, but the a priori
set the
size of the sample groups is determined by the the number of partitioning criteria to be applied. suited to production of reference values from selection could be applied to all situations.
2.5. Production of reference values from hospitalized populations has been described by Martin et al [17]. For each quantity, the reference sample group is isolated by statistical treatment of mixed populations of selected individuals, according to stated criteria. 2.6. In cases where population-based reference values are not available or do not apply, individual reference values may be collected as described by Winkel [18]. 2. 7. It must be emphasized that extremely valuable information will be obtained longitudinal studies of survivors within a given population [5].
by
3. Exclusion of individuals from the reference sample group 3.1. Many factors contribute to biological variability [19,20] and may entail the exclusion or partitioning of reference individuals. Moreover, the use to which the reference values are to be put will determine the exclusion criteria to be applied. For example, epidemiological surveys and preventive medicine programs require reference values from healthy individuals, thus requiring the exclusion of individuals suffering from systemic diseases and pathophysiological disorders. Another example is the situation where a clinical decision requires reference values which discriminate between specific disorders.
Thus, depending on the intended uses of the reference values and on the type of quantity measured, some or all of the following exclusion criteria should be applied. Otherwise, the measured values may show shift in location and/or increased dispersion.
Sl
For specific purposes, the criteria described below may be considered as partitioning criteria, to obtain reference values from well identified reference sample groups such as drinkers, smokers, women on oral contraceptives. pregnant women. obese individuals, etc. 3.3. Pathophysiological states Individuals suffering from systemic disease and pathophysiological disorders such as renal failure, congestive heart disease, chronic respiratory diseases, liver diseases, malabsorption syndromes, and nutritional anemias should be excluded by clinical examination, laboratory investigation and/or questionnaire at the time of interview and specimen collection [11,21]. The extensive list of exclusion criteria prepared by the Scandinavian Committee on Reference Values [22] may be used as a guide when defining the criteria to be used in an actual project. 3.4. Intake of pharmacologically active agents Individuals receiving agents which are taken for treatment of diseases, as replacement or supplementation therapy, or drug abuse should be subject to exclusion. The list of agents also includes oral contraceptives, alcohol, and tobacco. These substances may affect the physiology and the metabolism of individuals, and alter the blood and urine values of many quantities (see Part 3 of the EPTRV recommendations). An example of such an effect on the plasma value of some constituents of supposedly healthy individuals is given in Fig. 2. Since alcoholism induces major metabolic disturbances, drinkers with excessive intake must be excluded. Moreover,
1
I
II
30
I
70
M
Alkahne 2
1
I 460
600
L
1
,’ I L
I
I
1
110
130
150
phasphatase
I ,I CL90
Phosphates
6M Albumin
I 90
I
[U/I 1
I I lso rzt
[$I
/I]
750 I I
I
I 5 Glucose
[mmat?
II
, 0
Fig. 2. Influence of drug intake on reference intervals (2.5tb, 50th, 97.5th percentiles for 20 to 50-yr-old fasting men. Group 1, Subjects taking drugs; group 2, Subjects not taking drugs. From Siest and Henny
1251.
S8
in view of the variable effects of moderate alcohol intake in individuals, it is recommended to exclude also those who have ingested alcoholic beverages within 24 h prior to blood collection (see Part 3 of the EPTRV recommendations). 3.5. Modified physiological states Individuals should be excluded if they fall in any one of the following categories: - pregnancy (may induce important hormonal, metabolic, and physiological changes); - exercise or physical training (may give rise to short term or long term effects such as dehydration, tissue damage, and delayed puberty in adolescents); - psychological and mental disorders such as severe stress and depression (may be accompanied by hormonal and metabolic imbalances); _ food intake prior to blood collection may change the concentrations of serum components (see Part 3 of the EPTRV recommendations). 3.6. Other factors Obesity, hypertension or other factors yet to be identified may indicate that the subject is at risk for particular diseases. 4. Partitioning of the reference sample groups The need for partitioning the reference groups may differ with the quantities measured and the intended uses of the reference values. Subclassification should be limited to reference values which display significant differences in location or dispersion [17,23]. An example is given in Table I. 4. I. Age and sex Age should not necessarily be categorized by equal intervals. Age ranges should be chosen, taking into account the variation with age in the quantity measured; they
TABLE
I
Effect of partitioning concentration Category
criteria
on reference
(criteria)
1. Age (4-5 yr vs 30-60 yr *) (20-30 yr vs 60-100 yr *) 2. Male vs female (age 20-30) 3. Menopause (age 45-55) 4. Excess weight ( > 15% ideal) 5. Oral contraceptives (treated vs control) 6. Exercise 7. Meals
values for the quantity:
serum alkaline
Values ( f W) relative
to median
phosphatase,
of total population
Male
Female
-75% + 13% +19% -
- 79% +44%
+2.5% _ 0% 0%
catalytic
+21% + 10% -7.5% + 1.3% + Cl%
*
s9
should be particularly small over periods such as puberty and menopause. Bone age, height, and body mass are better indicators than actual age for categorizing children. Variations due to deviations from the ideal body mass should be distinguished from changes with age and/or sex (e.g. S-creatinine) [24]. 4.2. Genetic, socio-economic
and environmental
criteria
4.2.1. Subclassification, according to ethnic origins, geographical phology, or pigmentation may be of value for some quantities.
location,
mor-
4.2.2. Genetic markers such as blood groups (ABO) and histocompatibility antigens (HLA) may be more appropriate. The presence or absence of phenotypes of plasma proteins, tissue enzymes could be useful to obtain homogeneous reference sample groups (e.g. cY,-antitrypsin, apolipoprotein B, phenylalanine hydroxylase). 4.2.3. In some instances, adaptation of the individuals to their ecologic environment as to their socio-economic status may be the source of major differences: S-cholesterol concentration of natives of Asia and S-immunoglobulins of African blacks before and after emigration to Europe or America [26]. Also vegetarians may have much lower S-cholesterol and S-urate concentrations than people on a mixed diet. 4.2.4. Long-term dietary effects must also be considered separately from short-term ones which are described in Part 3 of the EPTRV recommendations.
the
4.3. Biological criteria 4.3.1. The haemodynamics, kidney perfusion, and hormonal balance are different when the subject is standing or supine (Part 3 of the EPTRV recommendations). Whenever indicated, separate reference sample groups should be selected from ambulant and hospitalized population (e.g. S-albumin, substance concentration). 4.3.2. For some quantities, it may be necessary to consider chronobiological factors as criteria for partitioning (e.g. S-cortisol, substance concentration). 4.4. Reference state The notion of reference state can be used to facilitate population comparisons and to study transferability of reference value data. For such comparisons, the influence of biological variations should be minimal. Most constituents are subject to the least biological variations between the ages of 20 and 30 yr after other variation factors have been excluded. To qualify for the Reference State, individuals shall be 20-30 years of age, of ideal body mass, fasted for 10 h, taking no medication, consuming less than 45 g alcohol per day, smoking less than 12 cigarettes per day, and with no apparent illness [27].
SlO
5. Conclusion Production of reference values from any population or individuals requires appropriate selection and often subclassification. This can only be made by careful description of the characteristics of the reference individuals and by the application of clearly stated criteria. It is emphasized that depending on the uses of the reference values and the type of quantity, part or all, of the guidelines outlined in this document should be used.
6. References 1 PetitCIerc C, Wilding P. Provisional Recommendation on the theory of reference values. Part 2. Selection of individuals for the production of reference values. Clin Chim Acta 1984 139:205F-213F; J CIin Chem Clin Biochem 1984;22:203-208. Republished in Clin Chem Newslett (Milan) 1985;5:91-96. 2 Solberg HE. Approved Recommendation (1986) on the theory of reference values. Part 1. The concept of reference Values. Ann Biol CIin 1986;45:237-241, Clin Chim Acta 1987;165:111-118; J Clin Chem Clin Biochem 1987;25:in press. 3 Solberg HE. Approved recommendation (1987) on the theory of reference values. Part 5. Statistical treatment of collected reference values. Determination of reference limits. J Clin Chem Chim Biochem 1987;25:submitted for publication. 4 Dybkar R, Solberg HE. Approved recommendation (1987) on the theory of reference values. Part 6. Presentation of observed values related to reference values. 1987;submitted for publication. 5 Grasbeck R. Health as seen from the laboratory. In: Grasbeck R. Alstrom T, eds. Reference values in laboratory medicine. The current state of the art, Chichester: John Wiley and Sons, 1981;17-23. 6 Larsen 0. Separation health and illness - a conceptual framework. In: Grasbeck R, Alstrom T, eds. The current state of the art, Chichester: John Wiley and Sons, 1981;33-43. 7 Munan L, Kelly A, Petitclerc C. Population serum urate levels and their correlate; the Sherbrooke regional study. Am J Epidemiol 1976;103:369-382. 8 Munan L. Population based reference data. In: Rubin M, Benson ES, eds. Logic and economics of clinical laboratory use, New York: Elsevier, 1978;117-128. 9 Gardner M, Scott R. Frequency distribution and reference values of plasma alkaline phosphatase activity in the adult population of a Scottish new town. J Clin Pathol 1978;31:1202-1206. 10 Gardner M, Scott R. Age- and sex-related reference ranges for eight plasma constituents derived from randomly selected adults in a Scottish new town. J Clin Pathol 1980;33:380-385. 11 Berg B, Nilsson JE, Solberg HE, Tryding N. Practical experience in selection and preparation of reference individuals: empirical testing of the provisional Scandinavian recommendations. In: Grasbeck R, Alstrom T, eds. Reference vales in laboratory medicine. The current state of the art. Chichester: John Wiley and Sons, 1981;55-64. 12 Schiele F, Guilmin AM, Detienne H, Siest G. Gamma-glutamyltransferase activity in plasma: statistical distributions, individual variations and reference intervals. CIin Chem 1977;25:1023-1028. 13 Siest G. La biologic de I’homme sain. In: Metais P, et al, eds. Biochimie clinique. Tome 2: Biochimie physiopathologique. Lyon: SIMEP. 14 Steinmetz J. Panek E, Siest G, Gueguen R. Factors affecting the concentration of triacylglycerols (triglycerides) in plasma: reference values for adults. Clin Chem 1979;25:924-932. 15 Reed AH, Cannon DC, Winkelman JW, Bhasin YP, Henry RJ, Pileggi VJ. Estimation of normal ranges from a controlled sample survey. 1. Sex and age-related influence on the SMA 12/60 screemng group of tests. Clin Chem 1972;18:57-66. 16 Wilding P, Rollason JG, Robinson D. Patterns of change for various biochemical constituents detected in well population screening. Clin Chim Acta 1972;41:375-387.
17 Martin HF, Hologgitas JV, DriscoU J, Fanger H, Gudzinowicz BJ, Bamett R, Weisbrot I. Reference values based on populations accessible to hospitals. Grasbeck R, Alstrom T, eds. Reference values in Laboratory medicine. The current state of the art. Chichester: Wiley, 1981;233-262. 18 Winkel P. The use of the subject as his own Referent. In: Grasbeck R, Alstrom T, eds. Reference values in laboratory medicine. The current state of the art. Chichester: Wiley, 1981:65-78. 19 Statland BE. Winkel P. Selected preanalytical sources of variation. Grasbeck R, Alstrom T, eds. Reference values in laboratory medicine. The current state of the art. Chichester: Wiley, 1981;127-137. 20 Bretaudiere JP, Buret J, Favre R, Gueguen R, PetitClerc C, Sachs C, Siest G, Vemet M, Zender R. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology: biological variations in laboratory tests. Ann Biol Clin 1979;37:229-239. 21 Drodowsky M, Sachs Ch. Factors to be taken into considerations for blood sampling in view of establishing reference values. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology (Vemet-Nyssen M, Blin G, Buret J, Drodowsky M, Gueguen R, Guize L, Panek E, PetitClerc C, Sachs C, Zender R). Ann Biol Clin 1980;38:251-265. 22 Alstrom T, Grasbeck R, Hjelm M, Skandsen S. Recommendations concerning the collection of reference values in clinical chemistry and activity report by the Committee on Reference Values of the Scandinavian Society for Clinical Chemistry and Clinical Physiology. Stand J Clin Lab Invest 1975;35:suppl 144. 23 Harris EK. Some theory of reference values. I. Stratified (categorized) normal ranges and a method for following and individual’s clinical laboratory values. Clin Chem 1975;21:1457-1464. 24 Munan L, Kelly A, PetitClerc C, Billon B. Association with body weight of selected chemical constituents in blood: epidemiologic data. Clin Chem 1978;24:772-777. 25 Siest G, Henny J. Production of reference values in healthy subjects. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology (Sachs Ch, Aellig A, Albert A, Blin G, Buret J, Daubrosse E, Drodowsky M. Gouget B, Gueguen R, Guize L, Henny J, PetitClerc C, Queralto J, Ramon F, Siest G, Vemet M). Ann Biol Clin 1981;39:235-244. 26 Wilding P, Bailey A. The normal range. In: Williams DL, Nunn RF. Marks V, eds. Scientific Foundations of Clinical Biochemistry, Vol. 1. London: Heinemann Medical Books Ltd. 1979;451-459. 27 Siest G, Schiele F, Henny J, Young DS. Interpretation of clinical laboratory tests, Davis: Biomedical Publications, 1985;18-22.
OF
IFCC Section (1987) Recent IFCC Publications
Published by SCIENCE PUBLISHERS on behalf of FEDERATION OF CLINICAL
ELSEVIER THE INTERNATIONAL
CHEMISTRY
The exclusive ‘I:; for all languages Federation of Clinical Chemistry
and
countries
is vested
in The
International
All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means. electronic, mechanical, photocopying. recording, or otherwise, without prior permission of the copyright owner.
IFCC Sections (1978) No. I No. 2
Clin. Chim. Acta (1978) Vol. 83, IWF-202F Vol. 87,&9F-46SF
1FCC Sections (1979) No. 1 No. 2 No. 3
Clin. Chim. Acta (1979) Vol. 95, 153F-188F Vol. 96,155F-204F Vol. 98. 127F-186F
IFCC Sections (1980) No. 1 No. 2 No. 3 No. 4
Clin. Chim. Acta (1980) Vol. 103, 239F-258F Vol. lO5,147F-172F Vol. 106.107F-l2OF Vol. 10&499F-539F
IFCC Section (1981)
Clin. Chim. Acta (1981) Vol. lO9,103F-124F
IFCC Sections (1982) No. 1 No. 2
Clin. Chim. Acta (1982) Vol. 119,35lF-362F Vol. l22,109F-123F
IFCC Sections (1983) No. 1 No. 2 No. 3 No. 4 No. 5
Clin. Chim. Acta (1983) Vol. l27,423F-448F Vol. 129, 245F-249F Vol. 131.349F-3591” Vol. 134.369F-386F Vol. 135, 313F-367F
IFCC Sections (1984) No. 1 No. 2 No. 3 Recent IFCC publications
Clin. Chim. Acta (1984) Vol. 137,95F-114F Vol. l37,369F-386F Vol. l39,203F-230F Vol. 143,69F-72F
IFCC Sections (1987) No. 1 No. 2 No. 3 No. 4
Clin. Chim. Acta (1987) Vol. 165.95-118 170, s3 170, s13 170. s33
Clinica Chimica Acta, 170 (1987) S3-S12 fi) 1987, Elsevier Science Publishers B.V. (Biomedical Division)
INTERNATIONAL
FEDERATION
s3
OF CLINICAL CHEMISTRY
’
SCIENTIFIC COMMITTEE, CLINICAL SECTION EXPERT PANEL ON THEORY OF REFERENCE VALUES (EPTRV) 2
Approved recommendation ( 1987) on the theory of reference values. Part 2. Selection of individuals for the production of reference values. Prepared
forpubiisation
by: C. PetitClerc, Serrwe de Biochimie, Hate/-Dieu de Montreal, Quebec, Canada and H. E. Solberg, Department of Clinical Chemistv, Rlkshospitalet, Oslo, Norwqv
Contents Preface ...................................... 1. introduction ................................ 2. Selection of reference individuals .................. 3. Exclusion of indi~du~s from the reference sample group 4. Partitioning of the reference sample groups ........... 5. Conclusion ................................. 6. References ..................................
. ..
s3 S4 S4 S6 S8 SlO SlO
Preface
This paper is the second in a series of Recommendations on the Theory of Reference Values. Other parts deal with: Part 1. The Concept of Reference Values [2]. Part 3. Preparation of Individuals and Collection of Specimens for the Production of Reference Values.
’ The exclusive 0 for all languages and countries is vested in The International Federation of Clinical Chemistry. ’ Members of the Expert Panel during the preparation of this R~o~endation were as follows: R. Grlsbeck (FI) 1970-1978 (Chairman 19751978), N. Montalbetti (IT) 1982-1986, C. PetitClerc (CA) 1977-1986, G. Siest (FR) 1973-1979, H.E. Solberg (NO) 1977-1986 (Chairman since 1979). D. Stamm (DE) 1979-1986, P. Wilding (US) 1976-1984. This paper is based on a Provisional Recommendation [I ] which has been revised to take account of the comments received, and it has been accepted by the Council of the lntemational Federation of Clinical Chemistry by mail ballot in March 1987. Correspondence and requests for reprints to: Claude PetitClerc, M.D., Ph.D.. Service de Biochimie, Hotel-Dieu de Montreal,~3840 St. Urbain Street, Montreal, Quebec, Canada H2W lT8.
s4
Part 4. Control of Analytical Variation in the Production, Transfer and Application of Reference Values. Part 5. Statistical Treatment of Collected Reference Values. Determination of Reference Limits [3]. Part 6. Presentation of Observed Values Related to Reference Values [4]. A guide to the Documents is also in preparation. The Expert Panel on Theory of Reference Values (EPTRV) was created in 1970 by the Committee on Standards (at present: Scientific Committee) of the Intemational Federation of CIinical Chemistry (IFCC). Its task was to develop a nomenclature and recommend procedures for the production or reference values and their treatment, and presentation of observed values in relation to reference data. The first document in the above-mentioned series describes the subject of reference values and defines various terms. It should be consulted prior to the reading of the present document for a thorough understanding. 1. Introduction 1.1. Traditionally, reference values from supposedly healthy subjects have been widely used in the clinical laboratory sciences. However, health is not a well-defined condition and has come to be regarded as relative. Often, the border between health and disease is diffuse, especially in relation to aging [5,6]. Reference values may be used to evaluate the state of health of individuals and populations, to identify people at risk for disease, to help in decision making in clinical medicine, and to be used for various scientific purposes. It must be realized that in selecting reference individuals, the criteria of health applied are dictated by the aim of the laboratory investigation. Thus, reference individuals may not always be healthy subjects.
The conditions under which the reference values are collected should be completely described and standardized, according to the intended use. The description should include: _ the characteristics of the reference individuals and the reference sample group, such as age, sex, body mass, genetic, ethnic, and socio-economic factors; _ the physiological and environmental conditions of the reference individuals and the time of specimen collection (see Part 3 of the EPTRV recommendations); _ the procedure for the collection and treatment of the specimen (see Part 3 of the EPTRV recommendations); _ the analytical methods and the quality control procedure (see Part 4 of the EPTRV recommendations). 2. Selection of reference individuals The selection of individuals approached from many angles, resources available.
for the production of reference values has been according to the different philosophies, needs, and
S5
2.1. Two types of selection are used (see Fig. 1): _ The ‘a posterior? (retrospective) selection of individuals from a large sample of population obtained randomly [7-111 or non-randomly [12-161 and followed by partitioning and exclusion, according to the characteristics of the reference sample group. The partitioning and exclusion criteria will differ depending on the type of quantity to be studied. - The ‘a priori’ (prospective) selection from a general population using established exclusion and partitioning criteria as determined by previous studies on the same population, or obtained from the literature.
2.2. A posteriori selection, ideally from large random-sample groups, is ideal for the study of exclusion and partitioning criteria; as such sample groups represent the
A posteriori selection
A
priori
F Exclusion
PartSAl
e statistical
treatment
Biological factors
Exclusion
Reference values
I?3tbOlOgy
f
PaltitiOll
I
StatiStiUl treatment
I ’ Reference values Fig. 1. The two types of selection
of reference
individuals
a posteriori
and a priori
selection.
S6
main elements of the general population (e.g., rural or urban environment, socioeconomic classes, ethnic groups, etc.). Few laboratories have access to the large population samples required and the resources to analyse them. 2.3. A priori selection is far more convenient but requires known, or arbitrarily partitioning and exclusion criteria. As more data become available from literature. this type of selection will be largerly favoured. 2.4. For both types of selection, the nature of the exclusion criteria and The a posteriori selection is more healthy individuals, but the a priori
set the
size of the sample groups is determined by the the number of partitioning criteria to be applied. suited to production of reference values from selection could be applied to all situations.
2.5. Production of reference values from hospitalized populations has been described by Martin et al [17]. For each quantity, the reference sample group is isolated by statistical treatment of mixed populations of selected individuals, according to stated criteria. 2.6. In cases where population-based reference values are not available or do not apply, individual reference values may be collected as described by Winkel [18]. 2. 7. It must be emphasized that extremely valuable information will be obtained longitudinal studies of survivors within a given population [5].
by
3. Exclusion of individuals from the reference sample group 3.1. Many factors contribute to biological variability [19,20] and may entail the exclusion or partitioning of reference individuals. Moreover, the use to which the reference values are to be put will determine the exclusion criteria to be applied. For example, epidemiological surveys and preventive medicine programs require reference values from healthy individuals, thus requiring the exclusion of individuals suffering from systemic diseases and pathophysiological disorders. Another example is the situation where a clinical decision requires reference values which discriminate between specific disorders.
Thus, depending on the intended uses of the reference values and on the type of quantity measured, some or all of the following exclusion criteria should be applied. Otherwise, the measured values may show shift in location and/or increased dispersion.
Sl
For specific purposes, the criteria described below may be considered as partitioning criteria, to obtain reference values from well identified reference sample groups such as drinkers, smokers, women on oral contraceptives. pregnant women. obese individuals, etc. 3.3. Pathophysiological states Individuals suffering from systemic disease and pathophysiological disorders such as renal failure, congestive heart disease, chronic respiratory diseases, liver diseases, malabsorption syndromes, and nutritional anemias should be excluded by clinical examination, laboratory investigation and/or questionnaire at the time of interview and specimen collection [11,21]. The extensive list of exclusion criteria prepared by the Scandinavian Committee on Reference Values [22] may be used as a guide when defining the criteria to be used in an actual project. 3.4. Intake of pharmacologically active agents Individuals receiving agents which are taken for treatment of diseases, as replacement or supplementation therapy, or drug abuse should be subject to exclusion. The list of agents also includes oral contraceptives, alcohol, and tobacco. These substances may affect the physiology and the metabolism of individuals, and alter the blood and urine values of many quantities (see Part 3 of the EPTRV recommendations). An example of such an effect on the plasma value of some constituents of supposedly healthy individuals is given in Fig. 2. Since alcoholism induces major metabolic disturbances, drinkers with excessive intake must be excluded. Moreover,
1
I
II
30
I
70
M
Alkahne 2
1
I 460
600
L
1
,’ I L
I
I
1
110
130
150
phasphatase
I ,I CL90
Phosphates
6M Albumin
I 90
I
[U/I 1
I I lso rzt
[$I
/I]
750 I I
I
I 5 Glucose
[mmat?
II
, 0
Fig. 2. Influence of drug intake on reference intervals (2.5tb, 50th, 97.5th percentiles for 20 to 50-yr-old fasting men. Group 1, Subjects taking drugs; group 2, Subjects not taking drugs. From Siest and Henny
1251.
S8
in view of the variable effects of moderate alcohol intake in individuals, it is recommended to exclude also those who have ingested alcoholic beverages within 24 h prior to blood collection (see Part 3 of the EPTRV recommendations). 3.5. Modified physiological states Individuals should be excluded if they fall in any one of the following categories: - pregnancy (may induce important hormonal, metabolic, and physiological changes); - exercise or physical training (may give rise to short term or long term effects such as dehydration, tissue damage, and delayed puberty in adolescents); - psychological and mental disorders such as severe stress and depression (may be accompanied by hormonal and metabolic imbalances); _ food intake prior to blood collection may change the concentrations of serum components (see Part 3 of the EPTRV recommendations). 3.6. Other factors Obesity, hypertension or other factors yet to be identified may indicate that the subject is at risk for particular diseases. 4. Partitioning of the reference sample groups The need for partitioning the reference groups may differ with the quantities measured and the intended uses of the reference values. Subclassification should be limited to reference values which display significant differences in location or dispersion [17,23]. An example is given in Table I. 4. I. Age and sex Age should not necessarily be categorized by equal intervals. Age ranges should be chosen, taking into account the variation with age in the quantity measured; they
TABLE
I
Effect of partitioning concentration Category
criteria
on reference
(criteria)
1. Age (4-5 yr vs 30-60 yr *) (20-30 yr vs 60-100 yr *) 2. Male vs female (age 20-30) 3. Menopause (age 45-55) 4. Excess weight ( > 15% ideal) 5. Oral contraceptives (treated vs control) 6. Exercise 7. Meals
values for the quantity:
serum alkaline
Values ( f W) relative
to median
phosphatase,
of total population
Male
Female
-75% + 13% +19% -
- 79% +44%
+2.5% _ 0% 0%
catalytic
+21% + 10% -7.5% + 1.3% + Cl%
*
s9
should be particularly small over periods such as puberty and menopause. Bone age, height, and body mass are better indicators than actual age for categorizing children. Variations due to deviations from the ideal body mass should be distinguished from changes with age and/or sex (e.g. S-creatinine) [24]. 4.2. Genetic, socio-economic
and environmental
criteria
4.2.1. Subclassification, according to ethnic origins, geographical phology, or pigmentation may be of value for some quantities.
location,
mor-
4.2.2. Genetic markers such as blood groups (ABO) and histocompatibility antigens (HLA) may be more appropriate. The presence or absence of phenotypes of plasma proteins, tissue enzymes could be useful to obtain homogeneous reference sample groups (e.g. cY,-antitrypsin, apolipoprotein B, phenylalanine hydroxylase). 4.2.3. In some instances, adaptation of the individuals to their ecologic environment as to their socio-economic status may be the source of major differences: S-cholesterol concentration of natives of Asia and S-immunoglobulins of African blacks before and after emigration to Europe or America [26]. Also vegetarians may have much lower S-cholesterol and S-urate concentrations than people on a mixed diet. 4.2.4. Long-term dietary effects must also be considered separately from short-term ones which are described in Part 3 of the EPTRV recommendations.
the
4.3. Biological criteria 4.3.1. The haemodynamics, kidney perfusion, and hormonal balance are different when the subject is standing or supine (Part 3 of the EPTRV recommendations). Whenever indicated, separate reference sample groups should be selected from ambulant and hospitalized population (e.g. S-albumin, substance concentration). 4.3.2. For some quantities, it may be necessary to consider chronobiological factors as criteria for partitioning (e.g. S-cortisol, substance concentration). 4.4. Reference state The notion of reference state can be used to facilitate population comparisons and to study transferability of reference value data. For such comparisons, the influence of biological variations should be minimal. Most constituents are subject to the least biological variations between the ages of 20 and 30 yr after other variation factors have been excluded. To qualify for the Reference State, individuals shall be 20-30 years of age, of ideal body mass, fasted for 10 h, taking no medication, consuming less than 45 g alcohol per day, smoking less than 12 cigarettes per day, and with no apparent illness [27].
SlO
5. Conclusion Production of reference values from any population or individuals requires appropriate selection and often subclassification. This can only be made by careful description of the characteristics of the reference individuals and by the application of clearly stated criteria. It is emphasized that depending on the uses of the reference values and the type of quantity, part or all, of the guidelines outlined in this document should be used.
6. References 1 PetitCIerc C, Wilding P. Provisional Recommendation on the theory of reference values. Part 2. Selection of individuals for the production of reference values. Clin Chim Acta 1984 139:205F-213F; J CIin Chem Clin Biochem 1984;22:203-208. Republished in Clin Chem Newslett (Milan) 1985;5:91-96. 2 Solberg HE. Approved Recommendation (1986) on the theory of reference values. Part 1. The concept of reference Values. Ann Biol CIin 1986;45:237-241, Clin Chim Acta 1987;165:111-118; J Clin Chem Clin Biochem 1987;25:in press. 3 Solberg HE. Approved recommendation (1987) on the theory of reference values. Part 5. Statistical treatment of collected reference values. Determination of reference limits. J Clin Chem Chim Biochem 1987;25:submitted for publication. 4 Dybkar R, Solberg HE. Approved recommendation (1987) on the theory of reference values. Part 6. Presentation of observed values related to reference values. 1987;submitted for publication. 5 Grasbeck R. Health as seen from the laboratory. In: Grasbeck R. Alstrom T, eds. Reference values in laboratory medicine. The current state of the art, Chichester: John Wiley and Sons, 1981;17-23. 6 Larsen 0. Separation health and illness - a conceptual framework. In: Grasbeck R, Alstrom T, eds. The current state of the art, Chichester: John Wiley and Sons, 1981;33-43. 7 Munan L, Kelly A, Petitclerc C. Population serum urate levels and their correlate; the Sherbrooke regional study. Am J Epidemiol 1976;103:369-382. 8 Munan L. Population based reference data. In: Rubin M, Benson ES, eds. Logic and economics of clinical laboratory use, New York: Elsevier, 1978;117-128. 9 Gardner M, Scott R. Frequency distribution and reference values of plasma alkaline phosphatase activity in the adult population of a Scottish new town. J Clin Pathol 1978;31:1202-1206. 10 Gardner M, Scott R. Age- and sex-related reference ranges for eight plasma constituents derived from randomly selected adults in a Scottish new town. J Clin Pathol 1980;33:380-385. 11 Berg B, Nilsson JE, Solberg HE, Tryding N. Practical experience in selection and preparation of reference individuals: empirical testing of the provisional Scandinavian recommendations. In: Grasbeck R, Alstrom T, eds. Reference vales in laboratory medicine. The current state of the art. Chichester: John Wiley and Sons, 1981;55-64. 12 Schiele F, Guilmin AM, Detienne H, Siest G. Gamma-glutamyltransferase activity in plasma: statistical distributions, individual variations and reference intervals. CIin Chem 1977;25:1023-1028. 13 Siest G. La biologic de I’homme sain. In: Metais P, et al, eds. Biochimie clinique. Tome 2: Biochimie physiopathologique. Lyon: SIMEP. 14 Steinmetz J. Panek E, Siest G, Gueguen R. Factors affecting the concentration of triacylglycerols (triglycerides) in plasma: reference values for adults. Clin Chem 1979;25:924-932. 15 Reed AH, Cannon DC, Winkelman JW, Bhasin YP, Henry RJ, Pileggi VJ. Estimation of normal ranges from a controlled sample survey. 1. Sex and age-related influence on the SMA 12/60 screemng group of tests. Clin Chem 1972;18:57-66. 16 Wilding P, Rollason JG, Robinson D. Patterns of change for various biochemical constituents detected in well population screening. Clin Chim Acta 1972;41:375-387.
17 Martin HF, Hologgitas JV, DriscoU J, Fanger H, Gudzinowicz BJ, Bamett R, Weisbrot I. Reference values based on populations accessible to hospitals. Grasbeck R, Alstrom T, eds. Reference values in Laboratory medicine. The current state of the art. Chichester: Wiley, 1981;233-262. 18 Winkel P. The use of the subject as his own Referent. In: Grasbeck R, Alstrom T, eds. Reference values in laboratory medicine. The current state of the art. Chichester: Wiley, 1981:65-78. 19 Statland BE. Winkel P. Selected preanalytical sources of variation. Grasbeck R, Alstrom T, eds. Reference values in laboratory medicine. The current state of the art. Chichester: Wiley, 1981;127-137. 20 Bretaudiere JP, Buret J, Favre R, Gueguen R, PetitClerc C, Sachs C, Siest G, Vemet M, Zender R. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology: biological variations in laboratory tests. Ann Biol Clin 1979;37:229-239. 21 Drodowsky M, Sachs Ch. Factors to be taken into considerations for blood sampling in view of establishing reference values. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology (Vemet-Nyssen M, Blin G, Buret J, Drodowsky M, Gueguen R, Guize L, Panek E, PetitClerc C, Sachs C, Zender R). Ann Biol Clin 1980;38:251-265. 22 Alstrom T, Grasbeck R, Hjelm M, Skandsen S. Recommendations concerning the collection of reference values in clinical chemistry and activity report by the Committee on Reference Values of the Scandinavian Society for Clinical Chemistry and Clinical Physiology. Stand J Clin Lab Invest 1975;35:suppl 144. 23 Harris EK. Some theory of reference values. I. Stratified (categorized) normal ranges and a method for following and individual’s clinical laboratory values. Clin Chem 1975;21:1457-1464. 24 Munan L, Kelly A, PetitClerc C, Billon B. Association with body weight of selected chemical constituents in blood: epidemiologic data. Clin Chem 1978;24:772-777. 25 Siest G, Henny J. Production of reference values in healthy subjects. Recommendations by the Scientific Committee on Reference Values of the French Society for Clinical Biology (Sachs Ch, Aellig A, Albert A, Blin G, Buret J, Daubrosse E, Drodowsky M. Gouget B, Gueguen R, Guize L, Henny J, PetitClerc C, Queralto J, Ramon F, Siest G, Vemet M). Ann Biol Clin 1981;39:235-244. 26 Wilding P, Bailey A. The normal range. In: Williams DL, Nunn RF. Marks V, eds. Scientific Foundations of Clinical Biochemistry, Vol. 1. London: Heinemann Medical Books Ltd. 1979;451-459. 27 Siest G, Schiele F, Henny J, Young DS. Interpretation of clinical laboratory tests, Davis: Biomedical Publications, 1985;18-22.