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SERUM ENZYME ASSAYS WITH COMMERCIAL KITS
1038 SERUM ENZYME ASSAYS WITH COMMERCIAL KITS
SiR,-Owing to increasing diversity of serum enzyme assays in diagnosis, many laboratories depend on reconstituted commercial kits. Those measuring reaction-rates optically at 340 nm. may contain errors. Occasional omissions, like cofactors, are immediately evident; but unsuspected contamination, apparently by the very enzyme to be measured, is increasingly common but far less obvious, and may be due
to
its presence in accessory indicator
enzymes prepared from a parent tissue. Repeated instances
which could have clinical consequences led us to examine kits from a well-known manufacturer (Boehringer Mannheim GmbH), particularly those for creatine phosphokinase (c.P.K.), as well as for aldolase (ALD), lactate dehydrogenase (L.D.H.), pyruvate kinase (P.K.), and aspartate (G.O.T.) and alanine (G.P.T.) aminotransferases.
used instead of serum in freshly prepared multiple-assay kits (noted with catalogue number), the following blank values in mu per ml. at 250C were obtained as perfectly linear recordings at 340 nm. for 20 minutes on a Unicam SP 800 B spectroWhen normal saline
was
photometer :
X-linked recessive (Duchenne) muscular dystrophy 2; so that an unlucky C.P.K. vial that adds or subtracts (if used as blank) some 6 units might decide a needless abstinence from reproduction, or worse, the appearance of dystrophic offspring in families falsely reassured. These findings are reported with just such instances in mind, with the suggestion that, since C.P.K. survives overnight posting in a filled tube of clotted bloody these assays be sent to a suitable laboratory. This investigation was supported by the Muscular Dystrophy Group of Great Britain. Research Laboratory, Knightswood Hospital, Glasgow G13 2XG.
IAN SMITH W. H. S. THOMSON.
BLOOD-LEAD IN CHILDREN
SIR,-IfDr Landrigan and others (March 29, p. 708), are right, children show ill-effects at blood-lead levels (48 {J.g. per 100 ml.) classified as acceptable " (40-80 (Lg. Pb per 100 ml.)3 for industrial settings, but are not regarded as applicable to the general population nor, in particular, to children. Acceptable levels for non-occupational exposure are customarily lower than for occupational exposure. "
Some data for adults may be relevant to hazards in children. I have determined the abundance of lead (together with most of the elements of the periodic table) 4 in blood for the U.K., excluding N. Ireland, for the year 1971. A total The blank value for one c..x, kit, read in quadruplicate 4 weeks storage at 4°C, remained constant at 8-16. Subtraction of each kit blank before calculation, moreover, gave very close correlation (0-9976) with serum C.P.K. values simultaneously assayed using the J. T. Baker multiple-assay vial (Diamed Diagnostics, Liverpool) with no blank value, though that of their single-assay vials ranged from nil to 2-04. Thus, where adult and pxdiatric specimens go to the same laboratory performing frequent C.P.K. assays, a simple correction gives dependable results.
weekly during
CONCENTRATIONS OF LEAD IN BLOOD FOR THE UNITED KINGDOM FOR NON-OCCUPATIONALLY EXPOSED ADULTS
In the separate pxdiatric or genetic laboratories of large teaching hospitals, where c.P.K. is rarely requested, boxes of 20 lyophilised single-assay vials are used instead of multiple-assay kits. The contents in the vial are dissolved in prepared buffer, serum is added, and the whole is trans-
ferred
to a silica cuvette for measurement of enzyme rate of change of absorbance at 340 nm. Since
activity by
saline blank in such a vial distorts conditions for subsequent serum assay, it might be assumed no blank is needed, or that a single blank is the same in all vials from the same box and with the same batch number. This is not so. The following blank values were obtained linearly (as above) from all such single-assay vials in complete boxes of 20, noted with the Boehringer catalogue number and reported as the recorded range with mean ±2 s.D.: a
All these values are multiplied if diluted serum is used. In G.o.T. and G.P.T. contamination is small; but the inexplicable L.D.H. vial, containing only pyruvate, N.A.D.H. and phosphate buffer, might influence diagnosis in theanginal infarct seen too late for other enzymes. In C.P.K., however, even a few units above or below a female upper limit of 37-7 mu per ml. at 25°Cmay confirm or deny a high risk of the carrier state for females in pedigrees of 1.
Sweetin, J. C., Thomson, W. H. S. Clinica chim. Acta, 1973, 48, 49.
of 151
samples of blood were collected randomly in relation geography except that samples from large cities and industrial settings were excluded. A second group of samples were collected from 2500 individuals for the same geographical coverage; aliquots (100 mg. dry blood) were removed from each sample and combined to form a master to
mix for U.K. blood. The results of the lead aspect of this study are presented in the table. For the 151 samples, a mean blood-lead level of 24 g. per 100 ml. was obtained which may be compared with a value of 26 (Lg. per 100 ml. for the master mix. The similarity between the two mean values may indicate that the data reflect similar lead distributions in both suites of samples. If blood-lead levels for children are enhanced relative to adults, then a significant number of children and adults in the U.K. population could be potentially exposed to levels of lead in blood which according to the findings of Landrigan et al. could be detrimental to health. While symptoms of lead intoxication in children may be recognised through various i.Q. and motor-response tests the insidiousness of lead intoxication and the vague nature of possible symptoms would not be 2. Thomson, W. H. S. ibid. 1969, 26, 207. 3. See Br. Med. J. 1968, ii, 501. 4. Hamilton, E. I. Science of the Total Environment,
1972/73, 1, 341.
SiR,-Owing to increasing diversity of serum enzyme assays in diagnosis, many laboratories depend on reconstituted commercial kits. Those measuring reaction-rates optically at 340 nm. may contain errors. Occasional omissions, like cofactors, are immediately evident; but unsuspected contamination, apparently by the very enzyme to be measured, is increasingly common but far less obvious, and may be due
to
its presence in accessory indicator
enzymes prepared from a parent tissue. Repeated instances
which could have clinical consequences led us to examine kits from a well-known manufacturer (Boehringer Mannheim GmbH), particularly those for creatine phosphokinase (c.P.K.), as well as for aldolase (ALD), lactate dehydrogenase (L.D.H.), pyruvate kinase (P.K.), and aspartate (G.O.T.) and alanine (G.P.T.) aminotransferases.
used instead of serum in freshly prepared multiple-assay kits (noted with catalogue number), the following blank values in mu per ml. at 250C were obtained as perfectly linear recordings at 340 nm. for 20 minutes on a Unicam SP 800 B spectroWhen normal saline
was
photometer :
X-linked recessive (Duchenne) muscular dystrophy 2; so that an unlucky C.P.K. vial that adds or subtracts (if used as blank) some 6 units might decide a needless abstinence from reproduction, or worse, the appearance of dystrophic offspring in families falsely reassured. These findings are reported with just such instances in mind, with the suggestion that, since C.P.K. survives overnight posting in a filled tube of clotted bloody these assays be sent to a suitable laboratory. This investigation was supported by the Muscular Dystrophy Group of Great Britain. Research Laboratory, Knightswood Hospital, Glasgow G13 2XG.
IAN SMITH W. H. S. THOMSON.
BLOOD-LEAD IN CHILDREN
SIR,-IfDr Landrigan and others (March 29, p. 708), are right, children show ill-effects at blood-lead levels (48 {J.g. per 100 ml.) classified as acceptable " (40-80 (Lg. Pb per 100 ml.)3 for industrial settings, but are not regarded as applicable to the general population nor, in particular, to children. Acceptable levels for non-occupational exposure are customarily lower than for occupational exposure. "
Some data for adults may be relevant to hazards in children. I have determined the abundance of lead (together with most of the elements of the periodic table) 4 in blood for the U.K., excluding N. Ireland, for the year 1971. A total The blank value for one c..x, kit, read in quadruplicate 4 weeks storage at 4°C, remained constant at 8-16. Subtraction of each kit blank before calculation, moreover, gave very close correlation (0-9976) with serum C.P.K. values simultaneously assayed using the J. T. Baker multiple-assay vial (Diamed Diagnostics, Liverpool) with no blank value, though that of their single-assay vials ranged from nil to 2-04. Thus, where adult and pxdiatric specimens go to the same laboratory performing frequent C.P.K. assays, a simple correction gives dependable results.
weekly during
CONCENTRATIONS OF LEAD IN BLOOD FOR THE UNITED KINGDOM FOR NON-OCCUPATIONALLY EXPOSED ADULTS
In the separate pxdiatric or genetic laboratories of large teaching hospitals, where c.P.K. is rarely requested, boxes of 20 lyophilised single-assay vials are used instead of multiple-assay kits. The contents in the vial are dissolved in prepared buffer, serum is added, and the whole is trans-
ferred
to a silica cuvette for measurement of enzyme rate of change of absorbance at 340 nm. Since
activity by
saline blank in such a vial distorts conditions for subsequent serum assay, it might be assumed no blank is needed, or that a single blank is the same in all vials from the same box and with the same batch number. This is not so. The following blank values were obtained linearly (as above) from all such single-assay vials in complete boxes of 20, noted with the Boehringer catalogue number and reported as the recorded range with mean ±2 s.D.: a
All these values are multiplied if diluted serum is used. In G.o.T. and G.P.T. contamination is small; but the inexplicable L.D.H. vial, containing only pyruvate, N.A.D.H. and phosphate buffer, might influence diagnosis in theanginal infarct seen too late for other enzymes. In C.P.K., however, even a few units above or below a female upper limit of 37-7 mu per ml. at 25°Cmay confirm or deny a high risk of the carrier state for females in pedigrees of 1.
Sweetin, J. C., Thomson, W. H. S. Clinica chim. Acta, 1973, 48, 49.
of 151
samples of blood were collected randomly in relation geography except that samples from large cities and industrial settings were excluded. A second group of samples were collected from 2500 individuals for the same geographical coverage; aliquots (100 mg. dry blood) were removed from each sample and combined to form a master to
mix for U.K. blood. The results of the lead aspect of this study are presented in the table. For the 151 samples, a mean blood-lead level of 24 g. per 100 ml. was obtained which may be compared with a value of 26 (Lg. per 100 ml. for the master mix. The similarity between the two mean values may indicate that the data reflect similar lead distributions in both suites of samples. If blood-lead levels for children are enhanced relative to adults, then a significant number of children and adults in the U.K. population could be potentially exposed to levels of lead in blood which according to the findings of Landrigan et al. could be detrimental to health. While symptoms of lead intoxication in children may be recognised through various i.Q. and motor-response tests the insidiousness of lead intoxication and the vague nature of possible symptoms would not be 2. Thomson, W. H. S. ibid. 1969, 26, 207. 3. See Br. Med. J. 1968, ii, 501. 4. Hamilton, E. I. Science of the Total Environment,
1972/73, 1, 341.