Original Article

Macroenzyme detection by polyethylene glycol precipitation

D Fraser Davidson and Dawn JM Watson

Abstract
Address Background The presence of macroenzymes can cause significant diagnostic
Biochemistry Department confusion and their detection can involve relatively cumbersome analytical
Crosshouse Hospital
procedures.
Kilmarnock KA2 0BE, UK
Methods Using a simplified polyethylene glycol precipitation technique and
Correspondence isoenzyme electrophoresis, this report describes the construction of reference ranges
Dr D Fraser Davidson of precipitable activity for each of seven commonly measured enzymes in plasma.
E-mail: fraser.davidson@aaaht.scot.nhs.uk
Results The proposed reference ranges are reported. Since introducing the
protocol, 12 cases of macroenzymaemia have been encountered. Three typical case
histories are described in some detail.
Conclusions The polyethylene glycol precipitation method has thus far proved to
be a simple and effective additional test for the detection of macroenzymes when the
plasma enzyme activity is elevated.
Ann Clin Biochem 2003; 40: 514–520

Introduction enzymes often involve complicated or cumbersome

Macroenzymes are enzymes in plasma that have
formed high-molecular-mass complexes, either by
self-polymerization or by association with other
plasma components.1 They have been described for
most routinely measured enzymes in the clinical
laboratory, and are usually complexes of normal
enzymes with an Ig, although other forms have been
reported.1,2 Immunoglobulin-associated forms are
referred to as type 1 macroenzymes; any others are
termed type 2.2 Although plasma activity may be
una¡ected or, perhaps, reduced by such e¡ects, in
some cases, Ig binding to circulating enzymes may
lead to an increased activity, probably by mechanisms
involving reduced inactivation, clearance or excre-
tion.1 There is little evidence that type 1 macro-
enzymes indicate the presence of disease. However,
they are indistinguishable from normal enzymes
during routine assay. The importance of their detec-
tion lies in their ability to cause diagnostic confusion
when plasma enzyme activities are elevated, leading
to unnecessary further investigative procedures.3,4
Although prevalence studies are rare, the presence
of a macroenzyme is not necessarily uncommon.
For example, macroamylase may be detected in
approximately one in 20 individuals with hyper-
amylasaemia.5 However, techniques to detect macro-
procedures requiring highly specialized chromato-
graphy, electrophoresis or ultracentrifugation equip-
ment not available to the average hospital laboratory.3
A relatively simple technique that has been success-
fully applied to the detection of macroamylase is that
of polyethylene glycol (PEG) precipitation.5,6 However,
there are virtually no reference range data for any
enzyme other than amylase. Hence, the purpose of the
present study was to examine the precipitation
technique with a view to constructing reference
ranges of PEG-precipitable activity (%PPA) for each of
seven routinely measured plasma enzymes, where
possible, to assess the homogeneity of the various
reference groups by isoenzyme electrophoresis, and to
report our experiences with the technique in patient
investigation.
Patients and methods
The Monovette blood collection system (Sarstedt Ltd,
Leicester, UK) was used throughout the study. Blood
specimens were received from inpatients, outpatients
and general practices throughout the local area, and
routine analyses were performed on lithium heparin
plasma.
Analyses of alanine transaminase (ALT), aspartate
transaminase (AST), alkaline phosphatase (ALP),

514 © 2003 The Association of Clinical Biochemists


amylase (AMY), creatine kinase (CK), g-glutamyl-
transpeptidase (GGT), lactate dehydrogenase (LDH),
albumin (ALB), total protein (TP), triglycerides (TG),
cholesterol (CHOL) and creatinine were carried out
using methods and equipment described previously.7
The upper reference limits for the plasma enzyme
activities were: ALT 50 U/L, AST 40 U/L, ALP 140 U/
L, AMY 100 U/L, CK 195 U/L, GGT 50 U/L and LDH
540 U/L. Measurement of Igs G, A and M was
performed on a Beckman Array protein system
[Beckman Instruments (UK) Ltd, High Wycombe, UK].
Laboratory results were scrutinized in order to
identify individuals, generally with elevated plasma
enzyme activities, from whom surplus unhaemolysed
plasma was retrieved. In each case, using request form
information, the data recorded for each patient
included age, sex and any clinical details provided.
Plasma from individuals known to be of high risk (e.g.
human immunode¢ciency virus, hepatitis B or C)
were not selected. Reference ranges were constructed
by selecting plasma from approximately 40 patients
for each enzyme. In order to avoid the possibility of the
inadvertent inclusion of samples containing macro-
enzyme, the homogeneity of each group was assessed
by examination of each distribution for the presence of
outliers, and, in the case of AST, ALT, ALP, CK and
LDH, by subjecting to isoenzyme electrophoresis the
four samples showing lowest and the four showing
highest %PPA. Any skewed distribution was normal-
ized by logarithmic or square root transformation as
required; after appropriate exclusion and investiga-
tion of any outliers, reference ranges were calculated
as plus and minus two standard deviations (SD) from
the mean.
To investigate the PEG precipitation process, pooled
human plasma was prepared from blood obtained from
patients with elevated Igs. Ten aliquots of this pooled
plasma were each mixed with an equal volume of PEG
6000 (240 g/L in 9 g/L saline) and, after vortex
mixing, were held variously for 1-10 min at room
temperature (218C) prior to centrifugation. Each
supernantant was removed, analysed for ALB, TP, TG,
CHOL, IgG, IgA and IgM and the results were
compared with those obtained from a 1:2 dilution of
the pooled plasma in 9 g/L saline.
In the construction of reference ranges for %PPA,
patients’ plasma (200 mL) was added to either 200 mL
of PEG 6000 solution (240 g/L in 9 g/L saline) or
200 mL saline (9 g/L) and held at room temperature for
10 min prior to centrifugation. For each patient, the
enzyme activities were measured on both the super-
natant and the saline dilution. %PPA was calculated
for each enzyme as:
%PPA ˆ 100 £ ‰…activitysaline ¡ activityPEG †=
…activitysaline †Š
Macroenzyme detection by polyethylene glycol precipitation 515
Inter-assay precision of %PPA measurement was
assessed by including a bovine serum-based commer-
cial quality control sample with each batch of analyses
(Humatrol N, cat. no. 13511, Human, Taunusstein,
Germany).
Isoenzyme electrophoresis for ALP, CK and LDH was
performed initially using the Corning-ACI system
(Corning Medical, Halstead, UK), and more recently by
the Sebia Hydrasys semi-automated electrophoresis
system (Sebia, Issy-les-Moulineaux, France). AST and
ALT isoenzymes were separated on Corning Universal
agarose gels (now available from Helena Biosciences
Ltd, Sunderland, UK; cat. no. HL-3-1161SA) and
visualized as described by Dawson et al.8 The AST and
ALT isoenzymes were viewed under ultraviolet light
(366 nm) as dark bands on a £uorescent background.
Results
Polyethylene glycol precipitation
The results of the study of PEG precipitation on pooled
plasma are shown in Table 1. PEG precipitation
removed all of the measurable IgG, IgM and approxi-
mately 80% of the IgA, CHOL, TG and GLOB, whereas
ALB was little a¡ected. The precipitation process was
complete within 1min at room temperature, and
further incubation produced no increase in analyte
removal.
Reference ranges
The reference data from which the proposed reference
ranges were calculated are summariz ed in Table 2.
There were only two outliers, which were excluded
from further calculation. One was from the amylase
group (%PPA ˆ 82) of 4.4 SD from the mean. The
second outlier arose in the LDH group (%PPA ˆ 100),
which was 4.1 SD above the mean. Isoenzyme
electrophoresis of this latter sample revealed a broad
band of activity in the LD4- LD3 region typical of
macroLDH,9,10 as shown schematically, together with
the other three highest and four lowest %PPA values,
in Fig. 1. Isoenzyme electrophoresis of AST and ALT
yielded only the cytoplasmic isoenzymes; ALP
Table 1. Effect of polyethylene glycol (PEG) precipitation on
proteins and lipids
Before PEG After PEG
Total protein (g/L) 86 44
Albumin (g/L) 38 34
Globulin (g/L) 48 10
Cholesterol (mmol/L) 5.4 0.8
Triglyceride (mmol/L) 1.94 0.4
IgG (g/L) 15.86 50.33
IgA (g/L) 11.6 1.9
IgM (g/L) 2.46 50.04
Ann Clin Biochem 2003; 40: 514–520
516 Davidson and Watson

Table 2. Summary of reference data from which proposed reference ranges for enzyme %PPA were calculated
Median (range) Proposed
Number of Mean (range) enzyme activity Mean (range) reference
Enzyme patients age (years) Clinical data (U/L) %PPA range (%)
AMY 39 (23F, 17M) 57 (25-85) Pancreatitis/abdominal pain (34); 239 (87-1444) 40.9 (18.0-56.0) 22-60
others (6)
ALP 43 (20F, 23M) 63 (30-88) Liver disease/alcoholism (14); 278 (122-974) 11.0* (0-40.0) **0-36
cancer (10); pancreatic disease (5);
chronic renal failure/
bone disease (5); others (9)
AST 40 (15F, 25M) 57 (9-86) Myocardial infarction/chest pain (14); 101 (48-364) 36.8 (17.0-55.0) 18-53
liver disease/alcoholism (16);
cancer (6); others (4)
ALT 40 (13F, 27M) 56 (9-83) Liver disease/alcoholism (12); 99 (52-486) 57.3 (39.0-78.0) 38-76
myocardial infarction/chest pain (9);
cancer (9); pancreatitis (3); congestive
cardiac failure (2); others (5)
CK 40 (14F, 26M) 64 (26-93) Myocardial infarction/chest pain (32); 233 (82-896) 21.0* (13.0-39.0) {12-37
post-operative (2); fracture (1);
hypothermia (1); others (4)
LDH 40 (19F, 21M) 67 (40-93) Myocardial infarction/chest pain (26); 894 (446-2230) 40.7 (9.0-74.0) 12-70
congestive cardiac failure (5);
post-operative (3); others (7)
GGT 40 (15F, 25M) 52 (27-83) Liver disease/alcoholism (20); 191 (48-1304) 25.8 (0-49.0) 0-51
epilepsy (2); others (18)
*Indicates median %PPA; **after square root transform; {after logarithmic transform. PPA ˆ polyethylene glycol precipitable activity; F ˆ female;
M ˆ male; AMY ˆ amylase, ALP ˆ alkaline phosphatase; AST ˆaspartate transaminase; ALT ˆalanine transaminase; CK ˆ creatine kinase;
LDH ˆ lactate dehydrogenase; GGT ˆ g-glutamyltranspeptidase.

showed only liver and/or bone bands of activity; and Precision

CK exhibited only the presence of MM or MM plus
MB. The distributions of %PPA for ALP and CK were
each positively skewed. These were normaliz ed by
square root and logarithmic transformations,
respectively.
The inter-assay coe¤cients of variation for %PPA
measurements with the Humatrol N quality control
specimen for each of the seven enzymes, over 12
separate batches (six in the case of GGT) are given in
Table 3.

Figure 1. Schematic representation of the lactate

dehydrogenase (LDH) isoenzyme electrophoretogram,
including a case of macroLDH (track 8). PPA ˆ
polyethylene glycol precipitable activity; MI ˆ myocardial
infarction; CCF ˆ congestive cardiac failure.

Ann Clin Biochem 2003; 40: 514–520

 Macroenzyme detection by polyethylene glycol precipitation 517

Table 3. Inter-assay precision of the polyethylene glycol precipitation, a request was always made to obtain a
precipitation technique using Humatrol N quality control serum suitable urine specimen for measurement of the
amylase/creatinine clearance ratio (ACCR).
Enzyme Mean activity (U/L) Mean %PPA CV (%) Since introducing the PEG precipitation test we
AMY 250 42.3 11.2 have investigated 57 patients for the presence of
ALP 426 47.0 5.3 macroenzymes. Including the one case (case 1) of
AST 97 32.5 5.9 macroamylasaemia (macroAMY) and one of macro-
ALT 115 78.4 3.4 LDH (case 9) encountered during the production of
CK 453 32.3 10.8 reference ranges as described, we have, at time of
LDH 1118 41.5 7.3 writing, identi¢ed 12 cases of macroenzymaemia
GGT 154 24.5 17.3 (positives/total tested): ¢ve with macroAMY (5/21);
PPA ˆ polyethylene glycol precipitable activity; CV ˆ coefŽ cient of three with macroAST (3/3); one macroLDH (1/3);
variation; AMY ˆ amylase, ALP ˆ alkaline phosphatase; AST ˆ one typical of type 1 macroCK1,3 plus one typical of
aspartate transaminase; ALT ˆalanine transaminase; CK ˆ creatine type 2 macroCK11,12 (2/18); and one composed mainly
kinase; LDH ˆ lactate dehydrogenase; GGT ˆ g-glutamyltranspepti- of an electrophoretically slow-moving band consistent
dase. with macroALP13,14 (1/5). No patients have, as yet,
been identi¢ed as having macroGGT (0/6) or
Patients macroALT (0/1). The positive cases, together with
their associated details, including results of ACCR
The screening protocol included both the PEG where available, are summarize d in Table 4. As can be
precipitation test and isoenzyme electrophoresis in seen from this table, each of these patients showed
each case of ALP, AST, ALT, CK and LDH examined. In %PPA values markedly above the proposed upper
the detection of macroamylase, in addition to PEG reference limits.

Table 4. Summary of results from patients with macroenzymes

Enzyme
activity ACCR
Case no. Enzyme Age, sex (U/L) %PPA Isoenzyme electrophoresis (%) Clinical details
1 AMY 66, F 439 82 NA Pancreatitis
2 AMY 27, M 294 84 0.51 Epigastric pain, nausea and
vomiting
3 AMY 39, M 935 79.1 NA Acute pancreatitis with persisting
hyperamylasaemia
4 AMY 22, F 310 83 1.40 2-week history of intermittent
epigastric pain
270 79
282 81
5 AMY 31, M 1426 96.6 0.03 Acute abdominal pain
1422 95.5
6 AST 52, F 115 71 Mainly origin band + small 4-week history of upper
anodal cytoplasmic band abdominal pain
7 AST 63, M 89 95 Origin band only Post-operative collapse
240 97
233 100
67, M 211 100
8 AST 41, F 602 97 Origin band only Immune neutropenia
9 LDH 81, F 1446 100 Three bands: LD1, LD2 + broad Congestive cardiac failure
band in LD4–LD3 position
10 CK 37, M 296 69.1 Two bands: MM + intermediate Palpitations and dyspepsia
between MM–MB positions
11 CK 59, M 567 66.1 MM + cathodic band + small MB Hepatic metastases
and BB
12 ALP 67, M 242 82.4 Mainly slow-moving anodic band Persistently elevated ALP
(mobility 40% of usual liver)
PPA ˆ polyethylene glycol precipitable activity; ACCR ˆ amylase/creatinine clearance ratio; M ˆ male; F ˆ female; AMY ˆ amylase;
AST ˆaspartate transaminase; LDH ˆ lactate dehydrogenase; CK ˆ creatine kinase; ALP ˆ alkaline phosphatase; NA ˆ not available.

Ann Clin Biochem 2003; 40: 514–520

518 Davidson and Watson
Of the 16 patients considered negative for
macroAMY by the PEG precipitation test (median
%PPA ˆ 35, range 24 - 47), ACCR results were avail-
able for 14 specimens from 13 patients and these were
also normal 5 (median ACCR ˆ 3.8%, range 2.5-
13.5%). Isoenzyme electrophoresis was not available
as a con¢rmatory test for macroGGT detection.
However, electrophoresis was performed on all
patients investigated for macroALP, macroALT and
macroLDH, and each negative showed %PPA values
within the proposed reference ranges. All three
patients investigated for the presence of macroAST
showed both elevated %PPA and a macroenzyme
band on electrophoresis, as described in Table 4.
Of the 16 patients considered negative for macroCK,
three showed %PPA values above the proposed upper
reference limit. These were 41.0, 41.4 and 44.0,
respectively. An isoenzyme electrophoresis result was
unavailable for the ¢rst of these three patients, whereas
the other two with higher values for %PPA showed no
evidence of the presence of macroCK. The next highest
value for CK %PPA was 66.1 (see Table 4, case 11). This
sample was from a 59-year-old gentleman with
histologically con¢rmed hepatic metastases from an
unknown primary. He manifested a distinctive
cathodic band on isoenzyme electrophoresis consis-
tent with the presence of macroCK type 2.1 Of those
shown to be positive for the presence of macroenzymes,
some of the case histories are described below.
Figure 2. Case 8: changes in plasma or serum aspartate transaminase (AST) activity with time of storage at 48C compared with that
of a control patient (see text).
Ann Clin Biochem 2003; 40: 514–520
Case 5
This 31-year-old gentleman was admitted as an emer-
gency with acute abdominal pain, pyrexia and
vomiting. He had a 5-year history of Crohn’s disease.
On admission, his urea and electrolytes, liver function
tests, glucose and calcium were normal. His plasma
AMY was 1779 U/L (normal 5100 U/L), which was
consistent with a presumed diagnosis of acute
pancreatitis. However, a subsequent abdominal
ultrasound examination was normal and showed no
evidence of acute pancreatitis, despite a further
elevated plasma AMY activity of 1426 U/L. Nine days
after admission his plasma AMY remained at1422 U/L
despite an abdominal computed tomography (CT)
scan which showed no evidence of pseudocyst forma-
tion or pancreatic in£ammation. A urine test for AMY
activity showed a result of only 33 U/L, an ACCR
of 0.03% (normal 41.6%),5 and measurement of
%PPA on two separate occasions yielded values of
96.6 and 95.5, all of which indicated the presence of
macroamylase.
Case 7
This 63-year-old gentleman was admitted to hospital
for routine removal of nasal polyps. His admission
urea and electrolytes and electrocardiogram (ECG)
were normal. Several hours after this operation he was
found collapsed. His plasma AST was 240 U/L (normal
540 U/L) with normal CK and ALT; a provisional

diagnosis of acute myocardial infarction was made. On
the following day his AST was 233 U/L, CK and ALT
remained within normal limits, his ECG was unre -
markable and he quickly recovered. Retrospective
measurement of AST on the blood specimen taken
more than 24 h previously, at time of admission,
showed a result of 89 U/L. PEG precipitation tests on
this specimen and the two subsequent ones yielded
%PPA values of 95, 97 and 100, respectively, whilst
AST electrophoresis con¢rmed the presence of
macroAST as the only visible band of activity. Inter-
estingly, a further test, which was performed 4 years
later, showed an AST of 211 U/L; a PEG precipitation of
100% and AST electrophoresis again con¢rmed the
presence of persisting macroAST.
Case 8
This 41-year-old lady had been noted to have a mild
degree of neutropenia during routine monitoring of
her latest pregnancy and was referred to a haematolo-
gist who was able to diagnose immune neutropenia by
demonstrating the presence of granulocyte -speci¢c
antibodies (IgG and IgM) in her serum. However,
during her investigations she was noted, over a 3-
month period, to have signi¢cantly elevated plasma
AST activities, of 414, 513 and 411 U/L, respectively,
whilst other liver function tests were within normal
limits. In response to these ¢ndings, and after delivery
of a healthy male infant, an abdominal ultrasound
examination was performed which was unremark-
able, indicating that the liver, kidneys, spleen and
pancreas were normal. Further blood specimens taken
1month later showed an ASTof 602 U/L and a %PPA of
97; AST isoenzyme electrophoresis con¢rmed the
presence of macroAST. Moreover, re-measurement of
AST on both plasma and serum specimens revealed a
marked decrease in activity with time of specimen
storage. More than 90% of the original AST activity
was lost during 6 days of refrigerated storage. By
comparison, a similarly treated control sample from a
54-year-old gentleman with liver disease (plasma AST
174 U/L, %PPA 44, AST isoenzyme electrophoresis
exhibiting mainly the anodal cytoplasmic isoenzyme
with a small cathodal mitochondrial band and no
macroAST) showed virtually no loss of activity over
the same period (see Fig. 2).
Discussion
Polyethylene glycol precipitation
The PEG precipitation process, as used in the present
study, is relatively speci¢c for the globulin fraction of
plasma, removing virtually all measurable Ig. The
e¡ect is rapid and complete within 1min of incubation
at room temperature.
Macroenzyme detection by polyethylene glycol precipitation 519
The e¡ects of PEG on plasma enzyme activity are
probably multifactorial. Firstly, since Igs are precipi-
tated, any enzyme that is Ig-bound will also be
removed, thereby allowing detection of type 1 macro-
enzymes. Secondly, there is signi¢cant removal of lipid
from plasma by PEG and the reagent has been applied
successfully to the selective precipitation of LDLs.15
Some additional forms of type 2 macroenzymes such
as GGT or ALP may be composed of lipid aggregates,1
and hence treatment with PEG may be expected to
remove these enzymes along with their attached lipid.
Thirdly, treatment of sample with PEG is capable of
precipitating macroCK type 2,11 as observed in case 11
of the present study (see Table 4). MacroCK type 2
is thought to represent macromolecular aggregates
of mitochondrial CK found in the serum of severely
ill patients, mainly those with malignant disease.16
However, Igs and lipids are not substantial parts of
macroCK type 2.11 Hence, the mechanism and poten-
tial variability of reduction in enzyme activity
following treatment with PEG in cases of macroCK
type 2 remains to be determined. A fourth possible
e¡ect of PEG is inhibition of enzyme activity. Even in
the absence of macroenzyme, as observed by isoen-
zyme electrophoresis, there is a wide variation in the
reference ranges for %PPA between each enzyme. One
possible reason for this may be variable enzyme inhi-
bition. Therefore, application of the test and reference
ranges to analytical systems and platforms other than
those employed in the present study would require
validation. However, notwithstanding the above
comments, the PEG precipitation test proved to be a
simple technique to perform.
It is not envisaged at the present time that the PEG
precipitation test be adopted as a substitute for iso-
enzyme electrophoresis in the investigation of patients
with possible macroenzymes. However, as experience
accumulates and, if the technique proves to be free of
false negatives, it should readily allow the search for
patients with macroenzymes to be conducted more
easily. It is interesting that two cases of previously
unsuspected macroenzymaemia were discovered
incidentally during the construction of reference
ranges for %PPA.
Reference ranges
The proposed reference range for AMY %PPA of 22-
60, as determined by the present study, is very similar
to, and consistent with, the threshold value for further
investigation of 57 as proposed in the recent extensive
study by Lawson.5 In the absence of amylase iso-
enzyme electrophoresis, the additional or con¢rma-
tory test available to us in the present study was the
ACCR. In the ¢ve cases positive for macroamylase
shown in Table 4, ACCR results were available for
three patients and these were also abnormal.
Ann Clin Biochem 2003; 40: 514–520
520 Davidson and Watson
It has been recently reported that plasma from four
control patients with elevated ALP activities due to
liver or bone disease, when mixed with an equal
volume of 24% (w/v) PEG 8000, precipitated 5%, 7%,
8% and 17% of the activity;17 this is consistent with
our proposed reference range for %PPA ALP of 0-36.
By comparison, in the same report, two samples from a
case of macroALP showed values of 73 and 82,
respectively;17 this is in agreement with our own
¢ndings (see Table 4, case 12).
The number of subjects used to construct each of the
reference ranges for %PPA may be considered to be
rather small. However, with the gradual accumulation
of data, more suitable cut-o¡ values for %PPA, below
which the existence of a macroenzyme has proven to
be unlikely, should emerge with experience. For
example, it is likely that the reference range for %PPA
for CK will require some alteration.
Patients
The description of case 5 above is typical of that of a
case of macroamylasaemia. However, detection of the
presence of macroamylase does not, by itself, necessa-
rily exclude a diagnosis of acute pancreatitis.5 This
appears to have been the situation with case 3 (see
Table 4). This same argument, that disease may be
present, though less likely, despite the existence of a
macroenzyme, applies to all individuals in whom one
has been detected.
Regarding case 7, that of macroAST, it is noteworthy
that the presence of macroenzyme was not a transient
phenomenon; hence, it is important to record this
information prominently in the patient’s notes in order
to help avoid diagnostic confusion, perhaps years in
the future. The most interesting feature of case 8 is the
rapid disappearance of AST activity in vitro following
sample collection and separation. Presumably, this is
due to the gradual precipitation of enzyme - Ig
complex during refrigerated storage. It will be inter-
esting to determine, as experience is accumulated, if
this is a common feature of samples containing
macroenzymes; if so, the observation may be incorpo-
rated into a future testing regime.
Acknowledgements
The authors wish to express their gratitude to the
patients referred to as numbers 5, 7 and 8 in the
present study for allowing their cases to be described
in some detail. In addition, they are grateful to Mr J
Williamson, Principal Biochemist, Mr M McClelland,
Laboratory Computer Systems Manager, and the sta¡
of the biochemistry, haematology, pathology and
medical records departments at Crosshouse Hospital
Ann Clin Biochem 2003; 40: 514–520
for their helpful assistance throughout this on-going
study.
References
1 Remaley AT, Wilding P. Macroenzymes: biochemical characterisa-
tion, clinical signiŽ cance, and laboratory detection. Clin Chem 1989;
35: 2261-70
2 Sturk A, Sanders GTB. Macroenzymes: composition, detection and
clinical relevance. J Clin Chem Clin Biochem 1990; 28: 65-81
3 Mif in TE, Bruns DE. University of Virginia case conference.
Macroamylase, macro creatine kinase, and other macroenzymes.
Clin Chem 1985; 31: 1743-8
4 Galasso PJ, Litin SC, O’Brien JF. The macroenzymes: a clinical
review. Mayo Clin Proc 1993; 68: 349-54
5 Lawson GJ. Prevalence of macroamylasaemia using polyethylene
glycol precipitation as a screening method. Ann Clin Biochem 2001;
38: 37-45
6 Levitt MD, Ellis C. A rapid and simple assay to determine if
macroamylase is the cause of hyperamylasemia. Gastroenterology
1982; 83: 378-82
7 Davidson DF. Effects of contamination of blood specimens with
liquid potassium-EDTA anticoagulant. Ann Clin Biochem 2002; 39:
273-80
8 Dawson CM, Connelly MS, Kennedy HJ, Howe GD, Tickner TR.
Investigation of a persistently raised serum AST. Ann Clin Biochem
1989; 26: 538-41
9 Biewenga J, Feltkamp TEW. Lactate dehydrogenase (LDH)-IgG3
immunoglobulin complexes in human serum. Clin Chim Acta 1975;
64: 101-16
10 Tozuka M, Katsuyama T. A case of immunoglobulin A-l conjugated
with lactate dehydrogenase-5 isoenzyme, causing an extremely
high enzyme activity in serum. Clin Chem 1996; 42: 1288-90
11 Stein W, Bohner J, Bahlinger M. Analytical patterns and
biochemical properties of macro creatine kinase type 2. Clin Chem
1985; 31: 1952-8
12 Lee KN, Csako G, Bernhardt P, Elin RJ. Relevance of macro
creatine kinase type 1 and type 2 isoenzymes to laboratory and
clinical data. Clin Chem 1994; 40: 1278-83
13 Nakagawa H, Umeki K, Yamanaka K, Kida N, Ohtaki S.
Macromolecular alkaline phosphatase and an immunoglobulin G
that inhibited alkaline phosphatase in a patient’s serum. Clin Chem
1983; 29: 375-8
14 Wenham PR, Chapman B, Smith AF. Two macromolecular
complexes between alkaline phosphatase and immunoglobulin A
in a patient’s serum. Clin Chem 1983; 29: 1845-9
15 Briggs CJ, Anderson D, Johnson P, Deegan T. Evaluation of the
polyethylene glycol precipitation method for the estimation of high-
density lipoprotein cholesterol. Ann Clin Biochem 1981; 18: 177-81
16 Stein W, Bohner J, Rann W, Maulbetsch R. Macro creatine kinase
type 2: results of a prospective study in hospitalised patients. Clin
Chem 1985; 31: 1959-64
17 Owen MC, Pike LS, George PM, Barclay ML, Florkowski CM.
Macro-alkaline phosphatase due to IgG-k complex: demonstration
with polyethylene glycol precipitation and immunoŽ xation. Ann Clin
Biochem 2002; 39: 523-5
Accepted for publication 28 April 2003