See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/40036960

The Beckman DxI 800 prolactin assay demonstrates superior specificity for
monomeric prolactin

Article  in  Clinical Chemistry and Laboratory Medicine · November 2009

DOI: 10.1515/CCLM.2010.038 · Source: PubMed

CITATIONS READS

8 564

4 authors, including:

Brendan Byrne Paula M. O'Shea

Beaumont Hospital Galway University Hospitals
17 PUBLICATIONS   278 CITATIONS    128 PUBLICATIONS   604 CITATIONS   

SEE PROFILE SEE PROFILE

William P Tormey
Ulster University
158 PUBLICATIONS   3,052 CITATIONS   

SEE PROFILE

Some of the authors of this publication are also working on these related projects:

The Irish Reference Interval Harmonisation Project View project

NATIONAL LABORATORY HANDBOOK Laboratory Testing View project

All content following this page was uploaded by William P Tormey on 20 May 2016.

The user has requested enhancement of the downloaded file.

 Article in press - uncorrected proof
Clin Chem Lab Med 2010;48(2):205–208
2010 by Walter de Gruyter • Berlin • New York. DOI 10.1515/CCLM.2010.038
Short Communication
The Beckman DxI 800 prolactin assay demonstrates superior
specificity for monomeric prolactin
Brendan Byrne*, Paula O’Shea, Patricia Barrett
and William Tormey
Department of Chemical Pathology, Beaumont Hospital,
Dublin, Ireland
Abstract
Background: Commercially available prolactin immunoas-
says detect macroprolactin to variable degrees. Best practice
requires laboratories to assess the cross-reactivity of their
prolactin assay with macroprolactin, and where appropriate,
introduce a screen for the presence of macroprolactin. Our
policy has been to reanalyse hyperprolactinaemic samples
following polyethylene glycol (PEG) precipitation and to
report the resultant value as the monomeric prolactin content
of the sample. The goal of this study was to determine the
need to continue PEG precipitation when prolactin measure-
ments with the Wallac AutoDELFIA were replaced by the
Beckman DxI 800.
Methods: A total of 317 apparently hyperprolactinaemic
samples were analysed for prolactin using the Beckman DxI
800 and results compared with those determined with the
PEG screening technique on the Wallac AutoDELFIA. Any
samples demonstrating a discordance of )25% were re-
analysed using gel filtration chromatography (GFC) for a
definitive result.
Results: The results indicate the Beckman DxI overestimates
the prolactin concentration in 1%–2% of hyperprolactinae-
mic samples. PEG precipitation prior to analysis with the
Wallac AutoDELFIA resulted in a 1% false negative diag-
nosis of hyperprolactinaemia.
Conclusions: Prolactin results can be reported directly from
the DxI. When results are discordant with the clinical pres-
entation, prolactin should be measured using GFC.
Clin Chem Lab Med 2010;48:205–8.
Keywords: hyperprolactinaemia; macroprolactin; macropro-
lactinaemia; screening; prolactin.
*Corresponding author: Brendan Byrne, Department of Chemical
Pathology, Beaumont Hospital, Dublin 9, Ireland
Phone: q353-1-8092668, Fax: q353-1-8093217,
E-mail: brendanbyrne@beaumont.ie
Received July 20, 2009; accepted October 2, 2009;
previously published online November 30, 2009
Unauthenticated
Download Date | 4/7/16 8:57 PM
Prolactin in the circulation exists in at least three forms. The
predominant form is the biologically active monomeric pro-
lactin (23 kDa), but it may also be found as a 50–60 kDa
form, and a species that is )100 kDa, commonly referred
to as macroprolactin. Hyperprolactinaemia is a biochemical
description with different laboratories reporting various
method-dependent reference intervals. The upper limit of the
reference interval quoted is up to 700 mIU/L with a non-
Gaussian, positive skew distribution. However, prolactino-
mas rarely have values reported in this range. Its clinical
presentation may vary from apparently asymptomatic, to
infertility and galactorrhoea. Prolactinomas may result in
headaches, blurred vision and hypopituitarism.
The occurrence of macroprolactinaemia is relatively com-
mon and may cause diagnostic confusion. Therefore, a reli-
able assay for monomeric prolactin concentrations is
clinically important. Most forms of macroprolactin are
thought to be prolactin complexed with IgG (1). The large
molecular mass results in the molecule being confined to the
vasculature, and is thought to be biologically inactive. All
commercially available prolactin immunoassays detect
macroprolactin to variable degrees (2), and the presence of
this pseudo-hyperprolactinaemia is a common cause of mis-
diagnoses (3). As a result, laboratories are advised to assess
the cross-reactivity of their prolactin assay with macropro-
lactin, and where appropriate, to introduce a screen for the
presence of macroprolactin (1, 4). The polyethylene glycol
(PEG) precipitation procedure is a valid screening test for
macroprolactin (4, 5).
Until recently, we used the Wallac AutoDELFIA
(PerkinElmer/Wallac, Turku, Finland) immunoassay analyser
to measure prolactin. This method is known to be highly
cross-reactive (16%–25%) with macroprolactin (2). There-
fore, all apparently hyperprolactinaemic samples required re-
analysis following PEG precipitation in accordance with
current best practise and we followed this procedure in our
laboratory. All laboratory reports of hyperprolactinaemia
contained an estimation of the monomeric prolactin content
in mIU/L together with an estimation of the macroprolactin
content of each sample in mIU/L.
Initial studies suggested that the Beckman prolactin assay
may have a relatively low cross-reactivity to macroprolactin
(-3%) (6, 7). We wished to switch our prolactin assay to
the Beckman DxI 800 immunoassay analyser (Beckman
Coulter, Fullerton, CA, USA). Thus, the goal of this study
was to determine whether it remained necessary to continue
2010/383
 Article in press - uncorrected proof
206 Byrne et al.: Screening for macroprolactin may not be required on the DxI
screening hyperprolactinaemic samples for macroprolactin
on the Beckman DxI 800.
Using the Beckman DxI 800, hyperprolactinaemia was
defined as a total prolactin concentration )566 mIU/L
(pre-menopausal females), )416 mIU/L (post-menopausal
females) and )278 mIU/L (males). Ethical permission for
this study was obtained in accordance with the declaration
of Helsinki and was approved by the Ethics and Medical
Research Committee of Beaumont Hospital, Dublin, Ireland.
As part of our initial assessment of the Beckman DxI pro-
lactin assay, 42 (5 males) patient samples in whom total
prolactin concentration (AutoDELFIA) varied from 89 to
500 mIU/L and whose prolactin values were within the
respective gender-specific reference interval (male
72–580 mIU/L, female 79–652 mIU/L) were compared to
the Beckman total prolactin concentration. The Passing-
Bablok regression equation obtained was Beckmans0.82
(AutoDELFIA)–15.6, indicating a negative bias relative to
the AutoDELFIA. However, all prolactin concentrations
were found to be within the Beckman prolactin assay gender-
specific reference interval. Therefore, there was 100% con-
cordance for classification of patients.
We then examined an additional 50 (11 males) patient
samples in which total prolactin (AutoDELFIA) ranged from
291 to 697 mIU/L, and whose post-PEG monomeric prolac-
tin (AutoDELFIA) ranged from 100 to 574 mIU/L. We com-
pared the post-PEG monomeric prolactin concentration with
the total prolactin concentration determined using the Beck-
man DxI prolactin assay. The Passing-Bablok regression
equation was Beckmans0.92(AutoDELFIA)q35.4, and
again there was 100% concordance in patient classification.
Based on these studies, we were satisfied that both assays
performed equally well when investigating patients who have
prolactin concentrations -700 mIU/L.
We also assessed the imprecision of both assays. For the
AutoDELFIA; the intra-assay imprecision for mean prolactin
concentrations of 112, 1973 and 3888 mIU/L was 1.2%,
0.9%, and 1.1%, respectively. The inter-assay imprecision at
the same concentrations of prolactin was 2.7%, 3.0% and
2.8%, respectively. The assay is traceable to the WHO 3rd
International Standard for prolactin 84/500.
For the Beckman DxI, the intra-assay imprecision for
mean prolactin concentrations of 135.8, 359.4 and 780
mIU/L was 1.5%, 3.9% and 3.2%, respectively. The inter-
assay imprecision at mean prolactin concentrations of 110,
309 and 542 mIU/L was 4.0%, 4.3% and 4.3%, respectively.
This compared favourably with the manufacturer’s claim for
inter-assay imprecision for prolactin concentrations at 89,
232, and 499 mIU/L of 6.9%, 3.3% and 4.2%, respectively.
This assay also is traceable to the WHO 3rd International
Standard for prolactin 84/500.
We obtained 317 apparently hyperprolactinaemic samples
following an initial Beckman analysis. All these samples
were also analysed undiluted with the AutoDELFIA, and
then re-analysed following PEG precipitation (8). The latter
technique had previously been validated in our laboratory
and was used to report patient results. Consequently, we used
Unauthenticated
Download Date | 4/7/16 8:57 PM
these results as our ‘‘in-house’’ reference method or ‘‘silver
standard’’ technique.
The post-PEG precipitation prolactin results obtained
using the AutoDELFIA were compared with the untreated
total prolactin values obtained using the Beckman assay. The
difference in prolactin concentrations determined using the
DxI and the post-PEG procedure using the Wallac assay was
calculated and expressed as a percentage. PEG precipitation
has been reported to co-precipitate up to 25% of monomeric
prolactin (8). Therefore, an arbitrary cut-off for ‘discordance’
of )25% was chosen. In cases where results obtained using
the post-PEG treatment on the AutoDELFIA were in rela-
tively close agreement with the untreated samples analysed
on the DxI, further analysis was deemed unnecessary. How-
ever, any samples demonstrating discordance )25%
required further assessment to determine which of the two
methods most accurately reflected the true situation. Gel fil-
tration chromatography (GFC) is considered the gold stan-
dard method for determining true monomeric prolactin
concentration. GFC analysis was kindly performed for us by
Mr. Fahie-Wilson and his group at Southend, UK.
Using our criteria, only 9 of 317 (2.8%) patient samples
demonstrated ‘discordance’ (range 27%–80%) when results
from the two methods were compared. Therefore, 97% of
the samples demonstrating an increased total prolactin value
on the DxI were deemed to be genuine cases of hyperpro-
lactinaemia based on the post-PEG precipitation values
obtained using the AutoDELFIA. This indicates that the DxI
has very low cross-reactivity with macroprolactin.
Of the nine samples showing discordance )25% between
the two methods, GFC was used to establish the true value
for each of these samples. The results are shown in Table 1.
The total prolactin result with the DxI approximated the
‘true’ monomeric prolactin value established by GFC in five
(56%) of these samples. The total prolactin value obtained
with the DxI markedly estimated concentrations in four of
nine (46%) patients when compared to the result obtained
with GFC. Using GFC, eight of these nine patients were
classified as genuinely hyperprolactinaemic. Classification of
patients as hyperprolactinaemic based on prolactin results
measured with the DxI was in accordance with that estab-
lished by GFC. However, three of these patients would have
been considered normoprolactinaemic had the post-PEG pre-
cipitated values obtained on the AutoDELFIA been used
alone.
GFC revealed that total prolactin concentrations obtained
with the DxI gave a more accurate reflection of the ‘true’,
or monomeric, prolactin concentration when compared with
the post-PEG treated samples analysed on the AutoDELFIA
in five of the nine discordant samples. The DxI underesti-
mated prolactin concentrations when compared with mono-
meric prolactin (GFC) in two of the nine cases. However, in
neither case would this have led to misdiagnosis or misclas-
sification of the patient had the DxI prolactin result been
reported.
In this study, the total prolactin concentration determined
with the DxI was falsely increased (range from 44% to 73%
 Article in press - uncorrected proof
Byrne et al.: Screening for macroprolactin may not be required on the DxI 207

Table 1 Post-PEG precipitated prolactin values obtained on the AutoDELFIA compared with samples analysed neat with the Beckman
DxI and monomeric prolactin values by GFC.

Patient Total PRL Total Monomeric Discordancea, % Monomeric % Macro

sample auto PRL PRL, mIU/L range 27–80 PRL, mIU/L PRL
no. DELFIA, DxI, post-PEG GFC ULR )400 GFC
mIU/L mIU/L ppt DELFIA
1 2830 992 354 64 937 77
2 1150 503 100 80 350 42
3 1230 1513 668 56 877 4
4 1470 684 492 28 781 40
5 1770 1207 832 31 711 22
6 920 822 600 27 703 4
7 1290 1477 600 59 1819 33
8 3490 1097 670 39 737 45
9 2010 831 330 60 798 74
a
Discordance (%) is the percentage difference of total prolactin on DxI and post-PEG prolactin on DELFIA. Of 317 potentially hyper-
prolactinaemic samples analysed, only nine samples gave discordant results ()25% different) when results from the two methods were
compared. Results from each of these samples are shown above along with the true monomeric prolactin value obtained using GFC.
PRL, prolactin.

higher) relative to the true monomeric prolactin result in four
of 317 (1.3%) of samples. Within this set of samples, one
patient had a normal monomeric prolactin concentration by
GFC, 350 mIU/L wRR -400 mIU/Lx and 503 mIU/L using
the DxI wRR -566 mIU/Lx. Therefore, this overestimation
would not have resulted in misdiagnosis or misclassification
if the total prolactin value from the DxI had been reported.
For the remaining patients, all had monomeric prolactin
(GFC) concentrations consistent with hyperprolactinaemia,
877, 711 and 737 mIU/L, respectively. The corresponding
results on the DxI were 1513, 1207 and 1097 mIU/L. In each
of these three patients the post-PEG precipitated prolactin
results (AutoDELFIA) of 668, 832, and 670 mIU/L more
closely approximated the ‘true’ monomeric prolactin.
The Association for Clinical Biochemistry (ACB) has sug-
gested that a threshold of 800 mIU/L be used for indicating
the need for imaging studies of the pituitary (9). However,
in women with galactorrhoea and/or oligomenorrhoea, a sus-
tained value of )600 mIU/L may also be appropriate for
indicating the need for imaging studies. Many clinicians use
a decision point of )1000 mIU/L in asymptomatic individ-
uals before imaging is considered. Based on these criteria,
the three patients described above may have been followed
up with imaging studies if the DxI results had been reported
directly from the analyser. However, given that the biochem-
ical threshold for imaging is arbitrarily defined and must take
account of the clinical context, some clinicians would con-
sider all three patients as genuine candidates for imaging.
The implication of this study is that potentially 1% of
patients may undergo further unnecessary diagnostic work-
up with added economic costs and additional psychological
stress for these patients. Of practical importance is the fact
that in 97% of cases, the results obtained on the DxI were
in agreement with the post-PEG precipitated values from the
AutoDELFIA. Furthermore, based on those samples that
showed discordance, no genuine cases of hyperprolactinae-
mia would have been misclassified had the total prolactin
value from the DxI alone been reported. However, this would
not have been true had the post-PEG prolactin values from
the AutoDELFIA been reported. Three patients had post-
PEG precipitated prolactin values of 354, 492 and 330
mIU/L, respectively, despite all three patients having true
hyperprolactinaemia when assessed using GFC. These results
suggest possible under reporting of hyperprolactinaemia in
three of 317 or 1% of patients investigated using the
AutoDELFIA in conjunction with PEG precipitation. It is
recognised that PEG treatment may result in an underesti-
mation of the monomeric prolactin content in a sample due
to co-precipitation of monomeric prolactin. For this reason,
specific post-PEG precipitation reference ranges are recom-
mended (3). However, it is quite possible that each of the
three patients described above would not have received the
necessary follow-up even if such reference ranges had been
employed. The most extreme of these results was from a
patient who had a post-PEG precipitation prolactin value of
600 mIU/L, but had a monomeric prolactin of 1819 using
GFC. Depending on the clinical presentation, this patient
may not have been properly investigated. It is important to
acknowledge that we have made an assumption that results
not showing significant discordance between the two meth-
ods accurately reflected the true prolactin status. To clarify
this issue, GFC would be required for all 317 samples.
Unfortunately, the high costs involved precluded us from per-
forming GFC on all samples.
Guidelines for the diagnosis and management of prolac-
tinomas were published by the Pituitary Society in 2006 (4).
They recommend the use of PEG precipitation to overcome
the interference from macroprolactin in assays where cross
reactivity with macroprolactin is high. Evidence from the
literature is supportive of this (1–3). However, our data sug-
gest that there is a small, but potentially significant risk,
associated with over reliance on the PEG precipitation pro-

Unauthenticated
Download Date | 4/7/16 8:57 PM
 Article in press - uncorrected proof
208 Byrne et al.: Screening for macroprolactin may not be required on the DxI
cedure. PEG treatment is non-specific for removal of large
molecular weight proteins. Our data demonstrate examples
where this may lead to serious misdiagnosis. PEG itself can
also interfere positively with assays on some immunoassay
analysers, including the Beckman system (10). In addition,
utilising testing procedures in a way that has not been
approved by the manufacturers may impact on the validity
of the CE marking and result in a breach of the in-vitro
diagnostics (IVD) directive.
In the interest of patient safety, it is better to accept a small
number of false positives. We recommend that prolactin
results should be reported directly from the DxI, without pri-
or PEG precipitation, and clinicians should be invited to con-
tact the laboratory if the value seems clinically incongruous.
In these cases definitive analysis using GFC should be
offered, before imaging is performed. To the best of our
knowledge, this is the first study to advocate this approach.
This simplifies the laboratory procedure for prolactin anal-
ysis, saving staff time and reducing costs without compro-
mising patient safety. Since conducting this study we have
presented our findings to local endocrinology teams and have
ceased screening for macroprolactin using the PEG precipi-
tation procedure. If a specific need is deemed necessary at
the multidisciplinary case conference, samples are analysed
using GFC. This protocol has been successfully employed
for more than 12 months by our neuroendocrinology service.
Acknowledgements
We wish to acknowledge the assistance provided by Mr. M.N.
Fahie-Wilson and the staff of Southend Hospital, Essex, UK, for
performing the GFC analysis.
Conflict of interest statement
Authors’ conflict of interest disclosure: The authors stated that
there are no conflicts of interest regarding the publication of this
article.
Research funding: None declared.
Unauthenticated
Download Date | 4/7/16 8:57 PM
View publication stats
Employment or leadership: None declared.
Honorarium: None declared.
References
1. Fahie-Wilson MN, John R, Ellis AR. Macroprolactin; high
molecular mass forms of circulating prolactin. Ann Clin Bio-
chem 2005;42:175–92.
2. Smith TP, Suliman AM, Fahie-Wilson MN, McKenna TJ.
Gross variability in the detection of prolactin in sera containing
big big prolactin (macroprolactin) by commercial immunoas-
says. J Clin Endocrinol Metab 2002;87:5410–5.
3. Suliman AM, Smith TP, Gibney J, McKenna TJ. Frequent mis-
diagnosis and mismanagement of hyperprolactinaemic patients
before the introduction of macroprolactin screening: application
of a new strict laboratory definition of macroprolactinaemia.
Clin Chem 2003;49:1504–9.
4. Casanueva FF, Molitch ME, Schlechte JA, Abs R, Bonert V,
Bronstein MD, et al. Guidelines of the Pituitary Society for the
diagnosis and management of prolactinoms. Clin Endocrinol
2006;65:265–73.
5. Gibney J, Smith TP, McKenna TJ. The impact on clinical prac-
tice of routine screening for macroprolactin. J Clin Endocrinol
Metab 2005;90:3927–32.
6. Dearman GO, Mackenzie PF, Fahie-Wilson MN. The Beckman
access prolactin assay and macroprolactin; prevalence and
detection of hyperprolactinaemia due to macroprolactin.
wabstract TP 1.69x. Clin Chim Acta 2005;355(Suppl):225.
7. Ellis MJ, Livesey JH, Soule SG. Macroprolactin, big-prolactin
and potential effects on the misdiagnosis of hyperprolactinemia
using the Beckman Coulter Access Prolactin assay. Clin Bio-
chem 2006;39:1028–34.
8. Kavanagh L, McKenna TJ, Fahie-Wilson MN, Gibney J, Smith
TP. Specificity and clinical utility of methods for the detection
of macroprolactin. Clin Chem 2006;52:1366–72.
9. Barth JH, Butler GE, Hammond PH. Biochemical investiga-
tions in Laboratory Medicine. ACB Ventura Publications 2001:
25–7.
10. Fahie-Wilson M, Dearman G. The Beckman access prolactin
assay and macroprolactin; prevalence and detection of hyper-
prolactinaemia due to macroprolactin. Clin Chem 2005;51
(Suppl A231).