Original Article

Protein and albumin-to-creatinine ratios in random urines

accurately predict 24 h protein and albumin loss
in patients with kidney disease

Mark Guy1, Joanna K Borzomato1,5, Ronald G Newall2, Philip A Kalra3 and Christopher P Price4
1
Department of Clinical Biochemistry, Salford Royal NHS Foundation Trust, Hope Hospital, Salford; 2Highover Park, Amersham, Bucks;
3
Department of Renal Medicine, Salford Royal NHS Foundation Trust, Salford; 4Department of Clinical Biochemistry, University of Oxford,
Oxford, UK; 5Present address: Department of Clinical Biochemistry, Royal Albert Edward Infirmary, Wigan, UK
Corresponding author: Mark Guy. Email: mark.guy@srft.nhs.uk

Abstract
Background: Random urine protein-to-creatinine (PCR) and albumin-to-creatinine (ACR) ratios have been proposed as
alternatives to 24 h urine measurements to simplify sample collection and overcome errors. The aim of this study was to
examine the ability of PCR and ACR to predict urinary 24 h protein and albumin loss, respectively, in patients with kidney
disease, and determine the most appropriate time of collection.
Methods: Eighty-three patients were recruited from a renal outpatient clinic. In a 24 h period, each collected an early-morning
urine (EMU), second and third voids, and the remaining urine passed that day. PCR and ACR were determined in random
urines and compared with the 24 h loss of protein and albumin, respectively.
Results: For all patients, median (range) 24 h urine protein and albumin losses were 220 (30– 15600) and 60 (,8 –10,557) mg,
respectively. Ratios derived from each of three random urines correlated well with 24 h protein or albumin loss (Spearman’s
rs . 0.87, P , 0.0001). Receiver operator characteristic (ROC) curve analysis showed PCR accurately predicted both an
abnormal 24 h urine protein !150 mg/24 h (areas under curves [AUC] 0.90–0.92) and significant proteinuria above 300 mg/
24 h (AUC between 0.97 and 1.00). ACR accurately predicted both an abnormal 24 h urine albumin !30 mg/24 h (AUC 0.98 to
0.99) and frank albuminuria at !300 mg/24 h or !700 mg/24 h (AUC between 0.99 and 1.00). EMU and random urines
performed equally well in predicting proteinuria and albuminuria from PCR and ACR, respectively.
Conclusions: By careful choice of cut-offs, both PCR and ACR can be used in patients with kidney disease to rule in or
rule out abnormal 24 h losses of protein and albumin. EMU and, importantly, random samples can be used as surrogates for
24 h urine collections.

Ann Clin Biochem 2009; 46: 468– 476. DOI: 10.1258/acb.2009.009001

Introduction urine collection, the variability of results obtained by this

The identification and quantitation of proteinuria is of impor-
tance in the initial diagnosis and subsequent follow-up and
monitoring of renal disease.1,2 Proteinuria is also recognized
as an independent risk factor for cardiovascular disease and
as a predictor of end organ damage.3 – 5 More specifically,
urine albumin measurement now has an established role in
the early identification and monitoring of diabetic nephropa-
thy6 and, relatively recently, has been identified as an indi-
cator of cardiovascular risk in renal, hypertensive and other
patients.7,8 Thus, urine protein or albumin measurements
now form a central part of the classification and guidelines
for chronic kidney disease (CKD).9 – 12
There is no ‘gold standard’ for identifying proteinuria
and, although it has been generally accepted that the best
measure of protein or albumin loss is that based on a 24 h
method make this view questionable. Quoted reference
ranges for 24 h protein loss vary, but most would regard a
protein loss of more than 150 mg/24 h as abnormal with sig-
nificant proteinuria being greater than 300 mg/24 h.13
Urine albumin measurements have been used as a more sen-
sitive measure and early marker for nephropathy6 and there is
now much support advocating the use of urine albumin
measurement rather than total protein14,15 because detection
of ‘low-level’ albuminuria, important in the therapeutic
approach to a patient, may be masked by tests that measure
total protein. Urine albumin losses in the normal individual
are usually ,30 mg/24 h (and often below 10 mg/24 h),
with 30–300 mg/24 h being within the ‘microalbuminuric’
range.16,17 Urinary losses above 300 mg/24 h would be
regarded as frank albuminuria (clinically overt proteinuria).

Annals of Clinical Biochemistry 2009; 46: 468 –476

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................................................................................................................................................
Twenty-four hour urine collections, however, can be cum-
bersome to perform and prone to errors in collection.18 For
this reason, the use of protein or albumin-to-creatinine ratios
(PCR or ACR) in random urine samples has been pro-
posed.19 ACRs have been evaluated extensively in patients
with diabetes20 – 22 and several studies have examined the
utility of PCRs in predicting proteinuria, primarily in hyper-
tensive pregnancy23,24 and paediatric settings.25,26 There
have been relatively few studies in renal disease27 – 30 and
there is little data at present which shows the ability of
ACR to predict proteinuria in non-diabetic patients. Most
studies have demonstrated a good relationship between cal-
culated ratios and 24 h protein loss and some have quoted
predictive values for identifying significant proteinuria.
Few, however, have indicated the confidence with which
creatinine ratios can rule in or rule out proteinuria. One sys-
tematic review of studies using random urine PCR con-
cluded that they provided evidence to rule out significant
proteinuria as defined by a 24 h measurement.13
The aim of this study was to assess the ability of PCR and
ACR from random urine samples to rule in or rule out pro-
teinuria or albuminuria in patients with kidney disease
attending a nephrology clinic. In addition, by collecting
urine at different points during the day, the most appropri-
ate random urine sample, if any, could be determined.
Patients
A total of 117 outpatients with various stages of CKD and
requiring 24 h urine total protein measurement as part of
their clinical investigation or management were enrolled
in the study between September and December 2005.
Patients were consecutive and recruited from one consult-
ant’s (PAK) nephrology clinic. Other parts of the study
have been reported elsewhere.31,32 The study was approved
by Greater Manchester Local Ethics Committees with signed
written consent being obtained from each patient.
Methods
Each patient was asked to collect urine in the 24 h period
immediately prior to their nephrology clinic appointment
as follows. After discarding the first sample of the day,
urine was collected in four separate containers: (i) second
void; (ii) third void; (iii) remaining urine passed that day;
and (iv) early-morning urine (EMU) passed the following
day. Urine was collected into containers without preserva-
tive and stored at 48C prior to analysis, which was usually
within 24– 48 h of receipt.
Urine protein, albumin and creatinine were measured on
each of samples (i), (ii) and (iv) and PCR and ACRs (both
mg/mmol) calculated. All four aliquots were then pooled,
the volume measured, and protein and albumin measure-
ments repeated to give the 24 h outputs of each.
All assays were performed on the Roche Integra 800
(Roche Diagnostics Ltd, Burgess Hill, UK) using reagent
kits and calibrators supplied by the manufacturer. Urine
protein was measured using pyrogallol red,33 urine creati-
nine by a kinetic Jaffe method34 and urine albumin by
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Guy et al. PCR and ACR in patients with kidney disease 469
immunoturbidimetry.35 The lower limits of detection for
urine protein and albumin assays were 20 and 8 mg/L,
respectively. Between-day coefficients of variation for the
urine protein assay were 2.2% and 4.3% at concentrations
of 161 and 550 mg/L, respectively. For the urine albumin
assay, they were 2.0% and 2.6% at concentrations of
26.7 and 97.5 mg/L, respectively and for urine creatinine
they were 1.7% and 1.8% at concentrations of 12.7 and
5.7 mmol/L, respectively. For the purposes of this study,
random urines with undetectable protein or albumin con-
centrations were, respectively, designated as having a PCR
of 5 mg/mmol and ACR of 0.5 mg/mmol. The 24 h urines
with undetectable albumin were designated an albumin
loss of 5 mg/24 h.
Statistical analyses on the data were performed using
Analyse-it for Excel (Analyse-it Software Ltd, Leeds, UK).
Spearman’s rank correlation was used to compare urine
PCR and ACR with 24 h protein and albumin loss, respect-
ively. The ability of urine PCR and ACR at various cut-offs
to predict abnormal 24 h protein or albumin loss was deter-
mined from receiver operator characteristic (ROC) curve
analysis and by calculation of sensitivities, specificities, pre-
dictive values and likelihood ratios (LRs). A negative likeli-
hood ratio (LR[2], false-positive rate/true negative rate) of
,0.1 and positive likelihood ratio (LR[þ], true positive rate/
false-negative rate) greater than 10 were taken to indicate
good test performance.36 Areas under ROC curves were
compared using the method of DeLong et al. 37
Results
Patient characteristics
Of the 117 patients who consented to be involved in the
study, 86 (73.5%) returned urine samples. Two urine
collections of ,500 mL in volume and one having a 24 h
urine creatinine output ,3.0 mmol were rejected on the
assumption that they might not have been complete 24 h
collections. The main contributory causes of CKD in the
remaining 83 patients were diabetic nephropathy (n ¼ 17),
hypertensive damage (n ¼ 13), renovascular (n ¼ 18),
glomerulonephritis/glomerulosclerosis (n ¼ 16) (including
IgA nephropathy [n ¼ 6]), tubulointerstitial damage (n ¼
5) and amyloidosis (n ¼ 1). Causes in 10 patients were not
certain or under investigation.
There were 25 patients with diabetes mellitus and 53 had
documented hypertension (10 of these patients were subop-
timally controlled). One patient had stage 1 CKD, with
11 (13%), 35 (42%), 27 (33%) and 9 (11%) patients having
stages 2, 3, 4 and 5, respectively. Seventy-two percent
were men and the median age for all patients was 67
years. For all patients, median (range) 24 h urinary protein
loss was 220 (30– 15,600) mg/24 h and median (range)
24 h albumin loss was 60 (,8– 10557) mg/24 h (Table 1).
Protein:creatinine ratio versus 24 h urine protein
There was a very good correlation between 24 h urine protein
loss and PCR in second and third voids and EMUs
(Figures 1a–c), and these correlations were very similar for
470 Annals of Clinical Biochemistry Volume 46 November 2009
................................................................................................................................................

(a) 10,000

albuminuria
Prevalence
Table 1 Patient characteristics and median (range) age, 24 h urine volumes and outputs of protein, albumin and creatinine. Prevalence data refers to patients with 24 h urinary loss of protein
1000

65.1
70.0
52.2
(%)

PCR (mg/mmol)
100

Prevalence
proteinuria 10

65.1
70.0
52.2
(%)

1
10 100 1000 10,000 100,000
60 (,8–10557)
66 (,8–10557)
Urine Albumin

40 (,8–1531)
Urine total protein (mg/24 h)
mg/24 h

(b) 10,000

1000
220 (30 – 15600)
220 (50 – 15600)

PCR (mg/mmol)
195 (30 – 1650)
Urine Protein

100
mg/24 h

10
1.57 (0.70 –2.9)

1.43 (0.96 –2.9)

Urine volume

1.60 (0.7 – 2.7)

1
10 100 1000 10,000 100,000
L/24 h

Urine total protein (mg/24 h)

(c) 10,000
7.7 (3.5 – 16.4)
8.5 (4.4 – 16.4)
6.6 (3.5 –9.6)
mmol/24 h
creatinine

1000
PCR (mg/mmol)
Urine

100

10
67 (28 –86)
67 (28 –86)
67 (33 –80)
Age (y)

1
10 100 1000 10,000 100,000
Urine total protein (mg/24 h)
5
9
4
5

Figure 1 Protein-to-creatinine ratio measured in random urines compared

with 24 h urine protein loss. (a) ¼ Second void, (b) ¼ third void, (c) ¼ early-
morning urine. Spearman rank correlation coefficients were (a) 0.90, (b)
27
19
8
4

0.91, (c) 0.87; P , 0.0001 for all

each of the three urine aliquots. ROC curve analysis showed

35
28
7
3
with CKD stage

PCR to be a good predictor of both abnormal urine protein

No. of patients
!150 mg/24 h or albumin !30 mg/24 h

loss !150 mg/24 h and clinically significant urine protein

11
9
2

loss above 300 mg/24 h (Table 2). There were no significant

2

differences between areas under curves (AUC) generated

from each of the three urine aliquots (P . 0.05 for all
CKD, chronic kidney disease
1
1
0
1

cases), suggesting they were equally good at predicting pro-

teinuria. The ability of PCR to predict or exclude significant
83
60
23
n

24 h protein loss of !500 mg was assessed for each aliquot

at various PCR cut-offs (Table 3). For each PCR cut-off, calcu-
lated LRs and predictive values were similar between each of
All patients

Females

the three urine aliquots. At cut-offs of 70 mg/mmol and

Males

below, PCR showed good ability in ruling out significant

proteinuria denoted by negative likelihood ratios (LR2) of

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 Guy et al. PCR and ACR in patients with kidney disease 471
................................................................................................................................................

Table 2 Areas under curves (AUC) generated from receiver operator (Figures 2a –c) and again correlations were very similar for
characteristic analysis showing the ability of protein-to-creatinine each of the three urine aliquots. ROC curve analysis
ratios (PCRs) from random urines to predict 24 h urine protein loss
!150, !300, !500 or 1000 mg/24 h
showed that ACR in each random urine adequately
predicted a 24 h albumin loss of !30, !300 and !700 mg
Urine total protein
(Table 5). EMU, and second and third voids were equally
loss (mg/24 h) Urine aliquot AUC 95% CI AUC
good at predicting albuminuria, as there was no significant
!150 Second void 0.91 0.85 –0.97 difference between AUCs generated from each of the three
Third void 0.92 0.87 –0.98
EMU 0.90 0.84 –0.97
urine aliquots (P . 0.05 for all cases). Using an ACR
!300 Second void 0.97 0.94 –1.00 cut-off of 30 mg/mmol, all three urine aliquots showed a
Third void 0.98 0.96 –1.00 similar good performance in being able to rule in or rule
EMU 0.97 0.93 –1.00 out significant albuminuria (!300 mg/day), denoted by
!500 Second void 1.00 0.99 –1.00
LRþ .10 and LR2 ,0.1, respectively (Table 6). All three
Third void 0.99 0.97 –1.00
EMU 0.99 0.98 –1.00 urine aliquots showed similar ability in predicting albumi-
!1000 Second void 0.99 0.97 –1.00 nuria for 24 h outputs of 300 and 700 mg. ACR cut-offs
Third void 0.99 0.97 –1.00 that gave optimum sensitivities and specificities for predict-
EMU 0.98 0.96 –1.00 ing albuminuria at these levels are shown in Table 7.
AUC, areas under curves; CI, confidence interval
(EMU, early-morning urine)
Total protein versus albumin
approximately 0.1 or below. The ability to rule in significant Figure 3 shows the relationship between albumin and total
proteinuria, denoted by positive likelihood ratios (LRþ) of protein in 24 h urines (n ¼ 83). At significant urinary total
approximately 10 or above, was seen at PCR cut-offs above protein losses (.1000 mg/24 h), there was a good corre-
50 mg/mmol. All three urine aliquots also showed similar lation (Spearman’s r ¼ 0.87, P , 0.0001) between the two
ability in predicting proteinuria for other 24 h outputs of with albumin contributing to the majority of the total
150, 300 and 1000 mg. PCR cut-offs that gave optimum protein present (median 89%, 95% range 67– 104%). With
sensitivities and specificities for predicting proteinuria at urinary losses .1000 mg/24 h, the Cusum linearity test
these levels are shown in Table 4. showed a linear relationship between the two (P . 0.1).
At lower urinary protein losses (,1000 mg/24 h), albumin
as a percentage of total protein was much more variable
Albumin:creatinine ratio versus 24 h urine albumin (Spearman’s r ¼ 0.71, P , 0.0001, median 27%, 95% range
For each patient, ACR from second and third voids and 2 –92%) and the linear relationship between the two was
EMUs correlated extremely well with 24 h albumin loss lost (Cusum linearity test, P , 0.01).

Table 3 Likelihood ratios (LRs) and predictive values for each urine aliquot in predicting a 24 h urine protein loss of !500 mg/24 h
PCR
cut-off Sensitivity Specificity
(mg/mmol) Urine (%) (%) PV1 PV2 LR1 LR2
10 (a) 100 6 0.36 1.00 1.1 0.0
(b) 100 9 0.37 1.00 1.1 0.0
(c) 100 7 0.37 1.00 1.1 0.0
15 (a) 100 19 0.40 1.00 1.2 0.0
(b) 100 22 0.41 1.00 1.3 0.0
(c) 100 35 0.45 1.00 1.5 0.0
20 (a) 100 30 0.43 1.00 1.4 0.0
(b) 100 41 0.48 1.00 1.7 0.0
(c) 100 54 0.54 1.00 2.2 0.0
30 (a) 100 56 0.55 1.00 2.3 0.0
(b) 100 69 0.63 1.00 3.2 0.0
(c) 100 76 0.69 1.00 4.2 0.0
50 (a) 100 87 0.81 1.00 7.7 0.0
(b) 93 85 0.77 0.96 6.3 0.1
(c) 100 87 0.81 1.00 7.7 0.0
70 (a) 100 96 0.94 1.00 27.0 0.0
(b) 90 98 0.96 0.95 48.4 0.1
(c) 83 100 1.00 0.93 high 0.1
100 (a) 69 100 1.00 0.86 high 0.3
(b) 70 100 1.00 0.86 high 0.3
(c) 69 100 1.00 0.86 high 0.3
200 (a) 38 100 1.00 0.75 high 0.6
(b) 38 100 1.00 0.75 high 0.6
(c) 28 100 1.00 0.72 high 0.7

PCR, protein-to-creatinine ratio

(a) ¼ second void, (b) ¼ third void, (c) ¼ early-morning urine. PVþ, PV2 ¼ positive, negative predictive values. LRþ, LR2 ¼ positive, negative LRs

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472 Annals of Clinical Biochemistry Volume 46 November 2009
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Table 4 Optimum PCR cut-offs for predicting 24 h urine protein Table 5 AUCs generated from ROC analysis showing the ability of
losses of 150, 300, 500 and 1000 mg ACRs from random urines to predict 24 h urine albumin losses !30,
!300 and !700 mg/24 h
Total
protein PCR Sensitivity Specificity Urine albumin
mg/24 h Urine cut-off (%) (%) LR1 LR2 loss (mg/24 h) Urine aliquot AUC 95% CI AUC
150 (a) 31 84 86 5.9 0.2 !30 Second void 0.98 0.96 –1.00
(b) 27 84 86 5.9 0.2 Third void 0.99 0.98 –1.00
(c) 23 78 79 3.7 0.3 EMU 0.98 0.96 –1.00
300 (a) 47 91 92 11.0 0.1 !300 Second void 1.00 0.99 –1.00
(b) 45 91 92 11.0 0.1 Third void 1.00 1.00 –1.00
(c) 50 94 94 15.0 0.1 EMU 1.00 1.00 –1.00
500 (a) 72 97 96 26.1 0.0 !700 Second void 1.00 1.00 –1.00
(b) 57 93 93 16.8 0.1 Third void 0.99 0.97 –1.00
(c) 60 93 94 16.8 0.1 EMU 1.00 0.99 –1.00
1000 (a) 123 93 93 12.8 0.1
(b) 115 93 93 12.8 0.1 AUC, areas under curves; CI, confidence interval; ACR,
(c) 105 93 93 12.8 0.1 albumin-to-creatinine ratio
(EMU, early-morning urine)
PCR, protein-to-creatinine ratio; LRþ, positive likelihood ratio; LR2,
negative likelihood ratio
(a) ¼ second void, (b) ¼ third void, (c) ¼ early-morning urine. LRþ, LR2 ¼ Twenty-nine out of 83 patients had evidence of clinically
positive, negative likelihood ratios significant proteinuria with urine protein !500 mg/24 h. Six
(a) 10,000
of these samples had 24 h urinary albumin losses ,300 mg
(range ,8– 196 mg). There were no patients with urine
1000
albumin !300 mg/24 h and urine protein , 500 mg/24 h.
Similarly 15 patients had evidence of heavy proteinuria
ACR (mg/mmol)

100 with urine protein !1000 mg/24 h with only one having
urine albumin levels lower than 700 mg/24 h (684 mg/
10 24 h). Three patients had urine albumin .700 mg/24 h
and urine protein lower than 1000 mg/24 h (values
1 between 740 and 780 mg/24 h). Table 8 shows optimum
sensitivities and specificities for random urine ACR in pre-
0 dicting 24 h protein outputs of 150, 300, 500 and 1000 mg.
1 10 100 1000 10,000 100,000
Urine albumin (mg/24 h)
(b) 10,000 Discussion
1000 Collection of 24 h urine samples is cumbersome and prone
to errors, hence analyte measurement in random urine is
ACR (mg/mmol)

100 often proposed with correction for variation in urine flow

rate by expressing results as a ratio to creatinine concen-
10
tration. It was not possible to formally assess completeness
1
of collection of 24 h samples, particularly as there is little
published data for patients with CKD. Although samples
0 were from a well-motivated group of patients who regularly
1 10 100 1000 10,000 100,000 made 24 h urine collections, a pragmatic approach was
Urine albumin (mg/24 h) adopted to reject samples that had either unusually low
(c) 10,000
urine volume or creatinine outputs, on the premise that
they may not have been a complete 24 h collection. As a
1000 result, only 71% of patients enrolled in this study returned
or provided viable samples highlighting the inconvenience
ACR (mg/mmol)

100 and errors associated with 24 h urine collection.

Several studies have assessed the benefits of using
10 random urine PCR as a surrogate for 24 h protein measure-
ment, but relatively few of these have involved patients with
1 kidney disease, or have examined the ability of PCRs or
ACRs to rule in or rule out abnormal protein loss. Most
0 studies have shown good correlations between PCR and
1 10 100 1000 10,000 100,000 24 h protein loss and have demonstrated adequate sensi-
Urine albumin (mg/24 h) tivities and specificities when PCR was used to predict
24 h proteinuria. However, cut-offs used to define abnormal
Figure 2 Albumin-to-creatinine ratio measured in random urines compared
PCR or abnormal protein loss have varied widely. Thus,
with 24 h urine albumin loss. (a) ¼ Second void, (b) ¼ third void, (c) ¼ early-
morning urine. Spearman rank correlation coefficients were (a) 0.97, (b) Gai et al. 27 in predicting a 24 h urine protein loss .150 mg
0.96, (c) 0.97; P , 0.0001 for all from a PCR ! 11.3 mg/mmol, obtained sensitivities and

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 Guy et al. PCR and ACR in patients with kidney disease 473
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Table 6 LRs and predictive values for each urine aliquot in predicting a 24 h urine albumin loss of !300 mg/day
ACR cut-off
(mg/mmol) Urine Sensitivity (%) Specificity (%) PV1 PV2 LR1 LR2
10 (a) 100 72 0.58 1.00 3.5 0.0
(b) 100 68 0.55 1.00 3.2 0.0
(c) 100 78 0.64 1.00 4.6 0.0
15 (a) 100 85 0.72 1.00 6.7 0.0
(b) 100 83 0.70 1.00 6.0 0.0
(c) 100 90 0.79 1.00 10.0 0.0
20 (a) 100 92 0.82 1.00 12.0 0.0
(b) 100 87 0.74 1.00 7.5 0.0
(c) 100 93 0.85 1.00 15.0 0.0
30 (a) 100 93 0.85 1.00 15.0 0.0
(b) 100 93 0.85 1.00 15.0 0.0
(c) 100 97 0.92 1.00 30.0 0.0
50 (a) 96 98 0.96 0.98 57.4 0.1
(b) 91 98 0.96 0.97 54.8 0.1
(c) 92 100 1.00 0.97 High 0.1
70 (a) 87 98 0.95 0.95 52.2 0.2
(b) 91 98 0.96 0.97 54.8 0.1
(c) 78 100 1.00 0.92 High 0.2
100 (a) 78 100 1.00 0.92 High 0.2
(b) 74 100 1.00 0.91 High 0.3
(c) 57 100 1.00 0.86 High 0.4
200 (a) 39 100 1.00 0.79 High 0.6
(b) 44 100 1.00 0.82 High 0.6
(c) 30 100 1.00 0.79 High 0.7

ACR, albumin-to-creatinine ratio; LR, likelihood ratio;

(a) ¼ second void, (b) ¼ third void, (c) ¼ early-morning urine. PVþ, PV2 ¼ positive, negative predictive values. LRþ, LR2 ¼ positive, negative LRs

Table 7 Optimum ACR cut-offs for predicting 24 h urine albumin 100,000

outputs of 30, 300, and 700 mg
Urine ACR
albumin cut- Sensitivity Specificity 10,000
mg/24 h Urine off (%) (%) LR1 LR2
Albumin (mg/24 h)

30 (a) 6 93 93 13.4 0.1

1000
(b) 6 96 97 27.9 0.0
(c) 4 96 97 27.9 0.0
300 (a) 50 96 98 57.4 0.0
(b) 43 96 98 57.4 0.0 100
(c) 40 100 100 high 0.0
700 (a) 109 94 99 62.1 0.1
(b) 80 88 92 11.7 0.1 10
(c) 65 94 97 31.0 0.1

ACR, albumin-to-creatinine ratio 1

(a) ¼ second void, (b) ¼ third void, (c) ¼ early-morning urine. LRþ, 1 10 100 1000 10,000 100,000
LR2 ¼ positive, negative likelihood ratios
Protein (mg/24 h)

Figure 3 Relationship between albumin and total protein in 24 h urines

specificities of 91% and 75%, respectively. Similarly Lane
et al.,30 using thresholds of between 300 mg/L and 3.0 g/L
to define abnormal 24 h proteinuria, obtained sensitivities
between 85% and 93% and specificities between 80% and
91% at varying PCR cut-offs.
Many studies have used a 24 h protein output of greater
than 300 mg to indicate clinically significant proteinuria.13
Recent guidelines have used higher levels of 500 mg/24 h
and 1 g/24 h (roughly equivalent to PCR of 50 and
100 mg/mmol, respectively) to denote significant proteinuria
and to define levels at which treatment should be insti-
tuted.9,12,38 In this study, we have demonstrated good corre-
lation between 24 h urine protein loss and PCR in second and
third voids and EMUs (Figures 1a–c). Furthermore, ROC
curve analysis at multiple 24 h protein cut-offs showed PCR
(n ¼ 83). Samples with undetectable albumin levels have been assigned a
24 h value of 5 mg
to be a good predictor of 24 h urine protein loss (Table 2).
Importantly, there was no statistical difference between
areas under the curves generated from EMU, second or
third voids suggesting any would be equally effective as a
substitute for the 24 h urine. In addition, the data show
that, by using an appropriate cut-off, PCR can be a reliable
tool for ruling in or ruling out abnormal 24 h protein loss.
For example, at PCR greater than 50 mg/mmol, there is
strong evidence to be able to rule in significant proteinuria
of 500 mg/24 h or more, demonstrated by LRþ greater
than 10. Likewise at PCR below 50 mg/mmol, there is
strong evidence to be able to rule out significant 24 h

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474 Annals of Clinical Biochemistry Volume 46 November 2009
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Table 8 Optimum albumin-to-creatinine ratio cut-offs for predicting Recent guidelines have recommended the use of urine
24 h urine protein outputs of 150, 300, 500, 1000 mg albumin measurement in preference to total protein.9,10,12
Total ACR Some also quote approximate equivalence points for urine
protein cut- Sensitivity Specificity protein and albumin. For example, the National Institute
mg/24 h Urine off (%) (%) LR1 LR2
for Health and Clinical Excellence12 quote ACRs of 30 and
150 (a) 6 72 72 2.6 0.4 70 mg/mmol to be roughly approximate to 24 h protein
(b) 7 72 76 3.0 0.4 outputs of 500 mg and 1 g, respectively. The Scottish
(c) 5 79 80 4.0 0.3
300 (a) 12 86 84 5.9 0.2
Intercollegiate Guidelines Network38 quote an ACR of
(b) 11 80 83 4.8 0.2 30 mg/mmol being equivalent to protein loss of 450 mg/
(c) 9 83 83 5.0 0.2 24 h. Thresholds for diagnosis and treatment of clinically
500 (a) 15 86 87 6.7 0.2 significant proteinuria have therefore been suggested at
(b) 17 86 87 6.7 0.2
ACR levels of 30 and 70 mg/mmol (approximately equival-
(c) 12 86 87 6.7 0.2
1000 (a) 109 93 96 21.2 0.1 ent to 24 h albumin outputs of 300 and 700 mg).9,10,12 Our
(b) 93 93 96 21.2 0.1 data suggest that either urine albumin or urine total
(c) 64 87 93 11.8 0.1 protein measurements would be equally good in assessing
or monitoring significant proteinuria, particularly for
ACR, albumin-to-creatinine ratio
(a) ¼ second void, (b) ¼ third void, (c) ¼ early - morning urine. LRþ, protein outputs greater than 1 g/24 h. Below 1 g/24 h, the
LR2 ¼ positive, negative likelihood ratios situation is less clear as the correlation between albumin
and total protein is more variable. This is highlighted by
the data in Table 8. Optimum ACR cut-offs for predicting
protein loss, as demonstrated by LR2 ,0.1 (Table 3). 24 h protein outputs at levels ,1 g/24 h are much lower
Similarly random urine PCR may be used effectively to than might be expected. Furthermore, six out of 29 patients
rule in or rule out other levels of proteinuria (Table 4). with significant proteinuria .500 mg/24 h had albumin
ACR now has an established role in the diagnosis outputs lower than the 300 mg/24 h threshold. If these
and monitoring of diabetic nephropathy.20 There are an data can be extrapolated to other patient populations, indi-
increasing number of studies using ACR in patients with viduals would have been potentially misclassified using
hypertension8,39 – 41 but there are few studies in other, non- guidelines based solely on ACR. This is in contrast to
diabetic groups. Most studies, however, have shown that Collier et al.,49 who found few discordant ACR and PCR
random urine ACR correlates well with 24 h albuminuria results in a study of 117 patients.
and demonstrate adequate sensitivity and specificity in pre- ACR measurements, however, have distinct advantages
dicting 24 h albuminuria.42 – 45 This study in renal outpati- due to the superior analytical sensitivity of assays. This is
ents has also shown good agreement between ACR and especially true for detecting early nephropathy in diabetics
24 h albumin loss (Figure 2) and ACR from each of the and possibly other high-risk groups, for example, patients
first, second and third voids predict 24 h albumin output with hypertension or cardiovascular disease.
equally well (Table 5). An ACR cut-off of 30 mg/mmol accu- This study has been limited by its relatively small size and
rately rules in or out significant albuminuria !300 mg/24 h, being restricted to one centre using hospital patients with
as denoted by LRþ .10 or LR2 ,0.1, respectively kidney disease. Owing to the small sample size, there
(Table 6). Lower cut-offs of between 4 and 6 mg/mmol were insufficient results to delineate the data by gender or
could be used to rule in or rule out albumin loss in the renal pathology. In diabetic patients, it has been demon-
‘microalbuminuric’ range, which is perhaps of greater strated that gender-specific cut-off values improve the diag-
importance in screening for renal disease (Table 7). nostic performance of ACR.21,50 Use of such cut-offs may
Most guidelines state that EMU is the preferred sample enhance the performance of ACR in patients with kidney
for ACR and PCR measurements, but this study has not disease. Further work needs to be undertaken to provide
found any advantage in using EMU over random such data and to establish whether our data can be extrapo-
samples. Reports in the literature are conflicting with lated to other patient groups.
some advocating EMU20,46 and others random urine In summary this study has shown that, in patients with
samples.18,47 There are benefits to both patients and clini- kidney disease, both random urine PCR and ACR accurately
cians if random ‘spot’ urines can be relied upon to accu- predict proteinuria and albuminuria, respectively. By
rately rule in or rule out proteinuria and provide an careful choice of cut-offs these ratios can be used to rule in
accurate quantitative measurement of proteinuria. or rule out abnormal 24 protein or albumin loss. If either
Albumin as a percentage of total protein is highly variable ACR or PCR is to be used as a first-line screening test for evi-
at lower levels of proteinuria (Figure 3) due to factors dence of renal disease, then the ratios could clearly be valu-
including a variable contribution from the tubular Tamm able as a rule-out test but early detection of proteinuria in
Horsfall glycoprotein, glomerular-filtered low molecular selected groups of patients may be best achieved using
weight proteins and, in some patients, paraproteins. As ACR. If the tests are to be used for monitoring the level of
urinary total protein levels approach and exceed 1 g/24 h, proteinuria or albuminuria in established renal disease,
the relative contribution from albumin increases and the they can then be used as surrogates for 24 h measurements.
ratio between albumin and total protein becomes relatively The finding that random urine aliquots are equally as accu-
constant. These findings are in agreement with other rate as EMU simplifies sample collection for both patients
published reports.14,48,49 and health-care workers.

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................................................................................................................................................
DECLARATIONS
Competing interests: RN and CP were consultants to Bayer
Corp., now Siemens., Healthcare Diagnostics Ltd, at the
time this project was initiated.
Funding: The study was supported by a grant from Siemens
Healthcare Diagnostics Ltd, USA.
Ethical approval: The study was approved by Greater
Manchester Local Ethics Committees (03/TG/063).
Guarantor: MG.
Contributorship: All authors contributed to the study and
to the writing of the paper.
Acknowledgements: None.
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(Accepted 22 June 2009)

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