Diabetes Research and Clinical Practice, 18 (1992) 83-87 0 1992 Elsevier Science Publishers B.V. All rights reserved 0168-8227/92/$05.00
83
DIABET 00678
Mitral-Test @: a qualitative dipstick test for micro-albuminuria P.F.M.J.
Spooren,
J.F.F.
Lekkerkerker
and I. Vermes
Department of Internal Medicine and Clinical Chemistry. Medisch Spectrum Twente, Enschede, Netherlands (Received 8 January 1992) (Revision accepted 2 May 1992)
Summary Micro-albuminuria is a very sensitive predictor of the development of renal disease in insulin- and non-insulin-dependent diabetes mellitus. A reliable dipstick test for routine screening for microalbuminuria is, therefore, desirable. Such a test has been developed by Boehringer Mannheim, Germany, and marketed as Mitral-Test@. It is an immunological slide-test with semi-quantitative properties. To evaluate its performance as a screening test we compared it with a turbidimetric immuno-assay. In 396 urine specimens from 132 patients, sensitivity was 91 y0 and specificity 96% for a discriminating albumin level of 20 mg/l. Correlation with quantitative values was reasonable (Y= 0.73). We also tested for micro-albuminuria, defined as mean albumin excretion rate of 2 20 ug/min, determined with the turbidimetric immuno-assay in timed overnight urines on three consecutive days, whereas Mitral-Test was considered to be positive for micro-albuminuria if the albumin concentration in one of the three urine samples was 2 20 mg/ml. In 132 patients, the sensitivity of Micra&Test was 82% and the specificity 86Yb. Albumin excretion rate in all false-negative results was < 50 ug/min. We therefore concluded that Mitral-Test is a useful qualitative screening test for micro-albuminuria in diabetic patients. Key words: Diabetic nephropathy;
Dipstick test; Micro-albuminuria
Introduction Micro-albuminuria is defined as a urinary albumin excretion rate (UAER) higher than normal (< 20 ug/min), but lower than the limit detectable by standard laboratory methods (approximately Correspondence to: P.F.M.J. Spooren, Department of Internal Medicine, Medisch Spectrum Twente, P.O. Box 50000,750O KA Enschede, Netherlands.
200 ug/min). This raised, although still low, level of urinary albumin has been shown to be a reliable predictor of the development of diabetic nephropathy in insulin-dependent diabetes mellitus (IDDM) and, to a lesser extent, in non-insulindependent diabetes mellitus (NIDDM) [ 1,2]. There are also indications that micro-albuminuria predicts nephropathy in other renally related disorders and atherosclerotic cardiovascular disease in NIDDM [3,4] and in non-diabetic subjects
84
[ 5,6]. Recent studies in patients suffering from IDDM suggest that aggressive therapy in the micro-albuminuric phase will arrest or even reverse incipient diabetic nephropathy [ 7,8]. Microalbuminuria is therefore an important parameter in the assessment of diabetic (and eventually also non-diabetic) renal and atherosclerotic disease. Recently, Boehringer Mannheim (Germany) introduced Micral-Test@. This test is an easily performed test consisting of a visually read reagent strip. In order to evaluate its performance we compared it with a standard quantitative method.
Materials and Methods In total, 140 patients with IDDM and NIDDM, randomly selected from our outpatient clinic, were asked to collect overnight urine samples on three consecutive days. Commencement and termination times of each sample were recorded on the container. Immediately after collection, the samples were sent to the laboratory where their albumin contents were determined by Mitral-Test and by turbidimetric immuno-assay (TIA). The latter was performed on the end-point determination principle with antisera purchased from Orion, Finland, on a Cobas Fara centrifugal analyser (Hofmann-La Roche, Basel, Switzerland) [ 93. In performing the Mitral-Test determinations the manufacturer’s instructions were followed meticulously. As it is a dipstick test based on immunological reactions, no pretreatment of the samples or additional equipment was necessary. There is a serial arrangement of several reagent pads which are in fluid communication with a reaction-controlling chromatographic process. The Mitral-Test strip is immersed in the urine sample for 5 s with the blue coloured buffer coating just above the surface of the fluid. During this time the sample fluid penetrates the buffer coating and subsequently the conjugate coating, which binds any albumin present in the urine sample to an antibody marked with the enzyme l3-galactosidase. The urine then passes through a capture
matrix, where the excess of marked antibodies is bound to immobilized albumin. Only the sample albumin bound to marked antibodies can reach the substrate film. Here, a colour reaction takes place: the marker enzyme B-galactosidase separates the purple dye chlorophenol red from the yellow substrate (chlorophenol red-galactoside) in a kinetic reaction. The resulting colour is proportional to the albumin concentration in the urine. The colour must be read exactly 5 min after dipping. Mitral-Test yields a range of semiquantitative results which are determined by comparing the strip colour with the five colour blocks on the vial label (Table 1). Both tests were performed by two qualified technicians from our department of Clinical Chemistry. Neither had prior knowledge of the results of the other test. In a preliminary study the results of the two examiners were almost the same for both methods. Therefore, we have neglected these differences in this project. In part A of the study the albumin concentration (mg/l) found in the urine sample with MicralTest was compared with that determined by the reference method. In part B the mean UAER (ug/min) of the three urine samples was determined by Mitral-Test and the reference method and the results compared. For this purpose Mitral-Test was regarded as a qualitative test for the presence or absence of micro-albuminuria. A positive reaction was defined as a mean UAER equal to or exceeding 20 ug/min. In part C the same criterion for a positive re-
TABLt Colours
1 of the dipstick
concentration
test and the corresponding
albumin
(Alb)
Colour
Alb (mg/U
Yellow
0
Light brown
ca. 10
Mid-brown
ca. 20
Brick-red
ca. 50
Burgundy
ca. 100 and more
85 action was applied for the reference method (i.e. mean UAER 2 20 ug/min). Mitral-Test was considered to be positive for micro-albuminuria if the albumin concentration in one of the three urine samples was equal to or greater than 20 mg/l. For statistical calculation linear correlation by the least-squares method was used.
Results Of the initial group of 140 patients, eight had to be excluded, four because of inaccuracies in timing of their collection period and four because of proved clinical albuminuria (defined as a mean UAER 2 200 pg/min, determined by the reference method, and all, incidentally, strongly positive for Mitral-Test). Three hundred and ninety-six urine specimens of 132 patients were, therefore, evalu-
1000
Part A of the study shows an acceptable correlation between the semi-quantitative results of the Mitral-Test and the quantitative results from the turbidimetric immuno-assay (I = 0.73) (Fig. 1). Sensitivity (91%) and specificity (96%) for a discriminating albumin level of 20 mg/l were both good (Table 2,A). In part B of the study the sensitivity of MicralTest for detection of micro-albuminuria (defined as a mean UAER of 2 20 ug/min) was 76 y0 and the specificity 99%. All false-negative results were obtained in patients with an UAER< 50 pg/min, measured by the reference method (Table 2,B). In part C of the study, we tested exclusively at an albumin level of 2 20 mg/l. The sensitivity of Mitral-Test in detecting a mean UAER 2 20 ug/ min increased to 82%) but the specificity decreased to 86% (Table 2,C). As in part B, all false-negative results from the reference method were less than 50 pg/min.
-
. .
. . t
.
i -- ._____^___ !
20-
.:
i
100 -
I
I
L
t
----------_c_-------------------
,
:. i -1 =
10-
: .
5-
1 .
;
l-
I
1
I
I
0
10
20
U-ALBUMIN
Fig. 1. Urinary albumin concentration
I
I
50
100
CONCENTRATION
determined by Mitral-Test
MICRAL-TEST
and by turbidimetric
(
IIlg/l )
immuno-assay
(TIA).
86 TABLE 2 A: albumin level 2 20 mg/l for TIA and Mitral-Test on each urine specimen B: mean UAER2 20 pg/min for TIA and Micra]-Test with triple and timed urine samples C: mean UAER 2 20 pg/min for TIA and one or more urine specimens 2 20 mg/l for Micral-Test
Sample/patient number True-positive True-negative False-positive False-negative Sensitivity f?,) Specificity (Y,)
A
B
C
396 121 247 11 12 91 96
132 25 98
132 21 85 14 6 82 8h
1 8 76 99
Discussion This study has shown an acceptable correlation between the semi-quantitative values of MicralTest and the quantitative values of the turbidimetric immuno-assay. It remains, however, at best a semi-quantitative test with a range of no more than five fixed concentrations. It cannot be regarded as a reliable method for quantitative determinations. Our results were better than those of some other recently published studies [ 10,l11. We attribute this, at least in part, to our meticulously following out manufacturer’s instructions. We found the accuracy of the result to be particularly sensitive to time. We found it necessary to immerse the strip for exactly 5 s, timed with a stopwatch, and to read the result exactly 5 min (with a maximum deviation of no more than 10 s) after dipping. Too long or too short waiting with the dipping and reading procedures means a wrong measurement of albumin concentration in the urine. A simple screening method like a dipstick test solves many of the practical problems facing the laboratory when confronted with large quantities of out-patient urine samples. In part B of the present study we compared the mean UAER obtained with the two methods, accepting the result as positive for micro-albuminuria when UAER
was 220 ug/min. But this is hardly a practical working method for the general practitioner. Unfortunately, the level of albumin in the urine is dependent on the volume and time of collection. In overnight urines the collecting time is mostly about 8 h and the volume of the order of 600 ml. In these cases an albumin level above 20 mg/l, as determined by TIA or an alternative quantitative method, is necessary before MicralTest can detect micro-albuminuria. This endpoint of 20 mg/l is therefore the most logical point for determining micro-albuminuria with MicralTest. At this level the sensitivity of Mitral-Test to a mean UAER of 2 20 ug/min increased to 82%) but the specificity decreased to 86% (Table l,C), which is a rather poor result. Moreover, only about 5096 of cases with a mean UAER range of 20-50 ug/min are detectable with Mitral-Test, so that some micro-albuminuric patients are likely to be missed if this is the only method employed, although patients with UAER’s in the 20-50 ug/ min range are less at risk of developing diabetic nephropathy [ 11. Another well-known rapid qualitative test for micro-albuminuria is Micro-Bumintest@ from Ames Laboratories. The results from this test are comparable to those from Micral-Test [ 10,121, but it suffers the practical disadvantage of a more complicated working method. Mitral-Test, because of its simplicity, is suitable for use in the general practitioner’s practice; the more complex Micro-Bumintest is not. Our conclusion was, therefore, that Mitral-Test is a good method for screening diabetic patients for micro-albuminuria, especially those patients with an UAER 2 50 ug/min. If a patient is positive for Mitral-test, however, further investigation with, for example, turbidimetric immuno-assay will be necessary for the quantitative determination of micro-albuminuria.
Acknowledgements We thank P.S. Page, M.D., for critically reading the manuscript and M. Grob and J.A. Schone-
87 wille for technical assistance. The Mitral-Test dipsticks were provided by Boehringer Mannheim, Germany. References 1 Mogensen, C.E. (1987) Microalbuminuria as a predictor of clinical diabetic nephropathy. Kidney Int. 31, 673-689. 2 Viberti, G.C. and Wiseman, M.J. (1986) The kidney in diabetes: significance of the early abnormalities. Clin. Endocrinol. Metab. 15, 753-782. 3 Jarrett, R.J., Viberti, G.C., Argyropoulos, A., Hill, R.D., Mahmud, U. and Nurrels, T.J. (1984) Microalbuminuria predicts mortality in non-insulin dependent diabetics. Diabetic Med. 1, 17-19. 4 Mogensen, C.E. (1984) Microalbuminuria predicts clinical proteinuria and early mortality in maturity-onset diabetes. N. Engl. J. Med. 310, 356-360. 5 Yudkin, J.S., Forrest, R.D. and Jackson, C.A. (1988) Microalbuminuria as predictor of vascular disease in nondiabetic subjects (Islington Diabetes survey). Lancet ii, 530-533.
6 Cassuto, D., Gibault, T., Altman, J.J. et al. (1991) Microalbuminuria in obese subjects. Int. J. Obesity 15 Suppl. 1, 37. 7 Feldt-Rasmussen, B., Mathiesen, E. and Deckert, T. (1986) Effects of two years of strict metabolic control on the progression of incipient nephropathy in insulindependent diabetes. Lancet ii, 1300-1304. 8 Mogensen, C.E. and Christensen, C.K. (1985) Blood pressure changes and renal function changes in incipient and overt diabetic nephropathy. Hypertension 7(H), 6473. 9 Harmoinen, A., Vuorinen, P. and Jokela, H. (1987) Turbidimetric measurement of microalbuminuria. Clin. Chim. Acta 166, 85-89. 10 Sullivan, K., Raskin, P., Banks, R. et al. (1991) Evaluation of new urine dipstick for the semi-quantitative detection of micro-albuminuria. Diabetes 40 Suppl. 1, 439A. 11 Bangstad, H.-J., Try, K., Dahl-Jorgensen, K. and Hanssen, K.F. (1991) New semiquantitative dipstick test for microalbuminuria. Diabetes Care 14, 1094-1097. 12 Tai, J. and Tze, W.J. (1990) Evaluation of MicroBumintest reagent tablets for screening of microalbuminuria. Diabetes Res. Clin. Pratt. 9, 137-142.
83
DIABET 00678
Mitral-Test @: a qualitative dipstick test for micro-albuminuria P.F.M.J.
Spooren,
J.F.F.
Lekkerkerker
and I. Vermes
Department of Internal Medicine and Clinical Chemistry. Medisch Spectrum Twente, Enschede, Netherlands (Received 8 January 1992) (Revision accepted 2 May 1992)
Summary Micro-albuminuria is a very sensitive predictor of the development of renal disease in insulin- and non-insulin-dependent diabetes mellitus. A reliable dipstick test for routine screening for microalbuminuria is, therefore, desirable. Such a test has been developed by Boehringer Mannheim, Germany, and marketed as Mitral-Test@. It is an immunological slide-test with semi-quantitative properties. To evaluate its performance as a screening test we compared it with a turbidimetric immuno-assay. In 396 urine specimens from 132 patients, sensitivity was 91 y0 and specificity 96% for a discriminating albumin level of 20 mg/l. Correlation with quantitative values was reasonable (Y= 0.73). We also tested for micro-albuminuria, defined as mean albumin excretion rate of 2 20 ug/min, determined with the turbidimetric immuno-assay in timed overnight urines on three consecutive days, whereas Mitral-Test was considered to be positive for micro-albuminuria if the albumin concentration in one of the three urine samples was 2 20 mg/ml. In 132 patients, the sensitivity of Micra&Test was 82% and the specificity 86Yb. Albumin excretion rate in all false-negative results was < 50 ug/min. We therefore concluded that Mitral-Test is a useful qualitative screening test for micro-albuminuria in diabetic patients. Key words: Diabetic nephropathy;
Dipstick test; Micro-albuminuria
Introduction Micro-albuminuria is defined as a urinary albumin excretion rate (UAER) higher than normal (< 20 ug/min), but lower than the limit detectable by standard laboratory methods (approximately Correspondence to: P.F.M.J. Spooren, Department of Internal Medicine, Medisch Spectrum Twente, P.O. Box 50000,750O KA Enschede, Netherlands.
200 ug/min). This raised, although still low, level of urinary albumin has been shown to be a reliable predictor of the development of diabetic nephropathy in insulin-dependent diabetes mellitus (IDDM) and, to a lesser extent, in non-insulindependent diabetes mellitus (NIDDM) [ 1,2]. There are also indications that micro-albuminuria predicts nephropathy in other renally related disorders and atherosclerotic cardiovascular disease in NIDDM [3,4] and in non-diabetic subjects
84
[ 5,6]. Recent studies in patients suffering from IDDM suggest that aggressive therapy in the micro-albuminuric phase will arrest or even reverse incipient diabetic nephropathy [ 7,8]. Microalbuminuria is therefore an important parameter in the assessment of diabetic (and eventually also non-diabetic) renal and atherosclerotic disease. Recently, Boehringer Mannheim (Germany) introduced Micral-Test@. This test is an easily performed test consisting of a visually read reagent strip. In order to evaluate its performance we compared it with a standard quantitative method.
Materials and Methods In total, 140 patients with IDDM and NIDDM, randomly selected from our outpatient clinic, were asked to collect overnight urine samples on three consecutive days. Commencement and termination times of each sample were recorded on the container. Immediately after collection, the samples were sent to the laboratory where their albumin contents were determined by Mitral-Test and by turbidimetric immuno-assay (TIA). The latter was performed on the end-point determination principle with antisera purchased from Orion, Finland, on a Cobas Fara centrifugal analyser (Hofmann-La Roche, Basel, Switzerland) [ 93. In performing the Mitral-Test determinations the manufacturer’s instructions were followed meticulously. As it is a dipstick test based on immunological reactions, no pretreatment of the samples or additional equipment was necessary. There is a serial arrangement of several reagent pads which are in fluid communication with a reaction-controlling chromatographic process. The Mitral-Test strip is immersed in the urine sample for 5 s with the blue coloured buffer coating just above the surface of the fluid. During this time the sample fluid penetrates the buffer coating and subsequently the conjugate coating, which binds any albumin present in the urine sample to an antibody marked with the enzyme l3-galactosidase. The urine then passes through a capture
matrix, where the excess of marked antibodies is bound to immobilized albumin. Only the sample albumin bound to marked antibodies can reach the substrate film. Here, a colour reaction takes place: the marker enzyme B-galactosidase separates the purple dye chlorophenol red from the yellow substrate (chlorophenol red-galactoside) in a kinetic reaction. The resulting colour is proportional to the albumin concentration in the urine. The colour must be read exactly 5 min after dipping. Mitral-Test yields a range of semiquantitative results which are determined by comparing the strip colour with the five colour blocks on the vial label (Table 1). Both tests were performed by two qualified technicians from our department of Clinical Chemistry. Neither had prior knowledge of the results of the other test. In a preliminary study the results of the two examiners were almost the same for both methods. Therefore, we have neglected these differences in this project. In part A of the study the albumin concentration (mg/l) found in the urine sample with MicralTest was compared with that determined by the reference method. In part B the mean UAER (ug/min) of the three urine samples was determined by Mitral-Test and the reference method and the results compared. For this purpose Mitral-Test was regarded as a qualitative test for the presence or absence of micro-albuminuria. A positive reaction was defined as a mean UAER equal to or exceeding 20 ug/min. In part C the same criterion for a positive re-
TABLt Colours
1 of the dipstick
concentration
test and the corresponding
albumin
(Alb)
Colour
Alb (mg/U
Yellow
0
Light brown
ca. 10
Mid-brown
ca. 20
Brick-red
ca. 50
Burgundy
ca. 100 and more
85 action was applied for the reference method (i.e. mean UAER 2 20 ug/min). Mitral-Test was considered to be positive for micro-albuminuria if the albumin concentration in one of the three urine samples was equal to or greater than 20 mg/l. For statistical calculation linear correlation by the least-squares method was used.
Results Of the initial group of 140 patients, eight had to be excluded, four because of inaccuracies in timing of their collection period and four because of proved clinical albuminuria (defined as a mean UAER 2 200 pg/min, determined by the reference method, and all, incidentally, strongly positive for Mitral-Test). Three hundred and ninety-six urine specimens of 132 patients were, therefore, evalu-
1000
Part A of the study shows an acceptable correlation between the semi-quantitative results of the Mitral-Test and the quantitative results from the turbidimetric immuno-assay (I = 0.73) (Fig. 1). Sensitivity (91%) and specificity (96%) for a discriminating albumin level of 20 mg/l were both good (Table 2,A). In part B of the study the sensitivity of MicralTest for detection of micro-albuminuria (defined as a mean UAER of 2 20 ug/min) was 76 y0 and the specificity 99%. All false-negative results were obtained in patients with an UAER< 50 pg/min, measured by the reference method (Table 2,B). In part C of the study, we tested exclusively at an albumin level of 2 20 mg/l. The sensitivity of Mitral-Test in detecting a mean UAER 2 20 ug/ min increased to 82%) but the specificity decreased to 86% (Table 2,C). As in part B, all false-negative results from the reference method were less than 50 pg/min.
-
. .
. . t
.
i -- ._____^___ !
20-
.:
i
100 -
I
I
L
t
----------_c_-------------------
,
:. i -1 =
10-
: .
5-
1 .
;
l-
I
1
I
I
0
10
20
U-ALBUMIN
Fig. 1. Urinary albumin concentration
I
I
50
100
CONCENTRATION
determined by Mitral-Test
MICRAL-TEST
and by turbidimetric
(
IIlg/l )
immuno-assay
(TIA).
86 TABLE 2 A: albumin level 2 20 mg/l for TIA and Mitral-Test on each urine specimen B: mean UAER2 20 pg/min for TIA and Micra]-Test with triple and timed urine samples C: mean UAER 2 20 pg/min for TIA and one or more urine specimens 2 20 mg/l for Micral-Test
Sample/patient number True-positive True-negative False-positive False-negative Sensitivity f?,) Specificity (Y,)
A
B
C
396 121 247 11 12 91 96
132 25 98
132 21 85 14 6 82 8h
1 8 76 99
Discussion This study has shown an acceptable correlation between the semi-quantitative values of MicralTest and the quantitative values of the turbidimetric immuno-assay. It remains, however, at best a semi-quantitative test with a range of no more than five fixed concentrations. It cannot be regarded as a reliable method for quantitative determinations. Our results were better than those of some other recently published studies [ 10,l11. We attribute this, at least in part, to our meticulously following out manufacturer’s instructions. We found the accuracy of the result to be particularly sensitive to time. We found it necessary to immerse the strip for exactly 5 s, timed with a stopwatch, and to read the result exactly 5 min (with a maximum deviation of no more than 10 s) after dipping. Too long or too short waiting with the dipping and reading procedures means a wrong measurement of albumin concentration in the urine. A simple screening method like a dipstick test solves many of the practical problems facing the laboratory when confronted with large quantities of out-patient urine samples. In part B of the present study we compared the mean UAER obtained with the two methods, accepting the result as positive for micro-albuminuria when UAER
was 220 ug/min. But this is hardly a practical working method for the general practitioner. Unfortunately, the level of albumin in the urine is dependent on the volume and time of collection. In overnight urines the collecting time is mostly about 8 h and the volume of the order of 600 ml. In these cases an albumin level above 20 mg/l, as determined by TIA or an alternative quantitative method, is necessary before MicralTest can detect micro-albuminuria. This endpoint of 20 mg/l is therefore the most logical point for determining micro-albuminuria with MicralTest. At this level the sensitivity of Mitral-Test to a mean UAER of 2 20 ug/min increased to 82%) but the specificity decreased to 86% (Table l,C), which is a rather poor result. Moreover, only about 5096 of cases with a mean UAER range of 20-50 ug/min are detectable with Mitral-Test, so that some micro-albuminuric patients are likely to be missed if this is the only method employed, although patients with UAER’s in the 20-50 ug/ min range are less at risk of developing diabetic nephropathy [ 11. Another well-known rapid qualitative test for micro-albuminuria is Micro-Bumintest@ from Ames Laboratories. The results from this test are comparable to those from Micral-Test [ 10,121, but it suffers the practical disadvantage of a more complicated working method. Mitral-Test, because of its simplicity, is suitable for use in the general practitioner’s practice; the more complex Micro-Bumintest is not. Our conclusion was, therefore, that Mitral-Test is a good method for screening diabetic patients for micro-albuminuria, especially those patients with an UAER 2 50 ug/min. If a patient is positive for Mitral-test, however, further investigation with, for example, turbidimetric immuno-assay will be necessary for the quantitative determination of micro-albuminuria.
Acknowledgements We thank P.S. Page, M.D., for critically reading the manuscript and M. Grob and J.A. Schone-
87 wille for technical assistance. The Mitral-Test dipsticks were provided by Boehringer Mannheim, Germany. References 1 Mogensen, C.E. (1987) Microalbuminuria as a predictor of clinical diabetic nephropathy. Kidney Int. 31, 673-689. 2 Viberti, G.C. and Wiseman, M.J. (1986) The kidney in diabetes: significance of the early abnormalities. Clin. Endocrinol. Metab. 15, 753-782. 3 Jarrett, R.J., Viberti, G.C., Argyropoulos, A., Hill, R.D., Mahmud, U. and Nurrels, T.J. (1984) Microalbuminuria predicts mortality in non-insulin dependent diabetics. Diabetic Med. 1, 17-19. 4 Mogensen, C.E. (1984) Microalbuminuria predicts clinical proteinuria and early mortality in maturity-onset diabetes. N. Engl. J. Med. 310, 356-360. 5 Yudkin, J.S., Forrest, R.D. and Jackson, C.A. (1988) Microalbuminuria as predictor of vascular disease in nondiabetic subjects (Islington Diabetes survey). Lancet ii, 530-533.
6 Cassuto, D., Gibault, T., Altman, J.J. et al. (1991) Microalbuminuria in obese subjects. Int. J. Obesity 15 Suppl. 1, 37. 7 Feldt-Rasmussen, B., Mathiesen, E. and Deckert, T. (1986) Effects of two years of strict metabolic control on the progression of incipient nephropathy in insulindependent diabetes. Lancet ii, 1300-1304. 8 Mogensen, C.E. and Christensen, C.K. (1985) Blood pressure changes and renal function changes in incipient and overt diabetic nephropathy. Hypertension 7(H), 6473. 9 Harmoinen, A., Vuorinen, P. and Jokela, H. (1987) Turbidimetric measurement of microalbuminuria. Clin. Chim. Acta 166, 85-89. 10 Sullivan, K., Raskin, P., Banks, R. et al. (1991) Evaluation of new urine dipstick for the semi-quantitative detection of micro-albuminuria. Diabetes 40 Suppl. 1, 439A. 11 Bangstad, H.-J., Try, K., Dahl-Jorgensen, K. and Hanssen, K.F. (1991) New semiquantitative dipstick test for microalbuminuria. Diabetes Care 14, 1094-1097. 12 Tai, J. and Tze, W.J. (1990) Evaluation of MicroBumintest reagent tablets for screening of microalbuminuria. Diabetes Res. Clin. Pratt. 9, 137-142.
