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Performance of a semi-quantitative whole blood test for human heart-type fatty acid-binding protein (H-FABP)
Clinical Biochemistry 38 (2005) 948 – 950
Performance of a semi-quantitative whole blood test for human heart-type fatty acid-binding protein (H-FABP) Masahito Hiura*, Osamu Nakajima, Toshizumi Mori, Katsuya Kitano Department of Cardiology, Hirakata-City Hospital, Kinnya Honmachi 2-14-1, Hirakata/Osaka 573-1013, Japan Received 15 December 2004; received in revised form 30 May 2005; accepted 13 June 2005 Available online 26 July 2005
Abstract Objectives: We evaluated the accuracy of visually reading the whole blood Rapicheck H-FABP panel test using the quantitative plasma H-FABP concentration as the reference. Subjects and methods: Consecutive patients with chest pain (n = 237) who were suspected of having acute myocardial infarction were recruited. The appearance of an evident test line within 5 min was given a grade of +3 (strongly positive), appearance within 15 min +2 (moderately positive) and the appearance of a weak test line within 15 min +1 (weakly positive). Results and discussion: The concordance rates were 91.8% for positive, 70.1% for negative and 80.2% for overall. Plasma H-FABP concentrations were above the cut-off value for 9.2% of negative (0) results. Fifty percent of weakly positive (+1) and 25.0% of moderately positive (+2) results had H-FABP concentrations lower than the cut-off value. All of the strongly positive (+3) were above the cut-off value. These results suggested that the false-positive and false-negative results of Rapicheck H-FABP were caused by over or underestimation in visual reading when the plasma H-FABP concentration was near the cut-off concentration. Conclusions: Low accuracy of visual reading of Rapicheck H-FABP was due to poor estimation by manual reading around the cut-off value. D 2005 The Canadian Society of Clinical Chemists. All rights reserved. Keywords: H-FABP; POCT; Visual reading; Whole blood test; Semi-quantitative
Introduction Human heart-type fatty acid-binding protein (H-FABP) is a biochemical marker for detecting myocardial injury such as acute myocardial infarction (AMI) [1]. Also, blood HFABP concentrations have been utilized for estimation of the infarct size [2– 4], ascertaining the success or failure of reperfusion therapy [5,6] and as an indicator of myocardial injury during open heart surgery [7,8]. Recently, it has been reported that elevated levels of H-FABP were also found in stroke and certain neurogenerative diseases [9,10]. In addition to the conventional sandwich-ELISA assay for the quantification of serum and/or plasma H-FABP concentrations [11], the Rapicheck H-FABP, a qualitative one-step immunochromatography system, was recently introduced for * Corresponding author. Fax: +81 728 47 2825. E-mail address: [email protected] (M. Hiura).
rapid point-of-care testing (POCT) of H-FABP in whole blood [12 – 14]. Because this device relies on visual reading, it may be difficult to interpret results where the test line does not appear clearly. The present study was undertaken to evaluate accuracy of Rapicheck H-FABP whole blood device with visual reading using corresponding quantitative plasma H-FABP concentrations as the reference.
Subjects and methods Subjects The subjects for this study consisted of 237 cases presenting with chest pain between April 2002 and March 2003. There were 145 males (age: 66.6 T 12.1 (mean T standard deviation (SD)) years old) and 92 females (72.2 T 14.1 years old). The subjects consisted of AMI (WHO’s
0009-9120/$ - see front matter D 2005 The Canadian Society of Clinical Chemists. All rights reserved. doi:10.1016/j.clinbiochem.2005.06.007
M. Hiura et al. / Clinical Biochemistry 38 (2005) 948 – 950 Table 1 Coincidence of judgment between visual reading and plasma H-FABP concentration measured by the sandwich-ELISA kit Visual reading, positive Plasma H-FABP 6.2 ng/mL, positive Plasma H-FABP <6.2 ng/mL, negative
Visual reading, negative
91.8% (101/110)
8.2% (9/110)
29.9% (38/127)
70.1% (89/127)
Overall coincidence rate was 80.2%.
criteria) in 29 cases, cardiac disease other than AMI in 138 cases and non-cardiac diseases in 70 cases. Informed consent was obtained from each patient in accordance with the criteria of the hospital’s internal reference board. Methods Grading of color intensity of test line In accordance with the manufacturer’s manual for Rapicheck H-FABP (Dainippon Pharmaceutical Co., Ltd., Osaka, Japan), whole blood samples (150 AL), collected in heparin-treated tubes (Venoject, Terumo, Tokyo, Japan), were dropped onto the designated part of the test plate, which was then left standing at room temperature for 15 min. The appearance of an indicator test line (in addition to the quality control line) within 5 min was graded +3 (strongly positive); appearance of a test line within 15 min was graded +2 (moderately positive); and the appearance of a weak test line within 15 min was graded +1 (weakly positive). The absence of a test line at 15 min was reported as 0 (negative). Plasma H-FABP concentration Simultaneous heparinized blood was centrifuged at 2500 rpm at room temperature for 10 min and used for quantifying H-FABP using an ELISA kit (MARKIT-M HFABP Dainippon Pharmaceutical Co., Ltd., Osaka, Japan) [11] in accordance with the manufacturer’s instructions.
949
of H-FABP concentrations in each category of was investigated, and the correlation between color intensity and plasma H-FABP concentration was analyzed. Statistical analysis McNemar analysis was performed for the evaluation of similarity of the discordant cells of judgment between visual reading and plasma H-FABP concentration. P value less than 0.05 was considered to be statistically significant.
Results Coincidence of H-FABP test results Table 1 summarizes the positive and negative coincidence between visual reading and quantitative H-FABP concentration. The overall coincidence rate based on the positive and negative coincidence rates was 80.2%. The similarity between visual reading and plasma H-FABP concentration in the discordant cells by McNemar analysis was statistically significant ( P < 0.001). Relationship between visual reading of color intensity and H-FABP concentration The mean plasma H-FABP concentration in visually negative cases was 4.0 ng/mL (SD: 1.7). The mean quantitative H-FABP concentration for cases classified as +1 was 7.4 ng/mL (SD: 3.6); cases classified as +2 and +3 were quantitatively 13.2 (SD: 8.8) and 77.6 (114.7) ng/mL, respectively. Table 2 shows that visual reading and quantitative H-FABP concentration was discordant in a 19.8% of cases. A large proportion of discordant cases were in the negative, +1 and +2 classifications.
Discussion Concordance of H-FABP test results Results from visual reading of the Rapicheck H-FABP was compared with the quantitative plasma concentration of H-FABP. Plasma H-FABP concentrations of 6.2 ng/mL were classified as positive [15]. H-FABP concentrations <6.2 ng/mL were classified as negative. The percentage of visually positive cases having positive H-FABP concentrations represented the positive concordance rate; the percentage of visually negative cases with negative H-FABP concentrations was designated as the negative coincidence rate. Other combination were termed discordant.
The study showed that results for the Rapicheck H-FABP were significantly different compared to quantitative plasma H-FABP measurements. Evidently the Rapicheck H-FABP test yielded unclear test lines that are difficult to interpret in cases that were weakly positive (+1 or +2) or negative. The finding that about half of cases classified visually as weakly Table 2 Agreement of visual reading and quantitative H-FABP concentration Visual reading Negative
Relationship between color intensity of test line and plasma H-FABP concentration Color intensity of the test line was analyzed in relation to the corresponding plasma H-FABP concentration. The range
Total number of cases Number of cases 62 ng/mL Number of cases <62 ng/mL Discordant rate (%)
Positive
0
+1
+2
+3
98 9 89 92
70 35 35 50.0
12 9 3 25.0
57 57 0 0
950
M. Hiura et al. / Clinical Biochemistry 38 (2005) 948 – 950
positive had quantitative H-FABP concentrations lower than the cut-off value indicates that visual reading of weakly positive reactions is difficult and accuracy of the test kit is low.
References [1] Tanaka T, Hirota Y, Sohmiya K, Nishimura S, Kawamura K. Serum and urinary human heart fatty acid-binding protein in acute myocardial infarction. Clin Biochem 1991;24:195 – 201. [2] Glatz JFC, Kleine AH, Van Nieuwenhoven FA, Hermens WT, Van Dieijen-Visser MP, Van der Vusse GJ, et al. Fatty-acid-binding protein as a plasma marker for the estimation of myocardial infarct size in humans. Br Heart J 1994;71:135 – 40. [3] Wodig KWH, Kragten JA, Hermens WT, Glatz JFC, Van DieijenVisser MP. Estimation of myocardial infract size from plasma myoglobin or fatty acid-binding protein. Influence of renal function. Eur J Clin Chem Clin Biochem 1997;35:191 – 8. [4] Aartsen WM, Pelsers MMAL, Hermens WT, Glatz JF, Daemen MJ, Smits JF. Heart fatty acid binding protein and cardiac troponin T plasma concentrations as markers for myocardial infarction after coronary artery ligation in mice. Pflugers Archiv Eur J Physiol 2000;439:416 – 22. [5] Knowlton AA, Apstein CS, Saouf R, Brecher P. Leakage of heart fatty acid-binding protein with ischemia and reperfusion in the rat. J Mol Cell Cardiol 1989;21:577 – 83. [6] Ishii J, Nagamura Y, Nomura M, Wang JH, Taga S, Kinoshita M, et al. Early detection of successful coronary reperfusion based on serum concentration of human heart-type cytoplasmic fatty acid-binding protein. Clin Chim Acta 1997;262:13 – 27. [7] Suzuki K, Sawa Y, Kadoba K, Takahashi T, Ichikawa H, Kagisaki K, et al. Early detection of cardiac damage with heart fatty acid-binding protein after cardiac operations. Ann Thorac Surg 1998;65:54 – 8.
[8] Hayashida N, Chihara S, Akasu K, Oda T, Tayama E, Kai E, et al. Plasma and urinary heart fatty acid-binding protein levels in patients undergoing cardiac surgery. Jpn Circ J 2000;64:18 – 22. [9] Steinacker P, Mollenhauer B, Bibl M, Cepek L, Esselmann H, Brechlin P, et al. Heart fatty acid binding protein as a potential diagnostic marker for neurodegenerative diseases. Neurosci Lett 2004;370:36 – 9. [10] Pelsers MMAL, Hanhoff T, Van der Voort D, Arts B, Peters M, Ponds R, et al. Brain- and heart-type fatty acid-binding proteins in the brain: tissue distribution and clinical utility. Clin Chem 2004;50: 1568 – 75. [11] Ohkaru Y, Asayama K, Ishii H, Nishimura S, Sunahara N, Tanaka T, et al. Development of a sandwich enzyme-linked immunosorbent assay for the determination of human heart fatty acid-binding protein in plasma and urine by using two different monoclonal antibodies specific for human heart type fatty acid-binding protein. J Immunol Methods 1995;178:99 – 111. [12] Watanabe T, Ohkubo Y, Matsuoka H, Kimura H, Sakai Y, Ohkaru Y, et al. Development of a simple whole blood panel test for detection of human heart-type fatty acid-binding protein. Clin Biochem 2001;34: 257 – 63. [13] Seino Y, Ogata K, Takano T, Ishii J, Hishida H, Morita H, et al. Use of a whole blood rapid panel test for heart-type fatty acid-binding protein in patients with acute chest pain: comparison with rapid troponin T and myoglobin tests. Am J Med 2003;115:185 – 90. [14] Seino Y, Tomita Y, Takano T, Ohbayashi K. Office cardiologists cooperative study on whole blood rapid panel tests in patients with suspicious acute myocardial infarction. Comparison between hearttype fatty acid-binding protein and troponin t tests. Circ J 2004;68:144 – 8. [15] Okamoto F, Sohmiya K, Ohkaru Y, Kawamura K, Asayama K, Kimura H, et al. Human heart-type cytoplasmic fatty acid-binding protein (H-FABP) for the diagnosis of acute myocardial infarction. Clinical evaluation of H-FABP in comparison with myoglobin and creatine kinase isoenzyme MB. Clin Chem Lab Med 2000;38: 231 – 8.
Performance of a semi-quantitative whole blood test for human heart-type fatty acid-binding protein (H-FABP) Masahito Hiura*, Osamu Nakajima, Toshizumi Mori, Katsuya Kitano Department of Cardiology, Hirakata-City Hospital, Kinnya Honmachi 2-14-1, Hirakata/Osaka 573-1013, Japan Received 15 December 2004; received in revised form 30 May 2005; accepted 13 June 2005 Available online 26 July 2005
Abstract Objectives: We evaluated the accuracy of visually reading the whole blood Rapicheck H-FABP panel test using the quantitative plasma H-FABP concentration as the reference. Subjects and methods: Consecutive patients with chest pain (n = 237) who were suspected of having acute myocardial infarction were recruited. The appearance of an evident test line within 5 min was given a grade of +3 (strongly positive), appearance within 15 min +2 (moderately positive) and the appearance of a weak test line within 15 min +1 (weakly positive). Results and discussion: The concordance rates were 91.8% for positive, 70.1% for negative and 80.2% for overall. Plasma H-FABP concentrations were above the cut-off value for 9.2% of negative (0) results. Fifty percent of weakly positive (+1) and 25.0% of moderately positive (+2) results had H-FABP concentrations lower than the cut-off value. All of the strongly positive (+3) were above the cut-off value. These results suggested that the false-positive and false-negative results of Rapicheck H-FABP were caused by over or underestimation in visual reading when the plasma H-FABP concentration was near the cut-off concentration. Conclusions: Low accuracy of visual reading of Rapicheck H-FABP was due to poor estimation by manual reading around the cut-off value. D 2005 The Canadian Society of Clinical Chemists. All rights reserved. Keywords: H-FABP; POCT; Visual reading; Whole blood test; Semi-quantitative
Introduction Human heart-type fatty acid-binding protein (H-FABP) is a biochemical marker for detecting myocardial injury such as acute myocardial infarction (AMI) [1]. Also, blood HFABP concentrations have been utilized for estimation of the infarct size [2– 4], ascertaining the success or failure of reperfusion therapy [5,6] and as an indicator of myocardial injury during open heart surgery [7,8]. Recently, it has been reported that elevated levels of H-FABP were also found in stroke and certain neurogenerative diseases [9,10]. In addition to the conventional sandwich-ELISA assay for the quantification of serum and/or plasma H-FABP concentrations [11], the Rapicheck H-FABP, a qualitative one-step immunochromatography system, was recently introduced for * Corresponding author. Fax: +81 728 47 2825. E-mail address: [email protected] (M. Hiura).
rapid point-of-care testing (POCT) of H-FABP in whole blood [12 – 14]. Because this device relies on visual reading, it may be difficult to interpret results where the test line does not appear clearly. The present study was undertaken to evaluate accuracy of Rapicheck H-FABP whole blood device with visual reading using corresponding quantitative plasma H-FABP concentrations as the reference.
Subjects and methods Subjects The subjects for this study consisted of 237 cases presenting with chest pain between April 2002 and March 2003. There were 145 males (age: 66.6 T 12.1 (mean T standard deviation (SD)) years old) and 92 females (72.2 T 14.1 years old). The subjects consisted of AMI (WHO’s
0009-9120/$ - see front matter D 2005 The Canadian Society of Clinical Chemists. All rights reserved. doi:10.1016/j.clinbiochem.2005.06.007
M. Hiura et al. / Clinical Biochemistry 38 (2005) 948 – 950 Table 1 Coincidence of judgment between visual reading and plasma H-FABP concentration measured by the sandwich-ELISA kit Visual reading, positive Plasma H-FABP 6.2 ng/mL, positive Plasma H-FABP <6.2 ng/mL, negative
Visual reading, negative
91.8% (101/110)
8.2% (9/110)
29.9% (38/127)
70.1% (89/127)
Overall coincidence rate was 80.2%.
criteria) in 29 cases, cardiac disease other than AMI in 138 cases and non-cardiac diseases in 70 cases. Informed consent was obtained from each patient in accordance with the criteria of the hospital’s internal reference board. Methods Grading of color intensity of test line In accordance with the manufacturer’s manual for Rapicheck H-FABP (Dainippon Pharmaceutical Co., Ltd., Osaka, Japan), whole blood samples (150 AL), collected in heparin-treated tubes (Venoject, Terumo, Tokyo, Japan), were dropped onto the designated part of the test plate, which was then left standing at room temperature for 15 min. The appearance of an indicator test line (in addition to the quality control line) within 5 min was graded +3 (strongly positive); appearance of a test line within 15 min was graded +2 (moderately positive); and the appearance of a weak test line within 15 min was graded +1 (weakly positive). The absence of a test line at 15 min was reported as 0 (negative). Plasma H-FABP concentration Simultaneous heparinized blood was centrifuged at 2500 rpm at room temperature for 10 min and used for quantifying H-FABP using an ELISA kit (MARKIT-M HFABP Dainippon Pharmaceutical Co., Ltd., Osaka, Japan) [11] in accordance with the manufacturer’s instructions.
949
of H-FABP concentrations in each category of was investigated, and the correlation between color intensity and plasma H-FABP concentration was analyzed. Statistical analysis McNemar analysis was performed for the evaluation of similarity of the discordant cells of judgment between visual reading and plasma H-FABP concentration. P value less than 0.05 was considered to be statistically significant.
Results Coincidence of H-FABP test results Table 1 summarizes the positive and negative coincidence between visual reading and quantitative H-FABP concentration. The overall coincidence rate based on the positive and negative coincidence rates was 80.2%. The similarity between visual reading and plasma H-FABP concentration in the discordant cells by McNemar analysis was statistically significant ( P < 0.001). Relationship between visual reading of color intensity and H-FABP concentration The mean plasma H-FABP concentration in visually negative cases was 4.0 ng/mL (SD: 1.7). The mean quantitative H-FABP concentration for cases classified as +1 was 7.4 ng/mL (SD: 3.6); cases classified as +2 and +3 were quantitatively 13.2 (SD: 8.8) and 77.6 (114.7) ng/mL, respectively. Table 2 shows that visual reading and quantitative H-FABP concentration was discordant in a 19.8% of cases. A large proportion of discordant cases were in the negative, +1 and +2 classifications.
Discussion Concordance of H-FABP test results Results from visual reading of the Rapicheck H-FABP was compared with the quantitative plasma concentration of H-FABP. Plasma H-FABP concentrations of 6.2 ng/mL were classified as positive [15]. H-FABP concentrations <6.2 ng/mL were classified as negative. The percentage of visually positive cases having positive H-FABP concentrations represented the positive concordance rate; the percentage of visually negative cases with negative H-FABP concentrations was designated as the negative coincidence rate. Other combination were termed discordant.
The study showed that results for the Rapicheck H-FABP were significantly different compared to quantitative plasma H-FABP measurements. Evidently the Rapicheck H-FABP test yielded unclear test lines that are difficult to interpret in cases that were weakly positive (+1 or +2) or negative. The finding that about half of cases classified visually as weakly Table 2 Agreement of visual reading and quantitative H-FABP concentration Visual reading Negative
Relationship between color intensity of test line and plasma H-FABP concentration Color intensity of the test line was analyzed in relation to the corresponding plasma H-FABP concentration. The range
Total number of cases Number of cases 62 ng/mL Number of cases <62 ng/mL Discordant rate (%)
Positive
0
+1
+2
+3
98 9 89 92
70 35 35 50.0
12 9 3 25.0
57 57 0 0
950
M. Hiura et al. / Clinical Biochemistry 38 (2005) 948 – 950
positive had quantitative H-FABP concentrations lower than the cut-off value indicates that visual reading of weakly positive reactions is difficult and accuracy of the test kit is low.
References [1] Tanaka T, Hirota Y, Sohmiya K, Nishimura S, Kawamura K. Serum and urinary human heart fatty acid-binding protein in acute myocardial infarction. Clin Biochem 1991;24:195 – 201. [2] Glatz JFC, Kleine AH, Van Nieuwenhoven FA, Hermens WT, Van Dieijen-Visser MP, Van der Vusse GJ, et al. Fatty-acid-binding protein as a plasma marker for the estimation of myocardial infarct size in humans. Br Heart J 1994;71:135 – 40. [3] Wodig KWH, Kragten JA, Hermens WT, Glatz JFC, Van DieijenVisser MP. Estimation of myocardial infract size from plasma myoglobin or fatty acid-binding protein. Influence of renal function. Eur J Clin Chem Clin Biochem 1997;35:191 – 8. [4] Aartsen WM, Pelsers MMAL, Hermens WT, Glatz JF, Daemen MJ, Smits JF. Heart fatty acid binding protein and cardiac troponin T plasma concentrations as markers for myocardial infarction after coronary artery ligation in mice. Pflugers Archiv Eur J Physiol 2000;439:416 – 22. [5] Knowlton AA, Apstein CS, Saouf R, Brecher P. Leakage of heart fatty acid-binding protein with ischemia and reperfusion in the rat. J Mol Cell Cardiol 1989;21:577 – 83. [6] Ishii J, Nagamura Y, Nomura M, Wang JH, Taga S, Kinoshita M, et al. Early detection of successful coronary reperfusion based on serum concentration of human heart-type cytoplasmic fatty acid-binding protein. Clin Chim Acta 1997;262:13 – 27. [7] Suzuki K, Sawa Y, Kadoba K, Takahashi T, Ichikawa H, Kagisaki K, et al. Early detection of cardiac damage with heart fatty acid-binding protein after cardiac operations. Ann Thorac Surg 1998;65:54 – 8.
[8] Hayashida N, Chihara S, Akasu K, Oda T, Tayama E, Kai E, et al. Plasma and urinary heart fatty acid-binding protein levels in patients undergoing cardiac surgery. Jpn Circ J 2000;64:18 – 22. [9] Steinacker P, Mollenhauer B, Bibl M, Cepek L, Esselmann H, Brechlin P, et al. Heart fatty acid binding protein as a potential diagnostic marker for neurodegenerative diseases. Neurosci Lett 2004;370:36 – 9. [10] Pelsers MMAL, Hanhoff T, Van der Voort D, Arts B, Peters M, Ponds R, et al. Brain- and heart-type fatty acid-binding proteins in the brain: tissue distribution and clinical utility. Clin Chem 2004;50: 1568 – 75. [11] Ohkaru Y, Asayama K, Ishii H, Nishimura S, Sunahara N, Tanaka T, et al. Development of a sandwich enzyme-linked immunosorbent assay for the determination of human heart fatty acid-binding protein in plasma and urine by using two different monoclonal antibodies specific for human heart type fatty acid-binding protein. J Immunol Methods 1995;178:99 – 111. [12] Watanabe T, Ohkubo Y, Matsuoka H, Kimura H, Sakai Y, Ohkaru Y, et al. Development of a simple whole blood panel test for detection of human heart-type fatty acid-binding protein. Clin Biochem 2001;34: 257 – 63. [13] Seino Y, Ogata K, Takano T, Ishii J, Hishida H, Morita H, et al. Use of a whole blood rapid panel test for heart-type fatty acid-binding protein in patients with acute chest pain: comparison with rapid troponin T and myoglobin tests. Am J Med 2003;115:185 – 90. [14] Seino Y, Tomita Y, Takano T, Ohbayashi K. Office cardiologists cooperative study on whole blood rapid panel tests in patients with suspicious acute myocardial infarction. Comparison between hearttype fatty acid-binding protein and troponin t tests. Circ J 2004;68:144 – 8. [15] Okamoto F, Sohmiya K, Ohkaru Y, Kawamura K, Asayama K, Kimura H, et al. Human heart-type cytoplasmic fatty acid-binding protein (H-FABP) for the diagnosis of acute myocardial infarction. Clinical evaluation of H-FABP in comparison with myoglobin and creatine kinase isoenzyme MB. Clin Chem Lab Med 2000;38: 231 – 8.