Scandinavian Journal of Clinical & Laboratory Investigation, 2014; 74: 524–526
ORIGINAL ARTICLE
Diagnosis of late presenting subarachnoid hemorrhage: Comparison of methods for cerebrospinal fluid ferritin Astrid Petersmann1, Anders Kallner2, Helma Preez1, Elisabeth Thein3 & Alexander Dressel3 1Institute
of Clinical Chemistry and Laboratory Medicine, University Medicine Greifswald, Greifswald, Germany, of Clinical Chemistry, Karolinska University Hospital, Stockholm, Sweden, and 3Department of Neurology, University Medicine Greifswald, Greifswald, Germany
2Department
Abstract Background. The majority of subarachnoid hemorrhage (SAH) is diagnosed using imaging techniques. The sensitivity of computed tomography scans decreases with increasing time after the bleeding event which can lead to false negative CT scans. Spectrophotometry and microscopic investigations of the cerebrospinal fluid (Csf) can provide additional diagnostic support, but may not be available for emergency diagnoses. Csf-Ferritin has been suggested as an alternative additional marker for SAH that present late and has a potency to be measured in a routine laboratory. Methods. A routine Ferritin chemiluminescent assay (Dimension Vista) was compared with a branded and CE-marked Csf-Ferritin nephelometric assay (BN ProSpec) using surplus routine patient samples. We calculated imprecision at pertinent concentrations, compared patient samples, and established reference intervals. Results. The standard deviation was about a third for the Dimension Vista assay compared to that of the BN ProSpec assay at the three tested concentrations. The correlation showed a systematic difference between the methods but the correlation was high (r 0.955). Accordingly, the reference intervals were higher for the BN ProSPec (2.7–16.8 mg/L) than for the Dimension Vista (2.0–12.6 mg/L). Conclusion. The precision of the Dimension Vista measurements was considerably better than that of the BN ProSpec. The Dimension Vista results correlated well with those of the comparative method, yielding slightly lower values. This is reflected in the reference intervals. These findings permit the use of the routinely available Ferritin assay of the Dimension Vista for measuring Csf-Ferritin and complementing the late diagnosis of SAH outside office hours of specialized Csf laboratories. Key Words: Minimal difference, routine emergency laboratory, Csf-Ferritin reference intervals
Introduction The majority of subarachnoid hemorrhage (SAH) and other intracranial bleedings are diagnosed using imaging techniques. Sensitivity of computed tomography (CT) scans decreases with time after the bleeding event, which can lead to false negative CT scans [1]. Spectrophotometry [2] and microscopic investigations of the cerebrospinal fluid (Csf) can provide additional diagnostic support [3,4]. However, both analytical procedures require experienced and specially trained technicians who may not be available for emergency diagnostics. Csf-Ferritin has been suggested as an additional marker for SAH patients who present at a time [5,6] when the CT is normalized and CSF bilirubin is
undetectable [7,8]. In comparison to the other methods Csf-Ferritin offers an objective measure that can easily be obtained in a clinical routine laboratory. We compared a method used for Ferritin, available on a large routine platform often used in routine emergency laboratories, to a validated and CE marked Csf-Ferritin method often used in specialized Csf laboratories with limited opening hours. The study design allows calculating the imprecision of the methods and reference intervals. Materials and methods Measurements of Csf-Ferritin concentration were performed with a nephelometric assay, on the BN
Correspondence: Astrid Petersmann, Institute of Clinical Chemistry and Laboratory Medicine, University Medicine of Greifswald, Ferdinand-SauerbruchStraße, 17475 Greifswald, Germany. Tel: 49 3834 86 5670. E-mail: [email protected] (Received 2 February 2014; accepted 30 March 2014) ISSN 0036-5513 print/ISSN 1502-7686 online © 2014 Informa Healthcare DOI: 10.3109/00365513.2014.913187
ProSpec (Siemens Healthcare Diagnostics, Eschborn, Germany) and chemiluminescence assay on the Dimension Vista. A routine Ferritin chemiluminescent assay (Dimension Vista, using the CE-marked urine ferritin concentration measurement calibration protocol) was compared with a branded and CE-marked CsfFerritin nephelometric assay (BN ProSpec). Both instruments were manufactured by Siemens Healthcare Diagnostics, Eschborn, Germany. Surplus routine patient samples were used throughout the study. The Ethics’ Committee of Greifswald approved the use of surplus material for evaluation and quality assurance purposes (III UV 39\03). We calculated imprecision at pertinent concentrations, compared patient samples, and established reference intervals. Three pools (pool 1, 2 and 3) of patient cerebrospinal fluid were made with approximate concentrations 11 mg/L, 17 mg/L and 20 mg/L (BN ProSpec) chosen to cover the diagnostically interesting interval. The pools were aliquoted and stored at 20oC until measurement. Each aliquot was thawed, mixed and centrifuged before measurement. Imprecision was estimated by measuring the pooled samples twice a day for five consecutive days. The imprecision was calculated from the total variance of these measurements. The imprecision of a measurement may be easier to use in a clinical context if it is expressed as a minimal difference (MD). The MD describes the smallest significant analytical difference between two adjacent measurements [9] and was calculated as MD k 2 SD2 k SD 2 . The variable k represents the coverage factor; k 2 corresponding to a level of confidence of 95%. Regression analysis was made by the PassingBablok approach and correlation was expressed as the Pearson correlation coefficient using 90 consecutive patient samples. Reference intervals were established as the central 95 percentile of the results from 100 patients. The inclusion criteria for samples were an intact bloodbrain barrier, no signs of bleeding or inflammation (defined by erythrocytes: 5 106/L, leucocytes: 5 106/L, lactate concentration: 2.5 mmol/L). These samples and results were not used in the method comparison. Statistical analysis was performed with Microsoft Excel 2010 and Analyse-it, v3.60 (Analyse-it Software, Ltd. Leeds, UK).
Method comparison of Csf-Ferritin 525 Table I. Imprecision data for Csf-Ferritin on BN ProSpec and Dimension Vista. The number of observations per pool was ten. The coverage factor for calculating the MD was k 2.
Pool 1 Mean mg/L SD mg/L %CV MD mg/L Pool 2 Mean mg/L SD mg/L %CV MD mg/L Pool 3 Mean mg/L SD mg/L %CV MD mg/L
BN
Vista
11.1 1.0 9.4 2.8
8.4 0.2 2.7 0.6
17.1 1.2 7.1 3.4
13.1 0.4 2.8 1.1
20.3 1.6 8.0 4.5
16.1 0.5 3.0 1.4
studied. This included 49 patients and was chosen to represent the clinically interesting interval. PassingBablok regression functions were calculated for the complete sample set (Y 0.928X 1.171; r 0.998) and for the subset (Y 0.805X 0.451; r 0.955), the latter displayed in Figure 1.
Results Imprecision (standard deviation) and the MD of the pools are given in Table I. The method comparison comprised 90 consecutive patient samples for which ferritin concentrations ranged from 2.5–4020 mg/L. A subset of samples with a ferritin concentration of up to 20 mg/L was
Figure 1. Regression (Passing-Bablok) and difference graph of the subset of results 20 mg/L. N 49. The dotted line is the equal line (Y X).
526 A. Petersmann et al. The reference intervals (central 95 percentile) were: 2.7–16.8 mg/L (BN ProSpec) and 2.0– 12.6 mg/L (Dimension Vista) based on 100 patients.
diagnosis of SAH outside office hours of specialized Csf laboratories. Acknowledgements
Discussion There are several different established markers to aid diagnosis of SAH, e.g. CT, Csf-cytology, Csf spectrometry and the ferritin concentration of the Csf. The markers are of importance at different times during the clinical diagnostic process. The Csf-Ferritin has been suggested as an alternative marker for SAH presenting late [5,7,8]. We validated a Ferritin assay applicable in a routine emergency laboratory by comparing it to a CEmarked Csf-Ferritin measurement method which often maybe performed only in specialized Csf laboratories with limited service hours. The precision of the Dimension Vista is superior to the BN ProSpec as demonstrated in Table I. Expressed as MD, two ferritin concentrations from the BN ProSpec can be considered different if differing more than 3.4 mg/L (pool 2; Table I), whereas the MD for the Dimension Vista is 1.1 mg/L. The Pearson correlation coefficient was high (0.998) for the entire data set and the subset (0.955) characterized by Csf-Ferritin 20 mg/L (Figure 1). The Dimension Vista gave results which were about 20% lower than those of the BN ProSpec. A similar difference was observed in the pooled samples. Accordingly, the reference intervals for the Dimension Vista are lower.
Conclusion The precision of the Dimension Vista measurements was considerably better than that of the BN ProSpec. The Dimension Vista results correlated well with those of the comparative method, yielding slightly lower values. This is reflected in the reference intervals. These findings permit the use of the routinely available Ferritin assay of the Dimension Vista for measuring Csf-Ferritin and complementing the late
None Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper. References [1] Edlow JA, Caplan LR. Avoiding pitfalls in the diagnosis of subarachnoid hemorrhage. N Eng J Med 2000;342:29–36. [2] Wahlgren NG, Bergstrom K. Determination of haem derivatives in the cerebrospinal fluid – a semi-quantitative method. J Neurol Neurosurg Psychiatry 1983;46:653–8. [3] Petzold A, Sharpe LT, Keir G. Spectrophotometry for cerebrospinal fluid pigment analysis. Neurocrit Care 2006;4:153–62. [4] Cruickshank A, Auld P, Beetham R, Burrows G, Egner W, Holbrook I, Keir G, Lewis E, Patel D, Watson I, White P. Revised national guidelines for analysis of cerebrospinal fluid for bilirubin in suspected subarachnoid haemorrhage. Ann Clin Biochem 2008;45:238–44. [5] Petzold A, Worthington V, Appleby I, Kerr M, Kitchen N, Smith M. Cerebrospinal fluid ferritin levels, a sensitive diagnostic test in delayed presenting subarachnoid haemorrhage. J Stroke Cerebrovasc Dis 2011;20:489–93. [6] Tumani H, Petzold A, Wick M, Kühn HJ, Uhr M, Otto M, Regeniter A, Brettschneider J. [Cerebrospinal fluid-based diagnostics of CT-negative subarachnoid haemorrhage]. [Article in German] Nervenarzt 2010;81:973–9. [7] Petzold A, Worthington V, Pritchard C, Appleby I, Kitchen N, Smith M. The longitudinal profile of bilirubin and ferritin in the cerebrospinal fluid following a subarachnoid hemorrhage: diagnostic implications. Neurocrit Care 2009;11:398–402. [8] Nagy K, Skagervik I, Tumani H, Petzold A, Wick M, Kühn H-J, Uhr M, Regeniter A, Brettschneider J, Otto M, Kraus J, Deisenhammer F, Lautner R, Blennow K, Shaw L, Zetterberg H, Mattsson N. Cerebrospinal fluid analyses for the diagnosis of subarachnoid haemorrhage and experience from a Swedish study. What method is preferable when diagnosing a subarachnoid haemorrhage? Clin Chem Lab Med 2013;51:2073–86. [9] Clinical and Laboratory Standards Institute (CLSI). Expression of Measurement Uncertainty in Laboratory Medicine; Approved Guideline. CLSI document EP39 (ISBN 1-56238-000-0). Wayne, PA: Clinical and Laboratory Standards Institute, 2012.
Copyright of Scandinavian Journal of Clinical & Laboratory Investigation is the property of Taylor & Francis Ltd and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
ORIGINAL ARTICLE
Diagnosis of late presenting subarachnoid hemorrhage: Comparison of methods for cerebrospinal fluid ferritin Astrid Petersmann1, Anders Kallner2, Helma Preez1, Elisabeth Thein3 & Alexander Dressel3 1Institute
of Clinical Chemistry and Laboratory Medicine, University Medicine Greifswald, Greifswald, Germany, of Clinical Chemistry, Karolinska University Hospital, Stockholm, Sweden, and 3Department of Neurology, University Medicine Greifswald, Greifswald, Germany
2Department
Abstract Background. The majority of subarachnoid hemorrhage (SAH) is diagnosed using imaging techniques. The sensitivity of computed tomography scans decreases with increasing time after the bleeding event which can lead to false negative CT scans. Spectrophotometry and microscopic investigations of the cerebrospinal fluid (Csf) can provide additional diagnostic support, but may not be available for emergency diagnoses. Csf-Ferritin has been suggested as an alternative additional marker for SAH that present late and has a potency to be measured in a routine laboratory. Methods. A routine Ferritin chemiluminescent assay (Dimension Vista) was compared with a branded and CE-marked Csf-Ferritin nephelometric assay (BN ProSpec) using surplus routine patient samples. We calculated imprecision at pertinent concentrations, compared patient samples, and established reference intervals. Results. The standard deviation was about a third for the Dimension Vista assay compared to that of the BN ProSpec assay at the three tested concentrations. The correlation showed a systematic difference between the methods but the correlation was high (r 0.955). Accordingly, the reference intervals were higher for the BN ProSPec (2.7–16.8 mg/L) than for the Dimension Vista (2.0–12.6 mg/L). Conclusion. The precision of the Dimension Vista measurements was considerably better than that of the BN ProSpec. The Dimension Vista results correlated well with those of the comparative method, yielding slightly lower values. This is reflected in the reference intervals. These findings permit the use of the routinely available Ferritin assay of the Dimension Vista for measuring Csf-Ferritin and complementing the late diagnosis of SAH outside office hours of specialized Csf laboratories. Key Words: Minimal difference, routine emergency laboratory, Csf-Ferritin reference intervals
Introduction The majority of subarachnoid hemorrhage (SAH) and other intracranial bleedings are diagnosed using imaging techniques. Sensitivity of computed tomography (CT) scans decreases with time after the bleeding event, which can lead to false negative CT scans [1]. Spectrophotometry [2] and microscopic investigations of the cerebrospinal fluid (Csf) can provide additional diagnostic support [3,4]. However, both analytical procedures require experienced and specially trained technicians who may not be available for emergency diagnostics. Csf-Ferritin has been suggested as an additional marker for SAH patients who present at a time [5,6] when the CT is normalized and CSF bilirubin is
undetectable [7,8]. In comparison to the other methods Csf-Ferritin offers an objective measure that can easily be obtained in a clinical routine laboratory. We compared a method used for Ferritin, available on a large routine platform often used in routine emergency laboratories, to a validated and CE marked Csf-Ferritin method often used in specialized Csf laboratories with limited opening hours. The study design allows calculating the imprecision of the methods and reference intervals. Materials and methods Measurements of Csf-Ferritin concentration were performed with a nephelometric assay, on the BN
Correspondence: Astrid Petersmann, Institute of Clinical Chemistry and Laboratory Medicine, University Medicine of Greifswald, Ferdinand-SauerbruchStraße, 17475 Greifswald, Germany. Tel: 49 3834 86 5670. E-mail: [email protected] (Received 2 February 2014; accepted 30 March 2014) ISSN 0036-5513 print/ISSN 1502-7686 online © 2014 Informa Healthcare DOI: 10.3109/00365513.2014.913187
ProSpec (Siemens Healthcare Diagnostics, Eschborn, Germany) and chemiluminescence assay on the Dimension Vista. A routine Ferritin chemiluminescent assay (Dimension Vista, using the CE-marked urine ferritin concentration measurement calibration protocol) was compared with a branded and CE-marked CsfFerritin nephelometric assay (BN ProSpec). Both instruments were manufactured by Siemens Healthcare Diagnostics, Eschborn, Germany. Surplus routine patient samples were used throughout the study. The Ethics’ Committee of Greifswald approved the use of surplus material for evaluation and quality assurance purposes (III UV 39\03). We calculated imprecision at pertinent concentrations, compared patient samples, and established reference intervals. Three pools (pool 1, 2 and 3) of patient cerebrospinal fluid were made with approximate concentrations 11 mg/L, 17 mg/L and 20 mg/L (BN ProSpec) chosen to cover the diagnostically interesting interval. The pools were aliquoted and stored at 20oC until measurement. Each aliquot was thawed, mixed and centrifuged before measurement. Imprecision was estimated by measuring the pooled samples twice a day for five consecutive days. The imprecision was calculated from the total variance of these measurements. The imprecision of a measurement may be easier to use in a clinical context if it is expressed as a minimal difference (MD). The MD describes the smallest significant analytical difference between two adjacent measurements [9] and was calculated as MD k 2 SD2 k SD 2 . The variable k represents the coverage factor; k 2 corresponding to a level of confidence of 95%. Regression analysis was made by the PassingBablok approach and correlation was expressed as the Pearson correlation coefficient using 90 consecutive patient samples. Reference intervals were established as the central 95 percentile of the results from 100 patients. The inclusion criteria for samples were an intact bloodbrain barrier, no signs of bleeding or inflammation (defined by erythrocytes: 5 106/L, leucocytes: 5 106/L, lactate concentration: 2.5 mmol/L). These samples and results were not used in the method comparison. Statistical analysis was performed with Microsoft Excel 2010 and Analyse-it, v3.60 (Analyse-it Software, Ltd. Leeds, UK).
Method comparison of Csf-Ferritin 525 Table I. Imprecision data for Csf-Ferritin on BN ProSpec and Dimension Vista. The number of observations per pool was ten. The coverage factor for calculating the MD was k 2.
Pool 1 Mean mg/L SD mg/L %CV MD mg/L Pool 2 Mean mg/L SD mg/L %CV MD mg/L Pool 3 Mean mg/L SD mg/L %CV MD mg/L
BN
Vista
11.1 1.0 9.4 2.8
8.4 0.2 2.7 0.6
17.1 1.2 7.1 3.4
13.1 0.4 2.8 1.1
20.3 1.6 8.0 4.5
16.1 0.5 3.0 1.4
studied. This included 49 patients and was chosen to represent the clinically interesting interval. PassingBablok regression functions were calculated for the complete sample set (Y 0.928X 1.171; r 0.998) and for the subset (Y 0.805X 0.451; r 0.955), the latter displayed in Figure 1.
Results Imprecision (standard deviation) and the MD of the pools are given in Table I. The method comparison comprised 90 consecutive patient samples for which ferritin concentrations ranged from 2.5–4020 mg/L. A subset of samples with a ferritin concentration of up to 20 mg/L was
Figure 1. Regression (Passing-Bablok) and difference graph of the subset of results 20 mg/L. N 49. The dotted line is the equal line (Y X).
526 A. Petersmann et al. The reference intervals (central 95 percentile) were: 2.7–16.8 mg/L (BN ProSpec) and 2.0– 12.6 mg/L (Dimension Vista) based on 100 patients.
diagnosis of SAH outside office hours of specialized Csf laboratories. Acknowledgements
Discussion There are several different established markers to aid diagnosis of SAH, e.g. CT, Csf-cytology, Csf spectrometry and the ferritin concentration of the Csf. The markers are of importance at different times during the clinical diagnostic process. The Csf-Ferritin has been suggested as an alternative marker for SAH presenting late [5,7,8]. We validated a Ferritin assay applicable in a routine emergency laboratory by comparing it to a CEmarked Csf-Ferritin measurement method which often maybe performed only in specialized Csf laboratories with limited service hours. The precision of the Dimension Vista is superior to the BN ProSpec as demonstrated in Table I. Expressed as MD, two ferritin concentrations from the BN ProSpec can be considered different if differing more than 3.4 mg/L (pool 2; Table I), whereas the MD for the Dimension Vista is 1.1 mg/L. The Pearson correlation coefficient was high (0.998) for the entire data set and the subset (0.955) characterized by Csf-Ferritin 20 mg/L (Figure 1). The Dimension Vista gave results which were about 20% lower than those of the BN ProSpec. A similar difference was observed in the pooled samples. Accordingly, the reference intervals for the Dimension Vista are lower.
Conclusion The precision of the Dimension Vista measurements was considerably better than that of the BN ProSpec. The Dimension Vista results correlated well with those of the comparative method, yielding slightly lower values. This is reflected in the reference intervals. These findings permit the use of the routinely available Ferritin assay of the Dimension Vista for measuring Csf-Ferritin and complementing the late
None Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper. References [1] Edlow JA, Caplan LR. Avoiding pitfalls in the diagnosis of subarachnoid hemorrhage. N Eng J Med 2000;342:29–36. [2] Wahlgren NG, Bergstrom K. Determination of haem derivatives in the cerebrospinal fluid – a semi-quantitative method. J Neurol Neurosurg Psychiatry 1983;46:653–8. [3] Petzold A, Sharpe LT, Keir G. Spectrophotometry for cerebrospinal fluid pigment analysis. Neurocrit Care 2006;4:153–62. [4] Cruickshank A, Auld P, Beetham R, Burrows G, Egner W, Holbrook I, Keir G, Lewis E, Patel D, Watson I, White P. Revised national guidelines for analysis of cerebrospinal fluid for bilirubin in suspected subarachnoid haemorrhage. Ann Clin Biochem 2008;45:238–44. [5] Petzold A, Worthington V, Appleby I, Kerr M, Kitchen N, Smith M. Cerebrospinal fluid ferritin levels, a sensitive diagnostic test in delayed presenting subarachnoid haemorrhage. J Stroke Cerebrovasc Dis 2011;20:489–93. [6] Tumani H, Petzold A, Wick M, Kühn HJ, Uhr M, Otto M, Regeniter A, Brettschneider J. [Cerebrospinal fluid-based diagnostics of CT-negative subarachnoid haemorrhage]. [Article in German] Nervenarzt 2010;81:973–9. [7] Petzold A, Worthington V, Pritchard C, Appleby I, Kitchen N, Smith M. The longitudinal profile of bilirubin and ferritin in the cerebrospinal fluid following a subarachnoid hemorrhage: diagnostic implications. Neurocrit Care 2009;11:398–402. [8] Nagy K, Skagervik I, Tumani H, Petzold A, Wick M, Kühn H-J, Uhr M, Regeniter A, Brettschneider J, Otto M, Kraus J, Deisenhammer F, Lautner R, Blennow K, Shaw L, Zetterberg H, Mattsson N. Cerebrospinal fluid analyses for the diagnosis of subarachnoid haemorrhage and experience from a Swedish study. What method is preferable when diagnosing a subarachnoid haemorrhage? Clin Chem Lab Med 2013;51:2073–86. [9] Clinical and Laboratory Standards Institute (CLSI). Expression of Measurement Uncertainty in Laboratory Medicine; Approved Guideline. CLSI document EP39 (ISBN 1-56238-000-0). Wayne, PA: Clinical and Laboratory Standards Institute, 2012.
Copyright of Scandinavian Journal of Clinical & Laboratory Investigation is the property of Taylor & Francis Ltd and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
