Liquid chromatography-tandem mass spectrometry analysis of 17-hydroxyprogesterone in dried blood spots revealed matrix effect on immunoassay
Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS...
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| Published in | Analytical and bioanalytical chemistry Vol. 411; no. 2; pp. 395 - 402 |
|---|---|
| Main Authors | , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Berlin/Heidelberg
Springer Berlin Heidelberg
01.01.2019
Springer Springer Nature B.V |
| Subjects | |
| Online Access | Get full text |
| ISSN | 1618-2642 1618-2650 1618-2650 |
| DOI | 10.1007/s00216-018-1449-0 |
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| Abstract | Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with
R
2
= 0.9994 for 17-OHP and
R
2
= 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (
R
2
= 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (
R
2
= 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. |
|---|---|
| AbstractList | Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) [less than or equal to] 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R.sup.2 = 0.9994 for 17-OHP and R.sup.2 = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R.sup.2 = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R.sup.2 = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R2 = 0.9994 for 17-OHP and R2 = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R2 = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R2 = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R 2 = 0.9994 for 17-OHP and R 2 = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples ( R 2 = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples ( R 2 = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R² = 0.9994 for 17-OHP and R² = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R² = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R² = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R = 0.9994 for 17-OHP and R = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R2 = 0.9994 for 17-OHP and R2 = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R2 = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R2 = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays.Immunoassays for measuring 17-hydroxyprogesterone (17-OHP) produce high rates of false positives that impact the identification of congenital adrenal hyperplasia (CAH) in neonates. A confirmatory test with high analytical specificity employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology is needed in newborn screening for CAH. 17-OHP and cortisol were extracted from dried blood spot (DBS) samples, resolved on a C18 column, and measured using tandem mass spectrometry. The results were compared with those determined using the AutoDELFIA immunoassay. The LC-MS/MS method had a limit of quantitation of 10.0 and 5.0 ng/mL for 17-OHP and cortisol, respectively. The method characteristics showed coefficient variation (%CV) ≤ 11.9% for both 17-OHP and cortisol, recoveries ranging from 83.1 to 101.5% for 17-OHP and from 95.1 to 102.8% for cortisol, and linearity with R2 = 0.9994 for 17-OHP and R2 = 0.9996 for cortisol, clinical sensitivity of 100.0% and a specificity of 96.4% as obtained by receiver operating characteristic analysis on 45 patient samples when 17-OHP > 39.1 ng/mL was selected as the cutoff value. Comparison between the LC-MS/MS and the AutoDELFIA immunoassay methods revealed a poor correlation for patient DBS samples (R2 = 0.6784); however, an excellent correlation was obtained for QC and proficiency test (PT) DBS samples (R2 = 0.9797). The LC-MS/MS method produced reliable results for 17-OHP and cortisol for the diagnosis of CAH. The AutoDELFIA immunoassay appears to be subject to matrix effects in the analysis for 17-OHP in DBS patient samples. The DBS samples of non-patient origin may not be suitable for assessing analytical accuracy of immunoassays. |
| Audience | Academic |
| Author | Morrissey, Mark Tavakoli, Norma P. Han, Liqiao Spink, David C. Cao, Zhimin (Tim) |
| Author_xml | – sequence: 1 givenname: Liqiao surname: Han fullname: Han, Liqiao organization: Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine – sequence: 2 givenname: Norma P. surname: Tavakoli fullname: Tavakoli, Norma P. organization: New York State Department of Health, Wadsworth Center – sequence: 3 givenname: Mark surname: Morrissey fullname: Morrissey, Mark organization: New York State Department of Health, Wadsworth Center – sequence: 4 givenname: David C. surname: Spink fullname: Spink, David C. organization: New York State Department of Health, Wadsworth Center – sequence: 5 givenname: Zhimin (Tim) surname: Cao fullname: Cao, Zhimin (Tim) email: tim.cao@health.ny.gov organization: New York State Department of Health, Wadsworth Center, College of Arts and Sciences, University at Albany, State University of New York |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/30456606$$D View this record in MEDLINE/PubMed |
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| CitedBy_id | crossref_primary_10_1007_s00216_019_02086_5 crossref_primary_10_1016_j_jchromb_2020_122117 crossref_primary_10_1111_cen_14422 crossref_primary_10_1016_j_jsbmb_2019_105389 crossref_primary_10_3390_ijns8040056 crossref_primary_10_1007_s00216_024_05411_9 crossref_primary_10_1038_s41598_024_66355_9 crossref_primary_10_1016_j_cca_2023_117304 crossref_primary_10_1210_endrev_bnab016 crossref_primary_10_1515_cclm_2024_0393 crossref_primary_10_1515_cclm_2023_0095 |
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| Keywords | Newborn screening 17-hydroxyprogesterone Cortisol Congenital adrenal hyperplasia Mass spectrometry |
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| PublicationDate_xml | – month: 01 year: 2019 text: 2019-01-01 day: 01 |
| PublicationDecade | 2010 |
| PublicationPlace | Berlin/Heidelberg |
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| PublicationTitle | Analytical and bioanalytical chemistry |
| PublicationTitleAbbrev | Anal Bioanal Chem |
| PublicationTitleAlternate | Anal Bioanal Chem |
| PublicationYear | 2019 |
| Publisher | Springer Berlin Heidelberg Springer Springer Nature B.V |
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publication-title: Clin Chem doi: 10.1373/clinchem.2003.027193 – ident: 1449_CR23 – volume: 7 start-page: 329 year: 2015 ident: 1449_CR11 publication-title: J Clin Res Pediatr Endocrinol doi: 10.4274/jcrpe.2180 – ident: 1449_CR22 – volume: 74 start-page: 662 year: 2009 ident: 1449_CR15 publication-title: Steroids doi: 10.1016/j.steroids.2009.02.008 – ident: 1449_CR8 |
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| SubjectTerms | 17-alpha-Hydroxyprogesterone - blood 17-alpha-Hydroxyprogesterone - chemistry 17-hydroxyprogesterone Adrenal glands Analytical Chemistry analytical specificity Biochemistry Blood Blood tests Characterization and Evaluation of Materials Chemical properties Chemistry Chemistry and Materials Science Chromatography Chromatography, Liquid Coefficient of variation Cortisol Food Science Hormones Humans Hydroxyprogesterone Hyperplasia Immunoassay Immunoassay - methods Immunoassays Infant, Newborn Laboratory Medicine Linearity Liquid chromatography Mass spectrometry Mass spectroscopy Mathematical analysis Medical screening Methods Molecular Structure Monitoring/Environmental Analysis Neonates patients Quantitation Research Paper Scientific imaging screening Sensitivity and Specificity Serologic Tests Spectroscopy Tandem Mass Spectrometry |
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| Title | Liquid chromatography-tandem mass spectrometry analysis of 17-hydroxyprogesterone in dried blood spots revealed matrix effect on immunoassay |
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