Benedetta Peruzzi , Serena Guerrieri , Tiziana Biagioli , Luisa Lanzilao , Sara Pratesi , Sara Bencini , Marinella Statello , Alessia Carraresi , Stefania Stefanelli , Martina Tonelli , Marco Brogi , Manuela Capone , Alessio Mazzoni , Anna Maria Grazia Gelli , Alessandra Fanelli , Roberto Caporale , Francesco Annunziato
{"title":"采用高效液相色谱法和流式细胞术相结合的方法分析胎儿产妇出血评估中的 HbF","authors":"Benedetta Peruzzi , Serena Guerrieri , Tiziana Biagioli , Luisa Lanzilao , Sara Pratesi , Sara Bencini , Marinella Statello , Alessia Carraresi , Stefania Stefanelli , Martina Tonelli , Marco Brogi , Manuela Capone , Alessio Mazzoni , Anna Maria Grazia Gelli , Alessandra Fanelli , Roberto Caporale , Francesco Annunziato","doi":"10.1016/j.plabm.2024.e00401","DOIUrl":null,"url":null,"abstract":"<div><h3>Introduction</h3><p>Recently, a flow cytometric (FC) based test has been developed for detection of circulating fetal cells to replace the less accurate and reproducible Kleihauer-Betke test.</p><p>FC test is easier to perform, it can distinguish the origin of fetal cells, but it is expensive and available in highly specialized laboratories. We evaluated the introduction of high-performance liquid chromatography (HPLC) approach as initial screening to identify patients who need an additional FC test to better discriminate the nature of haemoglobin-F (HbF) positive cells.</p></div><div><h3>Methods</h3><p>Blood samples from 130 pregnant women suspected to have fetomaternal haemorrhage were analysed with HPLC and FC methods. The cut-off for HbF HPLC concentration was calculated. Statistical analyses for the evaluation of HPLC as a screening method were performed. The positivity cut-off of HbF to be used as decision-making value to continue the investigation was calculated.</p></div><div><h3>Results</h3><p>An excellent agreement (R<sup>2</sup> > 0.90) was observed between the percentage of HbF obtained by HPLC and the percentage of fetal cells detected by FC. Results obtained from each assay were compared to define the HPLC threshold below which it is not necessary to continue the investigations, confirming the maternal nature of the HbF positive cells detected. Our study demonstrated that a cut-off of 1.0 % HbF obtained by HPLC was associated with the lowest rate of false negative results in our patient cohort.</p></div><div><h3>Conclusions</h3><p>This study provides a new FMH investigation approach that possibly leads to a reduction in times and costs of the analysis.</p></div>","PeriodicalId":20421,"journal":{"name":"Practical Laboratory Medicine","volume":"40 ","pages":"Article e00401"},"PeriodicalIF":1.7000,"publicationDate":"2024-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2352551724000477/pdfft?md5=8fe6e1a4d570fbec4024ab791d7c745f&pid=1-s2.0-S2352551724000477-main.pdf","citationCount":"0","resultStr":"{\"title\":\"HPLC and flow cytometry combined approach for HbF analysis in fetomaternal haemorrhage evaluation\",\"authors\":\"Benedetta Peruzzi , Serena Guerrieri , Tiziana Biagioli , Luisa Lanzilao , Sara Pratesi , Sara Bencini , Marinella Statello , Alessia Carraresi , Stefania Stefanelli , Martina Tonelli , Marco Brogi , Manuela Capone , Alessio Mazzoni , Anna Maria Grazia Gelli , Alessandra Fanelli , Roberto Caporale , Francesco Annunziato\",\"doi\":\"10.1016/j.plabm.2024.e00401\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Introduction</h3><p>Recently, a flow cytometric (FC) based test has been developed for detection of circulating fetal cells to replace the less accurate and reproducible Kleihauer-Betke test.</p><p>FC test is easier to perform, it can distinguish the origin of fetal cells, but it is expensive and available in highly specialized laboratories. We evaluated the introduction of high-performance liquid chromatography (HPLC) approach as initial screening to identify patients who need an additional FC test to better discriminate the nature of haemoglobin-F (HbF) positive cells.</p></div><div><h3>Methods</h3><p>Blood samples from 130 pregnant women suspected to have fetomaternal haemorrhage were analysed with HPLC and FC methods. The cut-off for HbF HPLC concentration was calculated. Statistical analyses for the evaluation of HPLC as a screening method were performed. The positivity cut-off of HbF to be used as decision-making value to continue the investigation was calculated.</p></div><div><h3>Results</h3><p>An excellent agreement (R<sup>2</sup> > 0.90) was observed between the percentage of HbF obtained by HPLC and the percentage of fetal cells detected by FC. Results obtained from each assay were compared to define the HPLC threshold below which it is not necessary to continue the investigations, confirming the maternal nature of the HbF positive cells detected. Our study demonstrated that a cut-off of 1.0 % HbF obtained by HPLC was associated with the lowest rate of false negative results in our patient cohort.</p></div><div><h3>Conclusions</h3><p>This study provides a new FMH investigation approach that possibly leads to a reduction in times and costs of the analysis.</p></div>\",\"PeriodicalId\":20421,\"journal\":{\"name\":\"Practical Laboratory Medicine\",\"volume\":\"40 \",\"pages\":\"Article e00401\"},\"PeriodicalIF\":1.7000,\"publicationDate\":\"2024-05-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S2352551724000477/pdfft?md5=8fe6e1a4d570fbec4024ab791d7c745f&pid=1-s2.0-S2352551724000477-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Practical Laboratory Medicine\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2352551724000477\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"MEDICAL LABORATORY TECHNOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Practical Laboratory Medicine","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2352551724000477","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"MEDICAL LABORATORY TECHNOLOGY","Score":null,"Total":0}
HPLC and flow cytometry combined approach for HbF analysis in fetomaternal haemorrhage evaluation
Introduction
Recently, a flow cytometric (FC) based test has been developed for detection of circulating fetal cells to replace the less accurate and reproducible Kleihauer-Betke test.
FC test is easier to perform, it can distinguish the origin of fetal cells, but it is expensive and available in highly specialized laboratories. We evaluated the introduction of high-performance liquid chromatography (HPLC) approach as initial screening to identify patients who need an additional FC test to better discriminate the nature of haemoglobin-F (HbF) positive cells.
Methods
Blood samples from 130 pregnant women suspected to have fetomaternal haemorrhage were analysed with HPLC and FC methods. The cut-off for HbF HPLC concentration was calculated. Statistical analyses for the evaluation of HPLC as a screening method were performed. The positivity cut-off of HbF to be used as decision-making value to continue the investigation was calculated.
Results
An excellent agreement (R2 > 0.90) was observed between the percentage of HbF obtained by HPLC and the percentage of fetal cells detected by FC. Results obtained from each assay were compared to define the HPLC threshold below which it is not necessary to continue the investigations, confirming the maternal nature of the HbF positive cells detected. Our study demonstrated that a cut-off of 1.0 % HbF obtained by HPLC was associated with the lowest rate of false negative results in our patient cohort.
Conclusions
This study provides a new FMH investigation approach that possibly leads to a reduction in times and costs of the analysis.
期刊介绍：
Practical Laboratory Medicine is a high-quality, peer-reviewed, international open-access journal publishing original research, new methods and critical evaluations, case reports and short papers in the fields of clinical chemistry and laboratory medicine. The objective of the journal is to provide practical information of immediate relevance to workers in clinical laboratories. The primary scope of the journal covers clinical chemistry, hematology, molecular biology and genetics relevant to laboratory medicine, microbiology, immunology, therapeutic drug monitoring and toxicology, laboratory management and informatics. We welcome papers which describe critical evaluations of biomarkers and their role in the diagnosis and treatment of clinically significant disease, validation of commercial and in-house IVD methods, method comparisons, interference reports, the development of new reagents and reference materials, reference range studies and regulatory compliance reports. Manuscripts describing the development of new methods applicable to laboratory medicine (including point-of-care testing) are particularly encouraged, even if preliminary or small scale.
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