Sofie D.H. Olsen , Astrid M. Kolte , Nina Bang , Maria Christine Krog , Rudi Steffensen , Henriette S. Nielsen , Marianne A. Jakobsen
{"title":"微嵌合检测用indel面板的研制","authors":"Sofie D.H. Olsen , Astrid M. Kolte , Nina Bang , Maria Christine Krog , Rudi Steffensen , Henriette S. Nielsen , Marianne A. Jakobsen","doi":"10.1016/j.yexmp.2022.104804","DOIUrl":null,"url":null,"abstract":"<div><h3>Objectives</h3><p>The aim of the study was to create a simple assay for microchimerism detection independent of sex and without HLA genotyping.</p></div><div><h3>Methods</h3><p><span><span>The method is based on detection of insertion or deletions utilizing a multiplex PCR followed by fragment analysis by </span>capillary electrophoresis, and probe-based qPCR assays. A total of 192 samples, taken either before pregnancy, during 1st trimester, or either during </span>2nd trimester<span><span> or at miscarriage, obtained from a cohort of 97 female patients with either primary or secondary recurrent pregnancy loss, were screened for fetal microchimerism by the </span>indel panel as well as an existing assay based on detection of the Y-chromosome marker; DYS14.</span></p></div><div><h3>Results</h3><p><span>The overall prevalence of DYS14 positive samples was 29% (55/192) whereas 32% (61/192) tested positive by the indel method. There was an overall agreement of 64% (122/192) between the results obtained by the two methods. A Fisher's Exact test showed no statistic significant difference in the prevalence of microchimerism detected by the two methods at any of the three times of sampling. The distribution of the number of positive wells detected by both methods were compared by a Mann-Whitney </span><em>U</em> test, which showed no statistically significant difference at any of the three times of sampling.</p></div><div><h3>Conclusion</h3><p>The data indicates that microchimerism can be detected efficiently by the indel method. This makes it possible to detect both female and male cells without the need of HLA-genotyping. Furthermore, the indel method has potential to be implemented as a routine analysis. This will remove the sex bias in future explorations of the role microchimerism plays in health and disease.</p></div>","PeriodicalId":12176,"journal":{"name":"Experimental and molecular pathology","volume":null,"pages":null},"PeriodicalIF":2.8000,"publicationDate":"2022-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"The development of an indel panel for microchimerism detection\",\"authors\":\"Sofie D.H. Olsen , Astrid M. Kolte , Nina Bang , Maria Christine Krog , Rudi Steffensen , Henriette S. Nielsen , Marianne A. Jakobsen\",\"doi\":\"10.1016/j.yexmp.2022.104804\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Objectives</h3><p>The aim of the study was to create a simple assay for microchimerism detection independent of sex and without HLA genotyping.</p></div><div><h3>Methods</h3><p><span><span>The method is based on detection of insertion or deletions utilizing a multiplex PCR followed by fragment analysis by </span>capillary electrophoresis, and probe-based qPCR assays. A total of 192 samples, taken either before pregnancy, during 1st trimester, or either during </span>2nd trimester<span><span> or at miscarriage, obtained from a cohort of 97 female patients with either primary or secondary recurrent pregnancy loss, were screened for fetal microchimerism by the </span>indel panel as well as an existing assay based on detection of the Y-chromosome marker; DYS14.</span></p></div><div><h3>Results</h3><p><span>The overall prevalence of DYS14 positive samples was 29% (55/192) whereas 32% (61/192) tested positive by the indel method. There was an overall agreement of 64% (122/192) between the results obtained by the two methods. A Fisher's Exact test showed no statistic significant difference in the prevalence of microchimerism detected by the two methods at any of the three times of sampling. The distribution of the number of positive wells detected by both methods were compared by a Mann-Whitney </span><em>U</em> test, which showed no statistically significant difference at any of the three times of sampling.</p></div><div><h3>Conclusion</h3><p>The data indicates that microchimerism can be detected efficiently by the indel method. This makes it possible to detect both female and male cells without the need of HLA-genotyping. Furthermore, the indel method has potential to be implemented as a routine analysis. This will remove the sex bias in future explorations of the role microchimerism plays in health and disease.</p></div>\",\"PeriodicalId\":12176,\"journal\":{\"name\":\"Experimental and molecular pathology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.8000,\"publicationDate\":\"2022-08-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Experimental and molecular pathology\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0014480022000673\",\"RegionNum\":4,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"PATHOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Experimental and molecular pathology","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0014480022000673","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"PATHOLOGY","Score":null,"Total":0}
The development of an indel panel for microchimerism detection
Objectives
The aim of the study was to create a simple assay for microchimerism detection independent of sex and without HLA genotyping.
Methods
The method is based on detection of insertion or deletions utilizing a multiplex PCR followed by fragment analysis by capillary electrophoresis, and probe-based qPCR assays. A total of 192 samples, taken either before pregnancy, during 1st trimester, or either during 2nd trimester or at miscarriage, obtained from a cohort of 97 female patients with either primary or secondary recurrent pregnancy loss, were screened for fetal microchimerism by the indel panel as well as an existing assay based on detection of the Y-chromosome marker; DYS14.
Results
The overall prevalence of DYS14 positive samples was 29% (55/192) whereas 32% (61/192) tested positive by the indel method. There was an overall agreement of 64% (122/192) between the results obtained by the two methods. A Fisher's Exact test showed no statistic significant difference in the prevalence of microchimerism detected by the two methods at any of the three times of sampling. The distribution of the number of positive wells detected by both methods were compared by a Mann-Whitney U test, which showed no statistically significant difference at any of the three times of sampling.
Conclusion
The data indicates that microchimerism can be detected efficiently by the indel method. This makes it possible to detect both female and male cells without the need of HLA-genotyping. Furthermore, the indel method has potential to be implemented as a routine analysis. This will remove the sex bias in future explorations of the role microchimerism plays in health and disease.
期刊介绍:
Under new editorial leadership, Experimental and Molecular Pathology presents original articles on disease processes in relation to structural and biochemical alterations in mammalian tissues and fluids and on the application of newer techniques of molecular biology to problems of pathology in humans and other animals. The journal also publishes selected interpretive synthesis reviews by bench level investigators working at the "cutting edge" of contemporary research in pathology. In addition, special thematic issues present original research reports that unravel some of Nature''s most jealously guarded secrets on the pathologic basis of disease.
Research Areas include: Stem cells; Neoangiogenesis; Molecular diagnostics; Polymerase chain reaction; In situ hybridization; DNA sequencing; Cell receptors; Carcinogenesis; Pathobiology of neoplasia; Complex infectious diseases; Transplantation; Cytokines; Flow cytomeric analysis; Inflammation; Cellular injury; Immunology and hypersensitivity; Athersclerosis.