Pub Date : 2025-11-12DOI: 10.1093/clinchem/hvaf121
Robert L Fitzgerald,Anya Umlauf,Raymond T Suhandynata,David J Grelotti,Marilyn A Huestis,Kyle F Mastropietro,Igor Grant,Thomas D Marcotte
BACKGROUNDSeveral US states have per se laws using 2 or 5 ng/mL of delta-9-tetrahydrocannabinol (THC) as cutpoints for driving under the influence of cannabis, while some have zero-tolerance statutes. These cutpoints are considered prima facia evidence of driving impairment.METHODSIn a cohort of people who regularly use cannabis (N = 190) we measured baseline concentrations of THC after instructing participants to abstain from cannabis for at least 48 hours. Baseline driving performance was evaluated using a driving simulator. We also measured blood THC concentrations serially following a smoking session (placebo or active cannabis).RESULTSForty-three percent of the participants exceeded zero-tolerance statutes (≥0.5 ng/mL) at baseline. Twenty-four percent had baseline blood THC concentrations that were ≥2 ng/mL and 5.3% were ≥5 ng/mL. The maximum observed baseline blood concentration was 16.2 ng/mL. Six hours after smoking active cannabis, the median (interquartile range) difference in THC concentrations compared with baseline was 0.5 (0-0.9) ng/mL; a 1-sample t-test comparing the mean change to 0 was significant (P < 0.001). There was no difference when comparing the mean change to 0 in the placebo group (P = 0.69). Simulated driving performance was not different between those who exceed zero tolerance and per se cutpoints vs those who are below these cutpoints (P > 0.05).CONCLUSIONSMany regular users of cannabis exceed zero tolerance and per se THC cutpoint concentrations days after their last use, risking legal consequences despite no evidence of impairment.
{"title":"Per Se Driving Under the Influence of Cannabis Statutes and Blood Delta-9-Tetrahydrocannabinol Concentrations following Short-Term Cannabis Abstinence.","authors":"Robert L Fitzgerald,Anya Umlauf,Raymond T Suhandynata,David J Grelotti,Marilyn A Huestis,Kyle F Mastropietro,Igor Grant,Thomas D Marcotte","doi":"10.1093/clinchem/hvaf121","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf121","url":null,"abstract":"BACKGROUNDSeveral US states have per se laws using 2 or 5 ng/mL of delta-9-tetrahydrocannabinol (THC) as cutpoints for driving under the influence of cannabis, while some have zero-tolerance statutes. These cutpoints are considered prima facia evidence of driving impairment.METHODSIn a cohort of people who regularly use cannabis (N = 190) we measured baseline concentrations of THC after instructing participants to abstain from cannabis for at least 48 hours. Baseline driving performance was evaluated using a driving simulator. We also measured blood THC concentrations serially following a smoking session (placebo or active cannabis).RESULTSForty-three percent of the participants exceeded zero-tolerance statutes (≥0.5 ng/mL) at baseline. Twenty-four percent had baseline blood THC concentrations that were ≥2 ng/mL and 5.3% were ≥5 ng/mL. The maximum observed baseline blood concentration was 16.2 ng/mL. Six hours after smoking active cannabis, the median (interquartile range) difference in THC concentrations compared with baseline was 0.5 (0-0.9) ng/mL; a 1-sample t-test comparing the mean change to 0 was significant (P < 0.001). There was no difference when comparing the mean change to 0 in the placebo group (P = 0.69). Simulated driving performance was not different between those who exceed zero tolerance and per se cutpoints vs those who are below these cutpoints (P > 0.05).CONCLUSIONSMany regular users of cannabis exceed zero tolerance and per se THC cutpoint concentrations days after their last use, risking legal consequences despite no evidence of impairment.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"1 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145491481","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-09DOI: 10.1093/clinchem/hvaf142
Albertus T J Wierenga, Lucy B Hesp, Arjan Simpelaar, Linde M Morsink, Carolien M Woolthuis, Jan Jacob Schuringa, Isidor Minovic, Gerwin Huls, André B Mulder
Background NPM1 is a disease-defining gene in the diagnosis of acute myeloid leukemia (AML) and is important for measurable residual disease (MRD) assessment. Over 50 different NPM1 mutations have been described, but only the 3 most common are routinely monitored during follow-up. Methods We developed a multiplex droplet digital polymerase chain reaction (PCR) assay for measurement of both variant allele frequencies (VAF) and mRNA transcripts of 10 different NPM1 mutations, using one generic probe, one generic NPM1 reverse primer, and 10 mutation-specific NPM1 forward primers. ABL1 expression and AP3B1 VAF were used as references. The performance of the assay was tested in diagnosis and follow-up samples from patients with an NPM1-mutated AML. Results Our assay shows negligible false-positive signals and high assay precision, leading to low limits of detection of at least 0.01%. The assay can easily be expanded to cover more NPM1 mutations by adding extra mutation-specific forward primers to the primer mix. Overall, a good correlation between mutant NPM1 expression and VAF was found. However, we also observed discrepant variable ABL1 expression levels, especially in AML patients with fms-related receptor tyrosine kinase 3-internal tandem duplications co-mutations. Conclusion We developed a robust and extremely flexible mRNA- and gDNA-based multiplex droplet digital PCR NPM1 assay. Because the AML tumor load is better reflected by mutant NPM1 VAF than expression level, we recommend using the gDNA-based mutant NPM1 MRD assay with a VAF detection limit of 0.01%. For MRD signals below 0.01%, our more sensitive mRNA-based method can be used, although further research has to prove its clinical impact.
{"title":"Validation of a Multiplex mRNA- and gDNA-Based Droplet Digital PCR Assay in Acute Myeloid Leukemia Patients with an NPM1 Mutation","authors":"Albertus T J Wierenga, Lucy B Hesp, Arjan Simpelaar, Linde M Morsink, Carolien M Woolthuis, Jan Jacob Schuringa, Isidor Minovic, Gerwin Huls, André B Mulder","doi":"10.1093/clinchem/hvaf142","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf142","url":null,"abstract":"Background NPM1 is a disease-defining gene in the diagnosis of acute myeloid leukemia (AML) and is important for measurable residual disease (MRD) assessment. Over 50 different NPM1 mutations have been described, but only the 3 most common are routinely monitored during follow-up. Methods We developed a multiplex droplet digital polymerase chain reaction (PCR) assay for measurement of both variant allele frequencies (VAF) and mRNA transcripts of 10 different NPM1 mutations, using one generic probe, one generic NPM1 reverse primer, and 10 mutation-specific NPM1 forward primers. ABL1 expression and AP3B1 VAF were used as references. The performance of the assay was tested in diagnosis and follow-up samples from patients with an NPM1-mutated AML. Results Our assay shows negligible false-positive signals and high assay precision, leading to low limits of detection of at least 0.01%. The assay can easily be expanded to cover more NPM1 mutations by adding extra mutation-specific forward primers to the primer mix. Overall, a good correlation between mutant NPM1 expression and VAF was found. However, we also observed discrepant variable ABL1 expression levels, especially in AML patients with fms-related receptor tyrosine kinase 3-internal tandem duplications co-mutations. Conclusion We developed a robust and extremely flexible mRNA- and gDNA-based multiplex droplet digital PCR NPM1 assay. Because the AML tumor load is better reflected by mutant NPM1 VAF than expression level, we recommend using the gDNA-based mutant NPM1 MRD assay with a VAF detection limit of 0.01%. For MRD signals below 0.01%, our more sensitive mRNA-based method can be used, although further research has to prove its clinical impact.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"20 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-11-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145472700","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-09DOI: 10.1093/clinchem/hvaf136
Jin Sun, Jie Tang, Lu Wei, Juan Geng, Rui Xiao, Niu Li, Shuyuan Li, Jian Wang, Qihua Fu, Ruen Yao, Tingting Yu
Background Duchenne and Becker muscular dystrophies are X-linked neuromuscular disorders caused by mutations in the dystrophin gene (DMD). Duplications account for approximately 10% of pathogenic variants, but their structural complexity and variable clinical impact present significant challenges in pathogenicity interpretation. Methods We retrospectively analyzed whole exome sequencing and multiplex ligation-dependent probe amplification data from 3842 individuals, identifying 39 patients with DMD duplications. These patients underwent whole genome sequencing (WGS) to characterize duplication patterns, breakpoint features, and haplotype structures. Optical genome mapping (OGM) was additionally performed in WGS-unresolved cases to identify exact haplotypes. We also compared the diagnostic performance of different platforms and further investigated genotype–phenotype correlations. Results DMD duplications exhibited substantial structural heterogeneity and were characterized into 4 major patterns: tandem duplication (58%), duplication-normal-duplication (16%), duplication-inversion-duplication (16%), and intricate duplication (10%). Recurrent complex arrangements were associated with high-homology repeats and often extended beyond DMD, complicating haplotype interpretation. WGS alone failed to resolve haplotypes in 34% (13/38) of cases, while OGM missed certain exon duplications and small fragments in 33% of cases (2/6). Integration of WGS and OGM enabled precise haplotype reconstruction and improved genotype–phenotype correlation. Conclusions Our findings broaden the molecular spectrum of DMD duplications, highlight their widespread structural complexity, and emphasize the importance of integrating multiple technologies to precisely delineate duplication haplotype structures and assess the pathogenicity of variants with uncertain significance. These findings provide valuable insights for DMD duplication detection, pathogenicity evaluation, and genetic counseling.
{"title":"Integrated Genotyping Strategies for Uncovering Detailed Haplotype Structures and Characterization of DMD Duplications","authors":"Jin Sun, Jie Tang, Lu Wei, Juan Geng, Rui Xiao, Niu Li, Shuyuan Li, Jian Wang, Qihua Fu, Ruen Yao, Tingting Yu","doi":"10.1093/clinchem/hvaf136","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf136","url":null,"abstract":"Background Duchenne and Becker muscular dystrophies are X-linked neuromuscular disorders caused by mutations in the dystrophin gene (DMD). Duplications account for approximately 10% of pathogenic variants, but their structural complexity and variable clinical impact present significant challenges in pathogenicity interpretation. Methods We retrospectively analyzed whole exome sequencing and multiplex ligation-dependent probe amplification data from 3842 individuals, identifying 39 patients with DMD duplications. These patients underwent whole genome sequencing (WGS) to characterize duplication patterns, breakpoint features, and haplotype structures. Optical genome mapping (OGM) was additionally performed in WGS-unresolved cases to identify exact haplotypes. We also compared the diagnostic performance of different platforms and further investigated genotype–phenotype correlations. Results DMD duplications exhibited substantial structural heterogeneity and were characterized into 4 major patterns: tandem duplication (58%), duplication-normal-duplication (16%), duplication-inversion-duplication (16%), and intricate duplication (10%). Recurrent complex arrangements were associated with high-homology repeats and often extended beyond DMD, complicating haplotype interpretation. WGS alone failed to resolve haplotypes in 34% (13/38) of cases, while OGM missed certain exon duplications and small fragments in 33% of cases (2/6). Integration of WGS and OGM enabled precise haplotype reconstruction and improved genotype–phenotype correlation. Conclusions Our findings broaden the molecular spectrum of DMD duplications, highlight their widespread structural complexity, and emphasize the importance of integrating multiple technologies to precisely delineate duplication haplotype structures and assess the pathogenicity of variants with uncertain significance. These findings provide valuable insights for DMD duplication detection, pathogenicity evaluation, and genetic counseling.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"10 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-11-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145472871","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1093/clinchem/hvaf094
Ruben Y Luo, Priscilla S W Yeung, Morgan W Mann, Lichao Zhang, Yifei K Yang, Andrew N Hoofnagle
Background: Protein biomarkers are routinely measured for disease diagnosis and prognosis in clinical laboratories. Since most assays focus on protein quantity, information about proteoforms is often not acquired. Proteoforms of a protein represent the complex integration of genetic polymorphism, alternative splicing of RNA transcripts, and post-translational modifications (PTMs) on the amino-acid backbone. A detailed analysis of the post-translationally modified proteoforms (PTMPs), which are influenced by pathophysiological conditions, may lead to more precise diagnosis and prognosis.
Content: This article first discusses the methodologies used to accurately detect and characterize PTMPs, i.e., immunoassays, electrophoresis, chromatography, and intact and proteolysis-aided mass spectrometry techniques. Then it reviews specific examples of PTMP biomarkers that have been successfully translated from biomarker discovery to clinical use. The examples include β2-transferrin for cerebrospinal fluid leak diagnosis, phosphorylated tau proteoforms for Alzheimer disease diagnosis, and fucosylated alpha-fetoprotein for hepatocellular carcinoma prognosis. In addition, the article provides prospective views of novel analytical technologies and promising new PTMP biomarkers entering clinical practice.
Summary: In summary, PTMs are controlled by biochemical processes to modulate the functions of proteins by expanding their chemical diversity. PTM alterations in proteins can be indicators for pathophysiological conditions. Advances in analytical technologies are deepening our understanding of PTMPs and paving the way for their translation to clinical use. As research continues to discover the clinical meaning of PTMP biomarkers, they are poised to become valuable additions to the clinical testing menu for precision medicine.
{"title":"Post-Translationally Modified Proteoforms as Biomarkers: From Discovery to Clinical Use.","authors":"Ruben Y Luo, Priscilla S W Yeung, Morgan W Mann, Lichao Zhang, Yifei K Yang, Andrew N Hoofnagle","doi":"10.1093/clinchem/hvaf094","DOIUrl":"10.1093/clinchem/hvaf094","url":null,"abstract":"<p><strong>Background: </strong>Protein biomarkers are routinely measured for disease diagnosis and prognosis in clinical laboratories. Since most assays focus on protein quantity, information about proteoforms is often not acquired. Proteoforms of a protein represent the complex integration of genetic polymorphism, alternative splicing of RNA transcripts, and post-translational modifications (PTMs) on the amino-acid backbone. A detailed analysis of the post-translationally modified proteoforms (PTMPs), which are influenced by pathophysiological conditions, may lead to more precise diagnosis and prognosis.</p><p><strong>Content: </strong>This article first discusses the methodologies used to accurately detect and characterize PTMPs, i.e., immunoassays, electrophoresis, chromatography, and intact and proteolysis-aided mass spectrometry techniques. Then it reviews specific examples of PTMP biomarkers that have been successfully translated from biomarker discovery to clinical use. The examples include β2-transferrin for cerebrospinal fluid leak diagnosis, phosphorylated tau proteoforms for Alzheimer disease diagnosis, and fucosylated alpha-fetoprotein for hepatocellular carcinoma prognosis. In addition, the article provides prospective views of novel analytical technologies and promising new PTMP biomarkers entering clinical practice.</p><p><strong>Summary: </strong>In summary, PTMs are controlled by biochemical processes to modulate the functions of proteins by expanding their chemical diversity. PTM alterations in proteins can be indicators for pathophysiological conditions. Advances in analytical technologies are deepening our understanding of PTMPs and paving the way for their translation to clinical use. As research continues to discover the clinical meaning of PTMP biomarkers, they are poised to become valuable additions to the clinical testing menu for precision medicine.</p>","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":" ","pages":"1111-1124"},"PeriodicalIF":6.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144945710","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1093/clinchem/hvaf096
Jorieke Weiden,Susan D P W M de Jonge-Peeters,Johannes M W van den Ouweland
{"title":"Excess Leading to Deficiency: An Unusual Cause of Cytopenia.","authors":"Jorieke Weiden,Susan D P W M de Jonge-Peeters,Johannes M W van den Ouweland","doi":"10.1093/clinchem/hvaf096","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf096","url":null,"abstract":"","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"28 1","pages":"1105-1108"},"PeriodicalIF":9.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145433996","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1093/clinchem/hvaf107
Brian D Adkins,Sharon K Germans,Charles F Timmons,Hung S Luu
{"title":"Commentary on Excess Leading to Deficiency: An Unusual Cause of Cytopenia.","authors":"Brian D Adkins,Sharon K Germans,Charles F Timmons,Hung S Luu","doi":"10.1093/clinchem/hvaf107","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf107","url":null,"abstract":"","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"35 1","pages":"1109"},"PeriodicalIF":9.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145433992","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1093/clinchem/hvaf084
Nicholas C Spies
{"title":"Embracing Generative Artificial Intelligence as a Support Tool for Clinical Decision-Making.","authors":"Nicholas C Spies","doi":"10.1093/clinchem/hvaf084","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf084","url":null,"abstract":"","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"22 1","pages":"1178-1179"},"PeriodicalIF":9.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145433994","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1093/clinchem/hvaf108
Ravinder Sodi
{"title":"Commentary on Excess Leading to Deficiency: An Unusual Cause of Cytopenia.","authors":"Ravinder Sodi","doi":"10.1093/clinchem/hvaf108","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf108","url":null,"abstract":"","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"88 1","pages":"1110"},"PeriodicalIF":9.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145433993","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BACKGROUNDDespite almost 3 decades of research, the mechanisms underlying the bidirectional trafficking of cells at the maternal-fetal interface that gives rise to microchimerism remain poorly understood. A major barrier to progress has been the lack of suitable detection methods capable of distinguishing maternal from fetal cells within the spatial context of the human placenta. To address this, we developed a novel detection method based on padlock probe technology to differentiate haploidentical cells in placental tissues.METHODSPadlock probes were designed to target single nucleotide polymorphisms (SNPs) present in messenger RNA transcripts. The assays were first validated in cell lines and subsequently applied to placental tissue to assess its ability to distinguish between maternal and fetal cells.RESULTSWe established a panel of 27 assays targeting 3 human leukocyte antigen-A alleles and 12 biallelic SNPs. The method demonstrated high specificity and sensitivity, detecting minor cell populations at dilutions as low as 1:10 000. Proof of concept was obtained in a decidua basalis specimen, showing the assays' capability to distinguish maternal and fetal cells within placental tissue.CONCLUSIONSWe present a novel, sex-unbiased methodology for the in situ visualization of haploidentical (microchimeric) cells. This approach enables the study of maternal-fetal cellular interactions within their native tissues at the maternal-fetal interface.
{"title":"Single Nucleotide Polymorphism Typing Going Spatial: In Situ Padlock Probes Targeting mRNA Variants to Identify Haploidentical Cells within the Tissue Environment.","authors":"Emiel Slaats,Katharina Schuch,Katja Sallinger,Julia Schönberger,Bernadette Luise Bramreiter,Jaqueline Anholts,Daniel Pitz Jacobsen,Anne Cathrine Staff,Amin El-Heliebi,Michael Eikmans,Thomas Kroneis","doi":"10.1093/clinchem/hvaf119","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf119","url":null,"abstract":"BACKGROUNDDespite almost 3 decades of research, the mechanisms underlying the bidirectional trafficking of cells at the maternal-fetal interface that gives rise to microchimerism remain poorly understood. A major barrier to progress has been the lack of suitable detection methods capable of distinguishing maternal from fetal cells within the spatial context of the human placenta. To address this, we developed a novel detection method based on padlock probe technology to differentiate haploidentical cells in placental tissues.METHODSPadlock probes were designed to target single nucleotide polymorphisms (SNPs) present in messenger RNA transcripts. The assays were first validated in cell lines and subsequently applied to placental tissue to assess its ability to distinguish between maternal and fetal cells.RESULTSWe established a panel of 27 assays targeting 3 human leukocyte antigen-A alleles and 12 biallelic SNPs. The method demonstrated high specificity and sensitivity, detecting minor cell populations at dilutions as low as 1:10 000. Proof of concept was obtained in a decidua basalis specimen, showing the assays' capability to distinguish maternal and fetal cells within placental tissue.CONCLUSIONSWe present a novel, sex-unbiased methodology for the in situ visualization of haploidentical (microchimeric) cells. This approach enables the study of maternal-fetal cellular interactions within their native tissues at the maternal-fetal interface.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"90 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145440691","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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