Annals of Laboratory Medicine最新文献

Background: Plasma oxalate measurements can be used for the screening and therapeutic monitoring of primary hyperoxaluria. We developed a gas chromatography-mass spectrometry (GC-MS) assay for plasma oxalate measurements with high sensitivity and suitable testing volumes for pediatric populations.

Methods: Plasma oxalate was extracted, derivatized, and analyzed by GC-MS. We measured the ion at m/z 261.10 to quantify oxalate and the ¹³C₂-oxalate ion (m/z: 263.15) as the internal standard. Method validation included determination of the linear range, limit of blank, limit of detection, lower limit of quantification, precision, recovery, carryover, interference, and dilution effect. The cut-off value between primary and non-primary hyperoxaluria in a pediatric population was analyzed.

Results: The detection limit was 0.78 μmol/L, and the linear range was up to 80.0 μmol/L. The between-day precision was 5.7% at 41.3 μmol/L and 13.1% at 1.6 μmol/L. The carryover was <0.2%. The recovery rate ranged from 90% to 110%. Interference analysis showed that Hb did not interfere with plasma oxalate quantification, whereas intralipids and bilirubin caused false elevation of oxalate concentrations. A cut-off of 13.9 μmol/L showed 63% specificity and 77% sensitivity, whereas a cut-off of 4.15 μmol/L showed 100% specificity and 20% sensitivity. The minimum required sample volume was 250 μL. The detected oxalate concentrations showed interference from instrument conditioning, sample preparation procedures, medications, and various clinical conditions.

Conclusions: GC-MS is a sensitive assay for quantifying plasma oxalate and is suitable for pediatric patients. Plasma oxalate concentrations should be interpreted in a clinical context.

A standardized protocol for the isolation of cell-free DNA (cfDNA) from cerebrospinal fluid (CSF) is lacking. Therefore, we established a cfDNA isolation protocol optimized for clinical CSF specimens, integrating acceptable modifications and using artificial CSF generated from remnant CSF spiked with reference cell-free tumor DNA (ctDNA). We compared the isolation yields of in vitro diagnostic (IVD)-certified column-based (CB) and magnetic bead-based (MB) isolation. Furthermore, we modified both methods, including pre- and post-elution steps. To confirm ctDNA integrity and quantify the variant allele frequency after isolation, we performed droplet digital PCR (ddPCR) targeting IDH1 R132C in the reference ctDNA. MB isolation had a higher yield than CB isolation (P<0.0001), and post-isolation vacuum increased the final concentration in both methods, with little effect on cfDNA integrity. Our study provides a protocol to maximize CSF-ctDNA concentrations in IVD testing and future studies.

Background: Marfan syndrome (MFS) is caused by fibrillin-1 gene (FBN1) variants. Mutational hotspots and/or well-established critical functional domains of FBN1 include cysteine residues, calcium-binding consensus sequences, and amino acids related to interdomain packaging. Previous guidelines for variant interpretation do not reflect the features of genes or related diseases. Using the Clinical Genome Resource (ClinGen) FBN1 variant curation expert panel (VCEP), we re-evaluated FBN1 germline variants reported as variants of uncertain significance (VUSs).

Methods: We re-evaluated 26 VUSs in FBN1 reported in 161 patients with MFS. We checked the variants in the Human Genome Mutation Database, ClinVar, and VarSome databases and assessed their allele frequencies using the gnomAD database. Patients' clinical information was reviewed.

Results: Four missense variants affecting cysteines (c.460T>C, c.1006T>C, c.5330G>C, and c.8020T>C) were reclassified as likely pathogenic and were assigned PM1_strong or PM1. Two intronic variants were reclassified as benign by granting BA1 (stand-alone). Four missense variants were reclassified as likely benign. BP5 criteria were applied in cases with an alternate molecular basis for disease, one of which (c.7231G>A) was discovered alongside a pathogenic de novo COL3A1 variant (c.1988G>T, p.Gly633Val).

Conclusions: Considering the high penetrance of FBN1 variants and clinical variability of MFS, the detection of pathogenic variants is important. The ClinGen FBN1 VCEP encompasses mutational hotspots and/or well-established critical functional domains and adjusts the criteria specifically for MFS; therefore, it is beneficial not only for identifying pathogenic FBN1 variants but also for distinguishing these variants from those that cause other connective tissue disorders with overlapping clinical features.

Although WHO declared the end of the public health emergency for coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), XBB lineages continue to evolve and emerge globally. In particular, XBB.1.5 and XBB.1.16 are raising concerns because of their high immune evasion, leading to apprehensions regarding vaccine efficacy reduction and potential reinfection. We aimed to investigate the COVID-19 outbreak in Korea and predict the likelihood of reinfection by testing neutralizing activity against live viruses from the S clade and 19 Omicron sublineages. We found a significant risk of infection with the currently prevalent XBB lineage for individuals who were either vaccinated early or infected during the initial Omicron outbreak. Vaccinated individuals were better equipped than unvaccinated individuals to produce neutralizing antibodies for other SARS-CoV-2 variants upon infection. Therefore, unvaccinated individuals do not easily develop neutralizing activity against other variants and face the highest risk of reinfection by the XBB lineage. Our study provides important information to facilitate the development of strategies for monitoring populations that would be the most susceptible to new COVID-19 outbreaks.

Background: Clinical chemistry tests are most widely used in clinical laboratories, and diverse measurement systems for these analyses are available in China. We evaluated the imprecision of clinical chemistry measurement systems based on internal QC (IQC) data.

Methods: IQC data for 27 general chemistry analytes were collected in February each year from 2013 to 2022. Four performance specifications were used to calculate pass rates for CVs of IQC data in 2022. Boxplots were drawn to analyze trends of CVs, and differences in CVs among different groups were assessed using the Mann-Whitney U-test or Kruskal-Wallis test.

Results: The number of participating laboratories increased significantly from 1,777 in 2013 to 5,425 in 2022. CVs significantly decreased for all 27 analytes, except creatine kinase and lipase. Triglycerides, total bilirubin, direct bilirubin, iron, and γ-glutamyl transferase achieved pass rates >80% for all goals. Nine analytes with pass rates <80% based on 1/3 allowable total error were further analyzed; the results indicated that closed systems exhibited lower CVs than open systems for all analytes, except total protein. For all nine analytes, differences were significant between tertiary hospitals and non-tertiary hospitals and between accredited and non-accredited laboratories.

Conclusions: The CVs of IQC data for clinical chemistry have seen a continuous overall improvement in China. However, there is ample room for imprecision improvement for several analytes, with stricter performance specifications.

Background: The mechanism and medical treatment target for degenerative aortic valve disease, including aortic stenosis, is not well studied. In this study, we investigated the effect of clonal hematopoiesis of indeterminate potential (CHIP) on the development of aortic valve sclerosis (AVS), a calcified aortic valve without significant stenosis.

Methods: Participants with AVS (valves ≥2 mm thick, high echogenicity, and a peak transaortic velocity of <2.5 m/sec) and an age- and sex-matched control group were enrolled. Twenty-four CHIP genes with common variants in cardiovascular disease were used to generate a next-generation sequencing panel. The primary endpoint was the CHIP detection rate between the AVS and control groups. Inverse-probability treatment weighting (IPTW) analysis was performed to adjust for differences in baseline characteristics.

Results: From April 2020 to April 2022, 187 participants (125 with AVS and 62 controls) were enrolled; the mean age was 72.6±8.5 yrs, and 54.5% were male. An average of 1.3 CHIP variants was observed. CHIP detection, defined by a variant allele frequency (VAF) of ≥0.5%, was similar between the groups. However, the AVS group had larger CHIP clones: 49 (39.2%) participants had a VAF of ≥1% (vs. 13 [21.0%] in the control group; P=0.020), and 25 (20.0%) had a VAF of ≥2% (vs. 4 [6.5%]; P=0.028). AVS is independently associated with a VAF of ≥1% (adjusted odds ratio: 2.44, 95% confidence interval: 1.11-5.36; P=0.027). This trend was concordant and clearer in the IPTW cohort.

Conclusions: Participants with AVS more commonly had larger CHIP clones than age- and sex-matched controls. Further studies are warranted to identify causality between AVS and CHIP.

Chimeric antigen receptor (CAR) T-cell therapy is a promising immunotherapy based on genetically engineered T cells derived from patients. The introduction of CAR T-cell therapy has changed the treatment paradigm of patients with B-cell lymphoid malignancies. However, challenging issues including managing life-threatening toxicities related to CAR T-cell infusion and resistance to CAR T-cell therapy, leading to progression or relapse, remain. This review summarizes the issues with currently approved CAR T-cell therapies for patients with relapsed or refractory B-cell lymphoid malignancies, including lymphoma and myeloma. We focus on unique toxicities after CAR T-cell therapy, such as cytokine-related events and hematological toxicities, and the mechanisms underlying post-CAR T-cell failure.

Circulating tumor DNA (ctDNA) has emerged as a promising tool for various clinical applications, including early diagnosis, therapeutic target identification, treatment response monitoring, prognosis evaluation, and minimal residual disease detection. Consequently, ctDNA assays have been incorporated into clinical practice. In this review, we offer an in-depth exploration of the clinical implementation of ctDNA assays. Notably, we examined existing evidence related to pre-analytical procedures, analytical components in current technologies, and result interpretation and reporting processes. The primary objective of this guidelines is to provide recommendations for the clinical utilization of ctDNA assays.