Towards Sensitive and Cost-Effective Chimerism Assays Using FABCASE: A Fast Approach for Estimating Assay Informativity

IF 2.3 4区医学 Q3 HEMATOLOGY International Journal of Laboratory Hematology Pub Date : 2025-03-16 DOI:10.1111/ijlh.14460

Matthijs Vynck

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Abstract

Background

Chimerism monitoring is part of the standard of care for patients following an allogeneic hematopoietic stem cell transplantation. There has recently been a move towards sensitive, high-throughput (next-generation) sequencing analysis of biallelic markers for this purpose. Determining the number and properties of the markers to include in an assay to achieve reliable yet cost-effective chimerism quantification is an underexposed but critical part of chimerism assay development, optimization, and validation.

Methods

We develop FABCASE (Fast and Accurate Biallelic Chimerism Assay Size Estimation), an approach to estimate the required number of markers to screen to obtain a given informativity rate. We explore several practical examples that illustrate the diverse applications of FABCASE beyond calculating the required number of markers to screen.

Results

FABCASE offers a more than four orders of magnitude speed improvement compared to a previously introduced Monte Carlo simulation approach, unlocking extensive in silico scenario analyses. We find that minor allele frequency (MAF) and informative rate estimation based on small sample series (tens) are reasonably accurate. MAFs may vary drastically between populations, and the number of required markers to attain a preset informativity rate is inflated (here, +28%) when not optimized. Marker subset selection from a pool of candidate markers is little impacted by small-to-medium MAF differences (0%–20% MAF). Prioritizing markers with uniform amplification efficiency reduces sequencing depth requirements and improves cost-effectiveness. A web graphical user interface facilitating marker set informativity evaluation is available at https://mvynck.shinyapps.io/FABCASE.

Conclusion

FABCASE facilitates the design, refinement, and implementation of sensitive and cost-effective chimerism assays. Due attention should be given to an assay's marker MAFs, sensitivities, and amplification efficiencies.

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使用FABCASE实现敏感和具有成本效益的嵌合分析：一种快速估计分析信息的方法。

背景：嵌合监测是同种异体造血干细胞移植后患者护理标准的一部分。最近有一种针对双等位基因标记的敏感、高通量（下一代）测序分析的趋势。确定用于测定的标记物的数量和性质，以实现可靠且具有成本效益的嵌合定量，是嵌合测定开发、优化和验证的一个未充分暴露但关键的部分。方法：我们开发了FABCASE（快速准确的双等位基因嵌合测定大小估计），这是一种估计需要筛选的标记数量以获得给定信息率的方法。我们探讨了几个实际的例子，这些例子说明了FABCASE在计算需要筛选的标记数量之外的各种应用。结果：与之前引入的蒙特卡罗模拟方法相比，FABCASE提供了超过四个数量级的速度改进，解锁了广泛的硅场景分析。我们发现小等位基因频率（MAF）和基于小样本序列（几十）的信息率估计是相当准确的。maf在种群之间可能会有很大的差异，并且在没有优化的情况下，达到预设信息性率所需的标记数量会膨胀（这里是+28%）。从候选标记池中选择标记子集几乎不受较小至中等MAF差异（0%-20% MAF）的影响。优先考虑具有均匀扩增效率的标记可降低测序深度要求并提高成本效益。web图形用户界面促进标记集信息性评估可在https://mvynck.shinyapps.io/FABCASE.Conclusion: FABCASE促进设计，改进，并实施敏感和具有成本效益的嵌合分析。应适当注意测定的标记物maf、灵敏度和扩增效率。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

International Journal of Laboratory Hematology 医学-血液学

CiteScore

4.50

自引率

6.70%

发文量

211

审稿时长

6-12 weeks

期刊介绍： The International Journal of Laboratory Hematology provides a forum for the communication of new developments, research topics and the practice of laboratory haematology. The journal publishes invited reviews, full length original articles, and correspondence. The International Journal of Laboratory Hematology is the official journal of the International Society for Laboratory Hematology, which addresses the following sub-disciplines: cellular analysis, flow cytometry, haemostasis and thrombosis, molecular diagnostics, haematology informatics, haemoglobinopathies, point of care testing, standards and guidelines. The journal was launched in 2006 as the successor to Clinical and Laboratory Hematology, which was first published in 1979. An active and positive editorial policy ensures that work of a high scientific standard is reported, in order to bridge the gap between practical and academic aspects of laboratory haematology.

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