Biology subjects

Sellers, B.

Publications and source records attributed to Sellers, B..

2 recordsLinked to original sources

bioRxiv2022

Multiparameter stimulation mapping of signaling states in single pediatric immune cells reveals heightened tonic activation during puberty

Cellular stimulation via factors such as cytokines followed by multiparameter single-cell measurements is a powerful approach to interrogate cellular functions. However, transforming such high-dimensional data into biological insights presents unique challenges, particularly given the extensive response heterogeneity among single cells, such as the presence of bimodal responding versus non-responding subpopulations upon stimulation. Here we present an unsupervised high-dimensional approach for analyzing stimulation responses at the single cell level (HDStIM) and apply it to evaluate how pediatric development may shape peripheral immune cell signaling states and responsiveness to stimulations in 42 subjects (age: 2 - 16). We show that in comparison to the conventional approach of assessing one marker at a time by averaging across single cells, HDStIM can effectively learn, in an unsupervised fashion, the multi-parameter signature of responding versus non-responding cells to accurately quantify responses within cell populations. HDStIM reveals that the extent of pre-stimulation/baseline activation of interferon-related and TCR signaling molecules in myeloid and T cells, respectively, increases during puberty. This suggests that puberty is marked by a heightened "tonic" activation state in these cells, perhaps to strengthen defense against pathogens during this period of human development.

immunology↗

bioRxiv2019

A Modified Injector and Sample Acquisition Protocol Can Improve Data Quality and Reduce Inter-Instrument Variability of the Helios Mass Cytometer

Mass cytometry is a powerful tool for high dimensional single cell characterization. Since the introduction of the first commercial CyTOF mass cytometer by DVS Sciences in 2009, mass cytometry technology has matured and become more widely utilized, with sequential platform upgrades designed to address specific limitations and to expand the capabilities of the platform. Fluidigms 3rd-generation Helios mass cytometer introduced a number of upgrades over the previous CyTOF2. One of these new features is a modified narrow bore sample injector that generates smaller ion clouds, which is expected to improve sensitivity and throughput. However, following rigorous testing we find that the narrow-bore sample injector may have unintended negative consequences on data quality and result in lower median signal intensities and higher coefficients of variation in antibody expression. We describe an alternative Helios acquisition protocol using a wider bore injector, which largely mitigates these data quality issues. We directly compare these two protocols in a multi-site study of 10 Helios instruments across 7 institutions and show that the modified protocol improves data quality and reduces inter-instrument variability. These findings highlight and address an important source of technical variability in mass cytometry experiments that is of particular relevance in the setting of multi-center studies.

immunology↗