easysep human cd45 depletion kit ii  (STEMCELL Technologies Inc)

Journal: Translational Lung Cancer Research

Article Title: Stem-like exhausted CD8 T cells in pleural effusions predict improved survival in non-small cell lung cancer (NSCLC) and mesothelioma

doi: 10.21037/tlcr-24-284

Figure Lengend Snippet: Mass cytometry based phenotypic profiling identifies heterogeneity of Tex stem and Tex term CD8 T cells in pleural effusions of NSCLC and mesothelioma patients. (A) UMAP projections of CD8 T cell derived from malignant pleural effusions of NSCLC patients (n=3) at the early timepoint. Normalized intensity of individual markers. Gated on DNA + cisplatin – CD45 + CD14 – CD16 – CD3 + gdTCR – . (B) Heatmap depicting normalized mean marker intensity measured by mass cytometry in each cluster of NSCLC patients. (C) Distribution of phenotypes in Tex term (top) and Tex stem (bottom) of NSCLC patients. Each bubble represents cells with specific phenotypes, with bubble size reflecting their frequency among total cells, and bubble color indicating distinct cluster types. (D) UMAP projections of CD8 T cell derived from malignant pleural effusions of mesothelioma patients (n=5) at the early timepoint. Normalized intensity of individual markers. Gated on DNA + cisplatin – CD45 + CD14 – CD16 – CD3 + gdTCR – of mesothelioma patients. (E) Heatmap depicting normalized mean marker intensity measured by mass cytometry in each cluster of mesothelioma patients. (F) Distribution of phenotypes in Tex term (top) and Tex stem cells (bottom) of mesothelioma patients. Each bubble represents cells with specific phenotypes, with bubble size reflecting their frequency among total cells, and bubble color indicating distinct cluster types. (G) Heatmap depicting normalized mean marker intensity measured by mass cytometry in Tex term and Tex stem cells. NSCLC, non-small cell lung cancer; Tex stem , stem-like exhausted CD8 T cells; Tex term , terminally exhausted CD8 T cells; UMAP, uniform manifold approximation and projection; SCM, memory stem cell; CM, central memory; EM, effector memory; EMRA, effector memory re-expressing CD45RA; MAIT, mucosal-associated invariant T.

Article Snippet: For some samples, CD45 + cells were positively selected using EasySep Human CD45 Depletion Kit (Stemcell, cat17898) following the manufacturer’s instructions before cryopreservation.

Techniques: Mass Cytometry, Derivative Assay, Marker, Expressing

Journal: bioRxiv

Article Title: Mapping the Cellular Biogeography of Human Bone Marrow Niches Using Single-Cell Transcriptomics and Proteomic Imaging

doi: 10.1101/2024.03.14.585083

Figure Lengend Snippet: A ) UMAP computed from 19,257 mesenchymal cells from 12 individuals showing different mesenchymal subsets, with RNAlo MSCs being excluded. B ) Dot plot showing normalized expression of key mesenchymal marker genes in MSC subsets. Rows and columns were manually ordered. C ) Dot plot showing the normalized expression of literature-derived marker genes for human MSCs including NT5E (CD73), THY1 (CD90), and ENG (CD105). NGFR (CD271) and MCAM (CD146) have also been described as canonical MSC markers. D ) CytoTRACE analysis projected onto the MSC UMAP showing the predicted differentiation score, where higher values imply the cell is more primitive. E) Boxplots showing the relative colony forming potential (number of colonies produced by each cell type divided by total colonies from each sample, each data point is one sample) of sorted Fibro-MSC (CD45-CD38-CD235ab-VECAD-PDPN+), THY1+ MSC (CD45-CD38-CD235ab-VECAD-PDPN-LEPR+ CD90+), Adipo-MSC (CD45-CD38-CD235ab-VECAD-PDPN-LEPR+ CD90-), and osteolineage cells (CD45-CD38-CD235ab-VECAD-PDPN-CD56+). P-values were computed as Fibro-MSC vs. all by Dunnett’s Multiple Comparisons test. F) Line plot showing the population doublings over the course of cell culturing for eight passages, where all cells were passaged every 7 days. P-values are computed as Fibro-MSC vs other MSCs by two-way ANOVA. G ) UMAP showing 3,874 endothelial cells from the 12 individuals. SEC – Sinusoidal Endothelial Cells, AEC – Arterial Endothelial Cells. H ) Dot plot showing normalized expression of top differentially expressed genes by adjusted p-value between endothelial subsets, as well as CDH5 , CD34 , and KDR which were manually selected as pan-endothelial markers. I ) MSCs from human bone marrow aspirate reported from De Jong et al., Nature Immunology , 2021 were reference mapped to our scRNA-Seq atlas and the cell type labels were predicted. J ) Human fetal bone marrow mesenchymal cells from Jardine et al., Nature , 2021 were reference mapped to our scRNA-Seq atlas and the cell labels were predicted.

Article Snippet: 20uL of RBC depleted cells were put on ice, while 67% of the remaining cells were used for immunomagnetic CD45 depletion using EasySep Human CD45 Depletion Kit II (STEMCELL # 17898) to isolate non-hematopoietic cells in an unbiased fashion, while 33% of cells were subjected to immunomagnetic CD34 enrichment using EasySep Human CD34 Positive Selection Kit II (STEMCELL #17856).

Techniques: Expressing, Marker, Derivative Assay, Produced, Cell Culture