Published May 22, 2026
| Version v1
Dataset
Open
Single-cell RNA-seq and qPCR evaluation of dissociation-induced biases in Gfap+ reactive astrocytes across middle cerebral artery occlusion and tauopathy models
Authors/Creators
-
1.
Czech Academy of Sciences, Institute of Biotechnology
- 2. Institute of Biotechnology, Czech Academy of Science, v.v.i.
Description
Dataset Description
This repository contains processed single-cell RNA sequencing (scRNA-seq) data generated from astrocyte enrichment experiments using three different isolation protocols.
These data support the publication titled: "Tissue dissociation workflows influence reactive astrocyte recovery in single-cell transcriptomics" (DOI: 10.1007/s12035-026-06278-w).
The raw data used as baseline for processed data, which are available in this repository, are publicly available on NCBI GEO and code used for processing is available on GitHub.
Experimental Overview
Capturing dynamic, morphologically complex cell states such as reactive astrocytes via standard single-cell workflows is heavily restricted by tissue-dissociation biases. To address this challenge, we developed and benchmarked an optimized, low-shear tissue processing workflow (WorthGentle) (a step-by-step protocol is available on Protocol.io.
The results were compared between protocols:
- WorthGentle – an optimized in-house protocol utilizing papain-based tissue dissociation, wide-bore pipette homogenization, and low-speed (60 x g) ovomucoid separation to isolate intact cell.
- Miltenyi – a standardized commercial protocol utilizing automated papain-based dissociation, and high-speed (3000 x g) density-gradient centrifugation for debris removal.
- WorthMech – an in-house high-shear control variant identical to WorthGentle, but utilizing standard-bore pipette tips to isolate the specific impact of physical shear stress.
Samples - cortical tissue was collected from female and male mouse models across three experimental groups:
- control - C57BL/6J mice
- permanent middle cerebral artery occlusion - C57BL/6J mice with tissues harvestd 7 days post-stroke
- tauopathy - transgenic mice expressing human P301S tau [Tg(Thy1-MAPT*P301S)2541Godt]
Data processing
The processed data provided in this repository are divided into two experiments:
- scRNA-seq - 1DP_01_seurat_annotated_all_cells.rds (all obtained cells), 1DP_02_seurat_annotated_astro.rds (astrocyte subset)
- single cell PCR (validation experiment) - singlecellPCR.zip