How I Mapped Brain Cell Changes in Alzheimer's Disease Using Single-Cell RNA Sequencing

Alzheimer's disease affects over 55 million people worldwide, yet the precise molecular changes happening inside individual brain cells remain poorly understood. I wanted to dig into that question - not at the tissue level, but at single-cell resolution. So I built a full scRNA-seq analysis pipeline in Python using Scanpy, working with a publicly available dataset of 63,608 nuclei from human prefrontal cortex tissue (sourced from CZ CELLxGENE). The donors spanned three Braak stages: 0 (cognitively normal), 2 (early Alzheimer's), and 6 (severe Alzheimer's). Here's what I found and how I found it. The Dataset The data came from a study on the molecular characterisation of selectively vulnerable neurons in AD. It covers the superior frontal gyrus, a prefrontal region known to be hit hard by neurodegeneration - and includes seven major brain cell types: Glutamatergic neurons GABAergic neurons Oligodendrocytes OPCs (oligodendrocyte precursor cells) Astrocytes Microglia Endothelial cells 31,997 genes. 63,608 cells. Three disease stages. A lot to work with. The Pipeline 1. Quality Control No dataset is clean out of the box. I filtered cells to keep only those with between 200 and 6,000 detected genes, and excluded anything with more than 20% mitochondrial gene content (high mitochondrial reads usually signal a dying or damaged cell). This removed around 2,809 low-quality cells. 2. Normalisation Library sizes were normalised to 10,000 counts per cell, followed by log1p transformation, standard practice that makes cells comparable regardless of how deeply they were sequenced. I then identified 5,607 highly variable genes to focus the downstream analysis. 3. Dimensionality Reduction PCA (50 components) → neighbourhood graph (10 neighbours, 20 PCs) → UMAP embedding. The UMAP is where the biology starts to become visible. All seven cell types separated into distinct clusters, with clear separation between neuronal subtypes and glial populations. 4. Differential Expression For t