Alterations in Germinal Center Formation and B Cell Activation during Severe Orientia tsutsugamushi Infection in Mice [Nanostring]

We performed NanoString analysis on total RNA extracted from spleens of mice infected with Orientia tsutsugamushi to investigate the transcriptomic signature of these cells throughout infection. Mouse spleen samples were collected from mock (D0) and D8 infected mice, and stored in RNALater (Ambion, Austin, TX) until extraction was performed. Total RNA was extracted by using the RNeasy Mini kit (Qiagen), and total RNA samples (3 mice/group, 200 ng/sample in ribonuclease-free water) were processed at the Baylor College of Medicine Genomic and RNA Expression Profiling Core (Houston, TX). We utilized the comercially available Mouse Immunology Panel NanoString kit. Gene expression profiling was performed by using the nCounter platform and the Mouse Immunology Panel NanoString kit, which comprised 561 genes and 14 housekeeping genes (NanoString Technologies, Seattle, WA). Raw data were then normalized and analyzed by ROSALIND (https://rosalind.onramp.bio/), with a HyperScale architecture developed by ROSALIND, Inc (San Diego, CA).