Single-cell Herpes simplex virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

Single-cell analyses of viral infections reveal heterogeneity that is not detected by traditional population-level studies. This study applies drop-based microfluidics to investigate the dynamics of HSV-1 infection of neurons at the single-cell level. We used micron-scale Matrigel beads, termed microgels, to culture individual murine Superior Cervical ganglia (SCG) neurons or epithelial cells. Microgel-cultured cells are subsequently encapsulated in individual media-in-oil droplets with a dual fluorescent-reporter HSV-1, enabling real-time observation of viral gene expression and replication. Infection within drops revealed that the kinetics of initial viral gene expression and replication were dependent on the inoculating dose. Notably, increasing inoculating doses led to earlier onset of viral gene expression and more frequent productive viral replication. These observations provide crucial insights into the complexity of HSV-1 infection in neurons and emphasize the importance of stud..., The microfluidic device containing hydrogel microgels with infected mature neurons were placed in a microscope stage top incubation chamber at 37°C. Images in Phase Contrast/YFP/RFP were taken every 15 min for 16 h. 
Percentage of YFP Positive Cells. To quantify the percentage of YFP positive cells, end-point images were analyzed using Fiji ImageJ. Cells were found in brightfield. The background intensity of the YFP channel was subtracted from the image, and the percentage of YFP positive cells was quantified. Experiments were repeated in triplicate. Cell counts for Vero cells embedded in-microgels ranged from 79 -280 cells/replicate, for Vero cells grown on-microgels from 165 - 390 cells/replicate, for Vero cells in-suspension from 228 - 456 cells/replicate, and for SCG neurons from 84 - 209 cells/replicate. Multiple comparison tests and one-way ANOVAs were performed in Matlab.
The Timing of YFP and RFP Detection. To quantify the timing of YFP and RFP detection, time-lapse images were ..., , # Single-cell Herpes Simplex Virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

## Description of the data and file structure

1.) The dataset titled \"Infection_percentage_raw_data_2-20-24.xlsx\" describes the infection condition, trial number, and number of YFP positive and negative cells counted.

2.) The dataset titled \"Raw_Timelapse_Data_2_27_24_V2.xlsx\" is the raw fluorescence data of the infected cells, organized by by the date of the experiment, the cell type analyzed, the inoculation concentration, the fluorescence channel (YFP or RFP) analyzed, the time point at which the image was taken, and the cell number analyzed. Data has not been background subtracted. Time is in hours post infection (hpi). NA values are timepoints where images were not acquired.