Imaging-based titration software for GFP-Celltag lentivirus
This repository contains a python script required for titering for GFP-CellTag Virus. For a detailed explanation, please refer to the paper: Guo et. al., CellTag Indexing: a genetic barcode-based multiplexing tool for single-cell technologies, bioRxiv, 2018 (new version under review).
For the CellTag experiment, titration of CellTag-lentivirus is required. Accurate assessment of viral titer can be performed via flow cytometry, we also provide an easy titering method based on the quantification of GFP signal with cell images acquired with a fluorescent microscope. Here, GFP positive cell ratio and CellTag virus titer can be easily calculated by quantifying trypsinized cell images.
The script needs python >=3.6.0 and python libraries listed below. Please install them before running script in this repository. We recommend to install them using anaconda.
numpy >= 1.15.4
scipy >= 1.1.0
pandas >= 0.23.4
scikit-image >= 0.13.0
matplotlib >=2.2.3
seaborn >= 0.9.0
For the detailed information about the CellTag-lentivirus transduction, please refer to the paper; Guo et. al., CellTag Indexing: a genetic barcode-based multiplexing tool for single-cell technologies, bioRxiv, 2018 (new version under review).
Images are acquired after trypsin treatment. Treat cells with trypsin or a reagent for cell dissociation. Please make sure that all cells are completely dissociated. Load cell suspension into a hemocytometer. Cell suspension can also be placed on a cell dish instead of using hemocytometer. Take pictures of the cells using a microscope. This script requires two kinds 2 types of images: an image in acquired in the bright field and an image of GFP fluorescent signal. Please see the example images in - https://github.com/morris-lab/CellTag-Titration/tree/master/test_data. Please use 4x~10x objective lens. The magnification should be small to acquire a large number of cells per image.
Clone the Github repository
git clone https://github.com/morris-lab/CellTag-Titration.git
cd CellTag-Titration
Open jupyter notebook.
jupyter notebook
The repository includes sample imaging dataset. Please run the jupyter notebook with this dataset first: - https://github.com/morris-lab/CellTag-Titration/blob/master/Quantify_images_of_trypsinized_cells_to_estimate_GFP_positive_ratio.ipynb