Example Multiplexed Error-Robust Fluorescence In Situ Hybridization (MERFISH) data from the mouse colon
Data files
Feb 04, 2025 version files 79.28 GB
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example_merfish_data.zip
79.28 GB
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README.md
3.64 KB
Abstract
Multiplexed Error-Robust Fluorescence In Situ Hybridization (MERFISH) is a massively multiplexed single-RNA-molecule imaging technique capable of spatially resolved single-cell transcriptomic profiling of thousands of genes in millions of cells within intact tissue slices. Initially introduced for brain tissues, MERFISH has since been extended to other tissues, where rapid RNA degradation during the preparation process can pose challenges. The companion paper to these example data outlines the application of MERFISH in one such challenging tissue, the mammalian gastrointestinal tract. The example data contained here represent raw MERFISH images and associated metadata necessary to illustrate the analysis pipeline that identifies RNAs from such data.
README: Imaging the intestinal transcriptome with Multiplexed Error-Robust Fluorescence In Situ Hybridization (MERFISH)
Paolo Cadinu
Jeffrey R. Moffitt
Boston Children's Hospital, 2025
Description
This repository contains a subset of a raw MERFISH measurement of the mouse gastrointestinal tract. The images, metadata, and example scripts are designed to illustrate the use of an existing analysis pipeline for MERFISH raw data decoding. These data are provided as a companion dataset to the detailed instructions and protocols provided in Cadinu, Yang, Xu, et al., Current Protocols 2025.
Main folder
This folder contains a set of raw image and metadata files derived from a MERFISH measurement.
-- .tif files represent image stacks taken from individual fields of view at different hybridization rounds and saved on one of two different cameras
-- .xml files contain metadata associated with the hardware settings used to collect the image stack
Settings folder
This folder contains files that describe the MERFISH measurement.
data organization.csv
This file details the organization of the raw data. It contains the following fields:
- bitName: Identifier for the readout oligo probe
- imageType: The base name of the image file
- imageRegExp: Regular expression defining the file name, constructed using the imageType, field of view number, and imaging camera ID
- bitNumber: Numeric identifier for the readout bit
- imagingRound: Hybridization round, ranging from -1 to 13
- color: Color channel associated with the image
- imagingCameraID: Identifier for the imaging camera
- frame: List of acquired frames
- zPos: List of z-stack plane positions relative to the coverslip surface, expressed in microns
- fiducialImageType: Analogous to imageType, but specific to fiducial beads
- fiducialRegExp: Regular expression for fiducial bead file names
- fiducialImagingRound: Hybridization round for fiducial beads
- fiducialFrame: List of acquired frames for fiducial beads
- fiducialColor: Channel associated with fiducial beads
- fiducialCameraID: Camera ID used for fiducial beads
Fibl2v1E1_codebook.csv
This file includes a list of genes and blanks present in the Fibl2v1E1 library, along with their corresponding binary barcodes. It contains the following fields:
- name: The gene name
- id: The Ensembl isoform ID
- barcode: The binary barcode associated with each gene
Transforms.matb
This file contains a set of microscope-specific geometrical transformations used to correct the chromatic aberration in the X-Y planes between the channels used to acquire the MERFISH signals. It is stored in a format accessible by the analysis code.
FPKM.matb
This file provides the abundance of all genes present in the library, as measured by bulk RNA-seq. It is stored in a format accessible by the analysis code. It includes the following fields:
- FPKM: abundance expressed in FPKM
- geneName: gene name
Example Script folder
Code in this folder is designed to be used with public analysis code stored here: https://github.com/ZhuangLab/MERFISH_analysis
startup.m
When placed in the directory from which MATLAB is launched, MATLAB will run this script. It will configure the necessary paths to all needed analysis. NOTE: This script must be updated to reflect paths specific to each computer on which it is run.
MERFISH_Decoding.m
This MATLAB file provides the complete routine for decoding a MERFISH dataset.
plot_barcodes_space.py
This Python script is used to visualize all the decoded barcodes in their spatial positions.