SeqFISH Pack

Applications

seqFISH is a highly sensitive technique that accurately detects low-copy number genes often not detected with single-cell RNAseq or immunostaining. Additionally, reverse transcription or PCR amplification often biases quantification in RT-PCR and RNA sequencing. Because seqFISH can be applied to any tissue type without preselecting genes, it enables the unbiased discovery of novel genes relevant to certain biological phenomena.

• Different methods of fluorescence in situ labeling:  seq-FISH, MER-FISH, seqFISH+, HCR-FISH
• Proteomics and spatial-omics applications
• Identify novel cell types
• Map genomic organization
• Map nuclear architecture
• Cell trajectories analyses
• Subcellular localization of transcripts and proteins
• Ligand-receptor pairs analyses
• Multiplex over 10,000 molecules for transcriptome and proteome imaging
• Cell-cell communication and signaling studies
• Tissue microenvironment impact on cell state changes and developmental trajectories
• Complex multicellular biological systems assays
• Study of complex biological phenomena
• Measure single cells’ phenotypic and genomic states in their respective spatial positions.
• DNA paint

Spatial transcriptomics principle

seqFISH generates accurate in situ quantification of mRNA levels [1]

SeqFISH and MERFISH use probe detection for single-cell spatial transcriptomics [1][2][3].

At first, in situ hybridization is done with one set of fluorescent FISH probes and a labeling dye. DNase is then used to remove the fluorophores, and the mRNA is hybridized with the identical FISH probes again but with a different labeling dye. The several rounds of hybridization and other dyes allow the barcoding of several genes in a single cell [4].

SeqFISH+ is the improved seqFISH technique ideal for spatial and biological processes studies of cells. It combines seqFISH with a confocal microscope generating super-resolution imaging and multiplexing of 10,000 genes in a single cell [5].

Multiplexed Error Robust Fluorescence In Situ Hybridization (MERFISH) is the improved single-molecule Fluorescence In Situ Hybridization (smFISH). The method massively parallelizes and simultaneously spatially identifies hundreds of thousands of RNA species. In addition, this method detects errors that can then be corrected in an error-robust way thanks to the use of some unassigned binary barcode. This is the main difference compared to seqFISH, which is coded in a color sequence [6].

Lab-on-chip technologies and microfluidic platforms improve seqFISH and MERFISH methods by reducing the cost and the experiment time, providing automation to the process, and improving reproducibility [7].

1. Shah, Sheel & Lubeck, Eric & Zhou, Wen & Cai, Long. (2016). In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus. Neuron. 92. 342-357.
2. Raj A, van Oudenaarden A. Nature, nurture, or chance: stochastic gene expression and its consequences. Cell. 2008;135:216–226.
3. Asp, M., Bergenstråhle, J., Lundeberg, J., Spatially Resolved Transcriptomes—Next Generation Tools for Tissue Exploration. BioEssays 2020, 42, 1900221.
4. Lubeck, E., Coskun, A., Zhiyentayev, T. et al. Single-cell in situ RNA profiling by sequential hybridization. Nat Methods 11, 360–361 (2014).
5. Eng, CH.L., Lawson, M., Zhu, Q. et al. Transcriptome-scale super-resolved imaging in tissues by RNA seqFISH+. Nature 568, 235–239 (2019).
6. Moffitt, J R, and X Zhuang. “RNA Imaging with Multiplexed Error-Robust Fluorescence In Situ Hybridization (MERFISH).” Methods in enzymology vol. 572 (2016): 1-49.
7. Rodriguez-Mateos, P., Azevedo, N.F., Almeida, C. et al. FISH and chips: a review of microfluidic platforms for FISH analysis. Med Microbiol Immunol 209, 373–391 (2020).

A typical pack contains:

• OB1 flow controller
• A flow sensor (MFS or a Coriolis flow sensor for the best performance)
• One or two 12:1 MUX distribution valves depending on the number of dyes you want to inject
• Tubings and luers
• Several Eppendorfs or Falcon reservoirs
• Microfluidic chips* advice and recommendation
• Automation and control software
• A user guide

*Chips are available in several materials, heights, widths, lengths, and shapes, with different numbers of inlets/outlets, and are compatible with different FISH methods. We can advise you on different chips to fit this pack.

Build your pack in three quick and easy steps:

1. Talk to our experts 
2. Tell them what you need to accomplish
3. Our team will advise a pack tailored to your needs