Plate-based single-cell transcriptomics
SORT-seq is a single-cell sequencing platform first published in 2016 by Muraro et al. and has since been cited over 700 times. SORT-seq is a partially robotized version of the lauded CEL-seq2 protocol.
See the publications that use SORT-seq here.
SORT-seq uses FACS to sort single cells into the wells of a 384-well cell-capture plate. Each well in the plate contains barcoded primers and other reagents. After barcoding, double-stranded cDNA is generated. Then we pool the contents of all wells, amplify the RNA, and prepare the sample into an Illumina sequencing library.
This sorting step makes it possible to analyze live single cells, population of interest, and cells too large for the 10x Genomics platform.
What you can do with SORT-seq
SORT-seq's flexible plate-based system makes it possible to screen small numbers of cells from many different samples without skyrocketing costs.
By combining single-cell flow cytometry data with transcriptomics data with strategies like GateID, we can find novel FACS gating strategies for desired cell types.
Key benefits of SORT-seq
- Modular and flexible due to the use of plates
Working with plates allows you to start small and upscale later. We can start by processing and sequencing one plate per condition. Once the data from the first plate looks good, we can go on to process the rest of the plates. This provides you with tremendous flexibility in scaling up your experiments.
- Compatible with all sample types
SORT-seq works even with rare or low quantities of cells. As long as you can fill at least part of a plate with FACS sorting, you can generate single-cell sequencing libraries. Also, SORT-seq can handle cells of all sizes. Cells that are too big for the 10x Genomics system (e.g. cardiomyocytes) can be analyzed with SORT-seq without any issues.
Step 1: FACS sorting
Begin by ordering our cell capture plates, which we ship worldwide. You then sort your single-cells into the 384-well cell-capture plates and send the plates back to us for processing. If you don't want us to process all the plates at once, just let us know which ones to start with.
Step 2: Plate processing
Once we've received your plates, we'll process them using the SORT-seq protocol. After barcoding the mRNA of your single-cells, we'll pool the material from one plate to generate one sequencing library per plate.
Step 3: Sequencing and data analysis
We'll send the sequencing libraries to one of our partners for sequencing. When we get the data back, we'll perform a preliminary data analysis, then send the data back to you.
"The service of Single Cell Discoveries guarantees a smooth process between customer and company. In case of problems or questions, the scientists reacted competently and immediately, so that answers could be found quickly and flexibly. The staff is very friendly in dealing with the customer. Many thanks for the great cooperation."
Prof. Dr. Stella Autenrieth & Susanne Pfrommer
German Cancer Research Center
As with all of our services, we provide ongoing support from our single-cell sequencing specialists. For each SORT-seq project, we provide the following:
- Regular email updates during plate processing
- Raw sequencing data (FASTQ files)
- A QC report per plate
- Mapped count tables for downstream analysis
- Preliminary clustering analysis with the Seurat pipeline.
- Preliminary differential gene expression analysis
- A follow-up meeting to discuss your questions and further data analysis needs
Can I use SORT-seq?
Use our single-cell sequencing buying guide to find out if SORT-seq matches your project.
Learn more about the results from our services - from a client perspective.
Colorectal Zebrafish Xenograft Model
Champalimaud Foundation | This research, conducted at the Champalimaud Foundation in Lisbon, is focused on the immune response after implantation of human cancer cells in zebrafish. The cells in the implanted tumors were characterized using SORT-seq. Here, we explain how SORT-seq contributed to this remarkable study. Cancer immunoediting is a process in which the immune…Continue reading
We'll need a minimum of two plates for each sample or condition. This ensures there is always a technical back-up plate available in case something goes wrong with the shipment or processing.
This mostly depends on your biological question. For cell type identification in a sample that contains only a few cell types, 2-3 plates are probably enough. If you are interested in subtle changes in the transcriptome, or if you are looking for rare cell populations, we may need more plates.
Yes, you can first process one plate (or set of plates) and then continue with the rest later on. Any minor technical batch effects should be negligible, especially if the biological differences between populations of cells in your sample are clear.
The first step is to reach out to us to set up a meeting with one of our specialists. During this meeting we'll discuss your experiment and help you define a pilot experiment. Next, you order the cell capture plates via our website. Once you have sorted single-cells into the plates, you ship them back to us for processing.
While this is possible in principle, it will likely require some batch effect correction. Different technologies will yield slightly different results due to technology-specific technical quirks and other biases. However, with batch effect correction and a clear biological difference between the different populations in your samples, it should be possible to mix with data from other platforms.
Not sure if SORT-seq is right for your project? Explore our other services.
Download pricing information
Ready to proceed? Use the form below to download pricing information on our SORT-seq services.
Head of Business Development
Not sure what technology is right for you?
Contact us for advice.