Single Cell NGS, 10X genomics
The CFG offers multiple workflows for single cell gene expression analysis at biosafety level 2 (MLII). The most recent acquisition is the 10X Genomics® Chromium controller, which offers a well-established and standardized platform for single cell analysis.
The Chromium platform typically is loaded with 10K viable cells and returns about 40% to 60% of this input, depending of the quality and robustness of the cells. Cell viability is thus essential for a successful single cell experiment. Sample preparation infra structure and cell viability testing, and 10x Genomics Chromium controllers, are fully operational and available at both the AMC and the VUmc locations, reducing cell viability loss due to transport to a minimum.
Single cell transcriptomics is the most common application for single cell analysis. This chemistry is based on poly-dT capture of mRNA molecules. Also alternative applications are possible using the Chromium controller. Currently we are implementing workflows for single cell ATAC sequencing and single nuclei sequencing. Sequencing the single cell libraries is performed at our (standard) NGS service .
sc-seq and protein detection: Cite-seq
Applying the Total-seq technology of Biolegend® combined with 10x Genomics expression solutions provides also the possibility to combine your single cell analysis with the detection of (surface) proteins.
Several of such so-called CITE-seq panels are available, among others oncogenic and immuno protein profiling panels. Custom prepared oligo tagged antibodies can however also be ordered.
Next to labeling your cells for surface markers, that may resemble classical histochemistry, Total-seq can also be applied for pooling samples. The latter methodology is called cell hashing and is often used when the cell heterogeneity is expected to be low or in order to reduce costs at a maximum (pilot).
Cell hashing is based on oligo (sample specific sequence) tagged antibodies for abundantly and general expressed surface proteins. For both CITE-seq and cell hashing we recommend to use Biolegend® version B or C reagents which are optimized for respectively 3’-expression version 3.1 or 5’-expression version 1.1 chemistry of 10x Genomics. Total-seq applications of Biolegend® are also offered for in particular mice.
Alternative options and consultancy
The Core Facility Genomics can also be your consultant for alternative applications regarding single cell technologies, although these might be not directly available at our laboratory.
We are happy to inform all customers about alternative methodologies such as spatial transcriptomics (e.g. 10x Genomics Visium or Nanostring’s digital spatial profiling) or Fluorescent Activated Cell Sorting based single cell sequencing.
Go to contact and pricing