InquiryGene expression is both temporally and spatially specific, and by taking samples at different times, single-cell transcriptome sequencing techniques can resolve changes in cell type and gene expression in the temporal dimension. However, the prerequisite for single-cell sequencing is that tissues must be mechanically separated or enzymatically digested into single-cell suspensions. This process inevitably loses information about the original location of cells in the tissue and breaks the communication network between cells, which makes it difficult to obtain information about the cell composition and gene expression status of different regions in the tissue, as well as the differential gene expression between different functional regions. Existing in situ expression mapping is mainly achieved by techniques such as reporter gene or in situ hybridization, but these methods are more difficult to implement and have low throughput, which limits the application of multiple samples. In contrast, spatial transcriptome technology can efficiently detect gene expression at spatial locations in tissues.
Our Spatial Transcriptome Sequencing Service of Frozen Samples
CD Genomics provides 10X Genomics' high-throughput Visium spatial gene expression solution, which can detect total mRNA in complete tissue sections, combine the spatial information of total mRNA with morphology, and map the location of all gene expression to obtain a complete gene expression profiles; the determination of distinct cell populations while preserving spatial location provides important information on the relationship between cell function, phenotype, and location in the tissue microenvironment.
Technical Principle
Frozen tissue sections are placed in the capture area of the Visium chip with spatial barcode. After HE staining and imaging, through tissue permeabilization, intracellular mRNA is released and captured by the probe with oligo-dT on the chip. The captured mRNA is reverse-transcribed to obtain the cDNA containing the spatial barcode sequence. After library construction and sequencing, the sequence of the mRNA transcription can be mapped to the original position in the tissue, thereby obtaining the location information of the gene expression. The core of the Visim spatial transcriptome is the chip: there are 4 capture areas on the chip and each capture area contains about 5,000 spots, each of that can capture 1 to 10 cells. Each spot contains millions of capture probes that can bind to mRNA, and each probe contains a unique barcode (spatial barcode) to mark the spatial position information.
Principle of spatial transcriptome gene expression chip (10xgenomics.com)
Spatial Transcriptome Sequencing Experiment Flow (10xgenomics.com)
Advantages of Spatial Transcriptomics Technology
It is a new direction in the field of transcriptomics research and a new approach in studying cell heterogeneity.
(1) Accurate localization: the probe effectively locates the spatial location of RNA in tissues and provides real-time understanding of the natural state of the transcriptome in tissues.
(2) Simple and easy to use: the chip design is simple and easy to operate, highly reproducible, and can quickly obtain high-resolution spatial transcriptome information.
(3) Wide applicability: effectively used in various biological fields such as developmental biology, tumor biology, brain neuroscience, and plant research.
Why Choose Us
1. Personalized solutions: The research needs of spatial transcriptome sequencing vary widely. Our professional scientific research team provides one-to-one project suggestions according to customer needs and customizes spatial gene expression solutions for customers.
2. Tissue preservation solution: Spatial transcriptome technology requires high sample quality. Fresh tissue samples need to be frozen and embedded immediately to better maintain the quality of sample RNA, thereby ensuring the accuracy of the experiment. We developed a tissue preservation solution by ourselves. Fresh tissue samples were isolated for 48 hours at 4°C, and the results of cell viability and cell morphology were not significantly affected. Effectively solve the adverse effects of sample collection, storage and transportation, and freezing and embedding processes.
3. Preparation of high-quality tissue sections: Our scientific research service team has accumulated experience in a large number of project samples, and has explored methods to prepare high-quality sections for various types of tissues.
4. Comprehensive bioinformatics analysis process and personalized data analysis.
Sample Type
OCT-embedded samples that can be used for tissue sections (bone tissue and other samples are not available)
Analysis Process
Application
Tumor heterogeneity;
Histomorphology;
Tissue development mechanism;
Response to therapeutic intervention;
Biomarker discovery;
Cell Atlas.
Q&A
Q1: Can quick-frozen tissues be mailed?
A: No, it is recommended that fresh samples be embedded with OCT.
Q2: Which method of sample preparation is preferred?
A: Fresh tissue materials are used for sample preparation, and the two methods of "freezing first and then embedding" and "embedding first and then freezing" are both acceptable.
Q3: Why is isopentane added when preparing frozen samples?
A: The boiling point of liquid nitrogen is very low. Tissues will boil when placed directly in liquid nitrogen, which will easily change the internal shape of the tissue or even fragment it, destroying the tissue structure. The boiling point of isopentane is not so low. Putting the tissue in after pre-cooling will not cause boiling, so the shape of the tissue will not change.
Q4: The tumor heterogeneity is very large, and I am very worried that the selected area is not representative. What should I do?
A: It is recommended to use immunohistochemistry/HE staining to screen the tissue area of interest.
Q5: Can bioinformatics analysis be associated with high-throughput single-cell RNA-Seq sequencing?
A: Yes, use seruat software for association analysis to achieve high-throughput single-cell data supplementary annotation spatial transcriptome results.
