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Single Cell Sequencing Service

Wuhan BioBank is one of the earliest institutions engaged in the development and service of single-cell  sequencing technology in China, and became the only officially certified single-cell sequencing co-construction laboratory in Central China by 10x Genomics in April 2021.

The platform has successfully carried out single-cell  sequencing for  thousands of samples from multiple species and tissues including human, mouse, bovine, fish and wheat, as well as spatial transcriptome sequencing of different tissues from human and mouse. The platform team has accumulated plenty of project experience and expertise and has supported a number of clients to publish high-level articles.

Single Cell Sequencing
Spatial transcriptome sequencing
Single Cell Multiome ATAC + Gene Expression
Single Cell Multiome ATAC + Gene Expression combines ATAC with transcriptome sequencing to identify complex cell populations through integrated analysis of epigenomic and gene expression profiles and detect cellular heterogeneity to uncover hidden information. Combining the discovery of regulatory elements with gene expression could help to explore the gene regulatory interactions that drive cell differentiation, development and disease. Sample types: animal tissues, single cell suspensions, nuclear suspensions, cell lines.   Experimental process 单细胞多组学ATAC+基因表达实验流程图 Analysis process Flow Chart of ATAC+Gene Expression Analysis in Single Cell Multiomics 　　 Application directions a. Biomarker discovery; b. Cell lineage and developmental program tracking; c. Cell heterogeneity and detection of rare cell populations; d. Gene regulatory networks; e. Response to therapeutic interventions.   Product advantages a. Efficiently distributing 500-10,000 nuclei per lane, analyzing up to 80,000 nuclei per run ; b. Extensible to run up to 8 samples in parallel; c. Nuclei recovery up to 65%; d. High sensitivity; e. Low microfluidic double rate (less than 1% per 1,000 nuclei); f. Suitable for studies with cell lines, primary cells, fresh and frozen samples.

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Single Cell ATAC
Single-cell ATAC-seq is to study chromatin accessibility at the single-cell level and is mainly used for the detection of transcriptional regulatory sequences, which can be combined with single-cell RNA-seq to analyse all actively transcribed regulatory sequences in the genome. This technique has been used in many research areas such as tumor heterogeneity studies, gene regulatory network analysis, cell lineage tracing, biomarker discovery, etc. ATAC-Seq (Assay for Transposase Accessible Chromatin with high-throughput sequencing) uses Tn5 transposase to cleave nuclear chromatin regions which is open at a specific space-time and then adds primers for high-throughput sequencing to obtain the regulatory sequences of all active transcripts in the genome at that specific space-time condition.   ATAC-Seq schematic diagram   The single cell ATAC-Seq technology relies on the microfluidic technology of the 10x Genomics platform, combining characteristics of different cells of sequence tags distinguished population with ATAC-Seq technology to build chromatin accessibility profiles for thousands of single cells. Sample types: cell lines, primary cells, fresh and frozen tissues, etc. Schematic diagram of 10x Genomics single-cell ATAC-Seq   Sample type Cell lines, primary cells, fresh and frozen tissues, etc. Experimental process 10x Genomics单细胞ATAC-Seq实验流程   Analysis process Flow chart of single-cell ATAC analysis 　　 Application directions a. Studying cell heterogeneity; b. Exploring biomarkers; c. Defining cell types; d. Building gene regulatory networks.   Product advantages a. Detecting open chromatin in transcriptional regulatory regions of single cells; b. Each channel can hold 500~10000 nuclei; c. Nuclei capture rate up to 65%; d. Suitable for the study of cell lines, primary cells, fresh and frozen samples; e. Using the official 10x software, single cell gene regulation can be deeply analyzed.

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Single Cell Immune Profiling
In 2015, 10x Genomics released the Chromium Single Cell System platform based on microfluidic and oil-drop wrapping technologies, enabling high-throughput single-cell transcriptome and single-cell V(D)J sequencing. This technology not only allows perfect matching of TCR/BCR duplexes to obtain TCR/BCR duplex composition, but also could be detailed into the single cell level while obtaining 5' end transcriptome information. New released Single Cell Immunoprofiling of 10x Genomics uses microfluidic chip to prepare single cell systems, selecting universal primers for the 5' end-joins and nested primers for the immune molecular constancy region for V(D)J enrichment to enable full-length sequencing of paired heavy and light chains (B cells) or alpha and beta chains (T cells) at the single cell level. Full-length sequencing is performed, thus providing an efficient technical platform for comprehensive and systematic studies of immunomic libraries, and is important for the study of the molecular mechanisms of disease onset and development of disease. Sample types Animal tissues, other body fluids such as blood, single cell suspensions, cell lines, etc.   Experimental process Experimental flow of 10x Genomics single cell immune group library Analysis process Flow Chart of Bioinformatics Analysis of Single Cell VDJ Immunogroup Bank   Application directions a. Identification of antibodies and discovery of new antibodies; b. Revealing the development of Ig libraries and mapping global Ig libraries; c. Studying infectious diseases, such as the evolutionary spectrum of antibodies in the development of infections; d. Studying the propensity for V(D)J recombination in immune disorders and to search for biomarkers of immune disorders; e. Early detection of cancer, and studying markers for the diagnosis of cancer progression and recurrence, and studying tumor immune mechanisms. Product advantages a. Identification and characterization of rare cell types and biomarkers; b. Analysis of tissue micro environment, disease progression and drug immune response; c. Large-scale antibody and TCR discovery against neoantigens d. Characterizing the immune response to infection by measuring immune cell phenotypes; e. Reduce single cell sequencing costs through large-scale single-cell V(D)J+ expression profiling sequencing; f. Provide flow cytometry sorting services to enable sorting for immune cells with purposes.

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Single Cell Gene Expression
Single cell RNA sequencing (scRNA-seq) is a new technology for high throughput sequencing of mRNA at the single cell level. The principle is to dissociate the sample tissue into  single cell suspension and then carry out high-throughput sequencing of the minute amounts of  mRNA in single cells after efficient amplification. Single cell transcriptome sequencing can solve the problem of cell heterogeneity that is obscured by conventional RNA-seq and help to discover new cell types, which is important for neurobiology research, immunology research and developmental biology research. Sample types: animal and plant tissue, blood and other body fluids, cell lines, single cell suspensions, plant protoplast suspensions,  animal and plant cell nucleus suspensions and so on.   Experimental process 　　10x Genomics单细胞转录组实验流程   Analysis process Flow Chart of Single-cell Transcriptome Bioinformatics Analysis 　　 Application directions a.Large-scale cell atlas construction; b. Identification of rare cell types & cell subpopulation refinement; c. Tumor heterogeneity and tumor microenvironment research; d. Immunological research; e. Cell development and differentiation research. 　　 Product advantages a. Real single cell level sequencing with high cell capture efficiency; b. The team is experienced in sample dissociation and could accept a wide range of sample types; c. One-stop service for sample processing, sorting, loading, sequencing and analysis; d. Short project period and wide range of technical applications.

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Spatial Gene Expression for FFPE
Considering traditional spatial transcriptome sequencing is limited to fresh frozen tissue sections, 10X Genomics introduces the new Visium spatial gene expression solution for FFPE samples (formalin-fixed paraffin-embedded), which breaks the barriers to obtain spatial gene expression information from FFPE tissue samples and get highly sensitive and specific expression and spatial location information of relevant genes from FFPE samples. FFPE sample gene expression is a pioneering solution that complements traditional pathology-driven analysis, enabling true discovery in a morphological context and further expanding the possibilities of spatial biology research.

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Spatial Gene Expression for Fresh Frozen
Principle of the technique When a frozen tissue slice is placed in the capture region of a Visium chip with spatial barcode and has been HE stained and imaged, the tissue is permeabilised and the intracellular mRNA is released and captured by a probe with oligo-dT on the chip. The captured mRNA starts reverse transcription and the resulting cDNA contains spatial barcode sequences. After constructing a library and sequencing, the mRNA transcribed sequences can be mapped to their original location in the tissue, thus obtaining information about the location of gene expression. The core of the Visim spatial transcriptome is the microarray: the formal library is constructed with four capture regions, each containing approximately 5,000 spots, each capable of capturing 1-10 cells, with a distance of 100 μm between spots and spot centroids. Each spot contains millions of capture probes that bind to mRNA, and each probe contains a unique barcode (barcode). Principle of space transcriptome gene expression chip   Sample type: OCT-embedded samples that can be used for tissue slicing (not available for samples such as bone tissue)   Experimental process   Experimental flow of space transcriptome sequencing   Analysis process 　　Space transcriptome sequencing analysis process Application directions a. Tumor heterogeneity; b. Histomorphology; c. Tissue developmental mechanisms; d. Response to intervention; e. Biomarker discovery; f. Cellular mapping. Product advantages a. Comprehensive understanding of disease complexity b. Discovery of new biomarkers and identification of new cell types and states c. Locating the spatial structure of cellular profiles d. Identify spatio-temporal gene expression patterns

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High throughput sequencing services

From sample processing to information analysis, Wuhan National Human Genetic Resources Library promises a meticulous service cycle and considerate after-sales experience, a strict and complete production management and service system and a large number of technical teams to provide better services and sequencing programs for scientific research experts in medical, agricultural and other multidisciplinary fields.

DNA sequencing services
RNA sequencing services
Epigenetic sequencing service
Macrogenome sequencing
Macrogenomic sequencing aims to sequence the genomes of microbial communities in specific environmental samples in order to analyse the genetic composition and function of microbial communities, to decipher the diversity and abundance of microbial communities, explore the relationship between microbes and their environment, microbes and hosts, and discover and study new genes with specific functions. Macrogenomics is a new approach to study microbial diversity, population structure, evolutionary relationships, functional activity, interactions and relationships with the environment, using the genome of the microbial community in environmental samples as the object of study and functional gene screening and/or sequencing as the means of study. This technique can be used to unravel the diversity and abundance of microbial populations, explore the relationships between microbes and their environment, microbes and their hosts, and discover and study new, functionally specific genes.   technology roadmap：   technical parameter： Sample requirements Sequencing strategy Lead time Sample type: Environmental microbial DNA samples Sample concentration: ≥50 ng/μl Total sample mass: ≥3 μg Sequencing mode: Illumina/MGI/Double-end sequencing Sequencing depth: 5-10G clean data 45 days   Product advantages. a. Technical advantages: PCR-free library construction for more accurate species annotation, and new splicing methods to reduce the false positive rate of gene annotation; b. Customised solutions: In addition to conventional analyses, a number of advanced analyses are available to provide the best solution for your project.

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Whole genome sequencing
Whole Genome Sequencing (WGS) is a genome-wide sequencing method that can decipher all common and rare variants, single nucleotide polymorphisms (SNP), insertion and deletion sites (InDel), structural variants (SV) and copy number variants (CNV), and thus reveal all types of genomic mutations and chromosomal rearrangement events. This will provide guidance for disease mechanism research and drug target screening. Applications include clinical medicine, population genetics, association analysis, evolutionary analysis and so on.   technology roadmap   technical parameter   Technical Parameters Sequencing strategy Lead time Sample type: Genomic DNA Sample concentration: ≥20 ng/μl (Qubit quantification) Total sample mass: ≥3.0 μg (total of 3 library builds, excluding consuming of sample assay, Qubit quantification) Sample purity:  OD260/280 =1.7~2.2 Electrophoresis requirements: clear primary bands, no degradation or light degradation, no serious RNA or protein contamination (latest electrophoresis results of the sample delivery date).   Sequencing mode: Illumina/MGI Sequencing depth: 30X recommended 45 days   Product advantages a. High accuracy: Using the Illumina /MGI platform by which more than a million sequences can be performed in a single run, with accurate counts ranging from several to hundreds of thousands copies; b. Wide detection range: single nucleotide polymorphism (SNP), insertion deletion (InDel), structural variation (SV), copy number variation (CNV) and other variants can be detected, and new variant sequences can be detected compared to microarrays; c. Low cost and less time consuming than whole human genome de novo sequencing.

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Exome sequencing
Whole Exome Sequencing (WES) refers to the use of sequence capture technology to capture and enrich DNA from exome regions for high-throughput sequencing, which can directly identify genetic variants associated with protein function variation. Exome sequencing has been applied to the search for disease-causing and susceptibility genes associated with a variety of complex diseases.                technology roadmap     technical parameter   Sample requirements Sequencing strategy Lead time Sample type: genomic DNA Total sample concentration: ≥20 ng/μl (Qubit quantification) Total sample mass: ≥3.0 μg (total of 3 times of library building, excluding consuming of sample assay, Qubit quantification) Sample purity: OD260/280 =1.7~2.2 Electrophoresis requirements: clear main band, no degradation or mild degradation, no serious RNA or protein contamination. (latest electrophoresis result of the sample delivery date)   Sequencing type: Illumina/MGI PE150. Sequencing depth: germ cell mutations ≥ 100X (10G). Somatic mutations ≥150X (15G)  45 days       Product advantages a. High specificity: Use mainstream kit, have stable capture efficiency, comprehensive exon region coverage and high capture specificity; b. Cost-effective: comprehensive functional annotation of exon regions, which can effectively identify disease- or trait-related regions and save research costs; c. High information efficiency: Compared with whole genome sequencing, it has deeper coverage and better data precision, less interference information and easy to analyse.

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Macro-transcriptome sequencing
Macro-transcriptomics studies the genome-wide transcription of organisms and the regulation of transcription at a holistic level for a specific environment and a specific period of time in a community. Macro-transcriptome sequencing can study in situ the composition of active strains and the expression of active genes in a microbial community of the specific habitat at a specific time and space. In combination with the detection of physicochemical factors, macro-transcriptome can study the differences in the composition of active components between microbial communities in space and time due to the differences in physicochemical and other indicators among the samples. Technical route:     　　Sample requirements： Sample type Sequencing strategy Lead time Sample concentration: ≥100 ng/μl Total sample  mass: ≥4 μg Sample purity: OD260/280=1.8~2.2, OD260/230=1.8~2.2; 28S: 18S≥1. Integrity:RIN≥7 Sequencing mode: Illumina/MGI PE150 Sequencing depth: 6-12G clean data 45 days   Product advantages: A. Technical advantages: the official ribosome removal kit can effectively remove rRNA and greatly improve the utilization rate of effective data; B. Personalized solutions: In addition to routine analysis, it can provide multiple advanced analysis to provide the best solution for your project.

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Long-strand non-coding RNA sequencing
lncRNAs are greater than 200bp in length and do not encode proteins, which are widely found in living organisms. lncRNAs regulate gene expression levels at multiple levels (epigenetic regulation, transcriptional regulation, and post-transcriptional regulation, etc.) and play an important role in life activities. lncRNA sequencing is a research method that uses a specific method to reduce the abundance of rRNAs in samples, during which a library to sequence the enriched lncRNA sequencing would be constructed and almost all the lncRNA information can be obtained at one time,  providing a comprehensive and in-depth analysis of lncRNA classes and functions to get lncRNA related information of specific biological processes (e.g. development, diseases, etc.) quickly and accurately.   Technical route:   technical parameter：   Sample type Sequencing strategy Lead time Sample type: Total RNA Sample concentration: ≥100 ng/μl Total sample mass: ≥5 μg Sample purity:OD260/280=1.8~2.2, OD260/230 = 1.8~2.2 Integrity:RIN≥7 Sequencing mode: Illumina/MGI PE150 Sequencing depth: 10G /12G clean data 45 days   Product advantages. a. Cost-effective: one library can obtain both mRNA and lncRNA information; b.Total solution: one-stop upstream sequencing & downstream functional validation and consulting services; c. Comprehensive services: standard analysis, advanced analysis and multi-omics association analysis are all available.  

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Macro-transcriptome sequencing
Macro-transcriptomics studies the genome-wide transcription of organisms and the regulation of transcription at a holistic level for a specific environment and a specific period of time in a community. Macro-transcriptome sequencing can study in situ the composition of active strains and the expression of active genes in a microbial community of the specific habitat at a specific time and space. In combination with the detection of physicochemical factors, macro-transcriptome can study the differences in the composition of active components between microbial communities in space and time due to the differences in physicochemical and other indicators among the samples.   　　technology roadmap：   　　Reference technology：   Sample requirements Sequencing strategy Lead time Sample type: Environmental microbial RNA samples Sample concentration: ≥100 ng/μl Total sample  mass: ≥4 μg Sample purity: OD260/280=1.8~2.2, OD260/230=1.8~2.2; 28S: 18S≥1. Integrity:RIN≥7 Sequencing mode: Illumina/MGI PE150 Sequencing depth: 6-12G clean data 45 days   　Product advantages: A. High coverage Through different sequencing schemes, almost all the transcriptome expression and structure information of samples can be obtained, and the transcriptome information of low abundance can be obtained at the same time; B. Multi-platform solutions: provide Illumina and Pacbio multi-platform solutions; C. No species restriction: the detection scope covers all microorganisms, animals and plants and human samples.

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Cell-based immune-related assays

The cellular immune analysis service mainly includes two functional modules: the spatial phenotypic analysis of more than 7 colors can be carried out at the pathological tissue level, and the cell phenotypic analysis of more than 20 colors can be completed at the cell level. A variety of mature technical schemes have been established to perfectly link the links of high-throughput flow sorting, single-cell sequencing, SMART-seq2, etc.
The number of analysis samples exceeds one million, and has accumulated rich experience in the fields of tumor genesis and development, new vaccine research and evaluation, micro-environment research and immune function evaluation in the fields of infection and inflammation. Provide customers with one-stop service from project scheme design, sample processing to detection and analysis.

Flow cytometry
Multiple immunofluorescence detection technology
Primary cell/type 3D organ culture
Patch clamp inspection service
Immune cell typing and functional assessment
Flow cytometry (FCM) is a multi-parameter, rapid quantitative analysis and sorting technique for cells or particles in a fast, linear and stable flow state using a laser as the light source. In addition to conventional apoptosis and cycling, it can be applied to immune cell typing, intracellular/extracellular cytokine and phosphorylated protein detection, stem cell phenotype identification, plant ploidy/genome size assessment, and high-throughput single cell sorting for downstream sequencing.

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Multiple cytokine expression assessment
Cytokines, as intercellular signaling molecules, mainly mediate and regulate the immune response and inflammatory reaction, and maintain the stability of the body's immune system by regulating the dynamic balance between immune promotion and immune suppression. The detection of cytokines is of great value for the diagnosis and treatment of certain diseases. Extracellular (serum, cell culture fluid, lavage fluid, etc.) cytokines reflect the level of secretion.     Instrument used: Cytoflex S (4 laser 13 colors), Cytoflex LX (6 laser 21 colors)    Application areas: immunology, hematology, oncology, bone marrow and organ transplantation, antitumor drug research, vaccine research, basic medical research, etc. Project advantages Flow cytometry extracellular factor assay (superior to Elisa, more than 10 cytokines simultaneously in 5 hours can be detected with only 25 μL sample, which greatly saves the amount of sample, time cost and experimental cost, and has high sensitivity and specificity) Lead time： 项目类型 项目内容 服务周期 Detection of secretory cytokine concentrations in body fluids Th1/Th2/Th9/Th17/Th22/anti-inflammatory, pro-inflammatory cytokines (IL-2, IL-6, IL-10, IFN-γ, TNF-α, IL-5, IL-13, IL-4, IL-17A, IL-17F, IL-22, IL-9, CXCL8 (IL-8), CXCL10 (IP-10), CCL11 (Eotaxin), CCL17 (TARC), CCL2 (MCP-1), CCL5 (RANTES), CCL3 (MIP-1α), CXCL9 (MIG), CXCL5 (ENA-78), CCL20 (MIP-3α), CXCL1 (GROα), CXCL11 (I-TAC), etc. ) 1-3 working days Note: The above lead time is under the condition of reagents provided by Party A( the client ). If Party B (the company )'s previous reagents have been ran out ,another 4-5 weeks is needed in addition for purches related reagents .

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High throughput cell sorting
Flow cytometry sorting is based on flow cytometry analysis, which can "pick out" the target cells of interest as the material for subsequent experimental operations such as culture amplification, protein identification, and sequencing. The challenge for sorting is to obtain cells with high viability, purity and recovery in a fast and accurate way.     Instrument used: Moflo XDP (3 laser 10 colors) Application areas: immunology, hematology, oncology, bone marrow and organ transplantation, antitumor drug research, vaccine research, basic medical research, etc. Project advantages. In the case of cellular raw material acquisition based on single cell sequencing, SMART Seq2, RNA Seq and other sequencing, flow cytometry sorting is the best choice for efficient screening of high viability and high purity target cells. Through genome sequencing, single cell sequencing, proteomics and bioconfidence platform, we can achieve one-stop service from raw material to report issuance. Lead time： Project type  Project Content Service period High throughput cell flow cytometry sorting   Stable transfer cell line screening      1-3 working days Antigen-Specific B-cell Screening    1-3 working days 96-1536 well plate single cell sorting 1-3 working days Multi-color, four-way sorting 1-3 working days Sorting of target cells for downstream sequencing, single cell sequencing, proteomics, etc.     1-3 working days  

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Flow cytometry special service
Extracellular vesicles (EV) are considered as potential non-invasive biomarkers in disease diagnosis and mediators in drug delivery with a high degree of heterogeneity. Transcriptome studies of single EVs are valuable to better define EV characteristics, EV subgroups and clinical applications. For EV identification and concentration assessment, it is a critical step for single cell sequencing to be successfully brought online. The use of flow cytometry-based means enables the size-specific localization of EVs, thus allowing the enumeration of integrity vesicles as well as the adjustment of appropriate concentrations for subsequent single-cell sequencing.

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Customized multiplex immunofluorescence panel
Multiplex immunohistochemistry is a tyramine dignal amplification (TSA)-based immunostaining technique that can multiplex multiple molecular markers on a slide. We can provide customized solutions for each project, and quickly achieve quantitative pathology analysis results with up to 9 color multiplex staining  through standardized preparation and high-throughput tissue section panoramic quantitative analysis system .We can perform qualitative, conformal and quantitative analysis of tissues, cells and proteins in the images with the help of post-staining antibody elution technology, spectral signal recognition technology and AI intelligent quantitative pathology analysis software. Finally ,the data results are visualized in the form of figures.

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Ready-to-use multiplex immunofluorescence panel
Product introduction This product is used for tumor immune microenvironment assessment and clinical development and translational research of tumor immune drugs, covering multiple dimensions such as tumor immunotherapy targets and tumor immune cells. Through high specificity multiplex fluorescence staining, spectral imaging and high throughput image analysis technologies, we can accurately capture tumor microenvironment data to empower anti-tumor drug development.     Sample image of product BB02     Service products and cycle Product Number     Indicators Tissue Phenotype Data Analysis Lead time BB01 CD3、CD20、CD68、 CD163、Ki67、pan-CK、DAPI T cells, B cells, M1 macrophages, M2 macrophages Standard analysis. 1. tissue partitioning: tumor and interstitial. 2. cell typing: defining the different cell types separately and calculating the number, density and percentage of cells. Extended analysis. 1. Evaluation of expression indicators TPS and CPS; 2  statistics on the number, density, and percentage of immune checkpoint molecules expressed in different types of cells. 3. spatial analysis.   10 working days BB02 CD4、CD8、FOXP3、 PD-1、PD-L1、pan-CK、DAPI Helper T cells, killer T cells BB03 CD8、CD68、CD56、 PD-1、PD-L1、pan-CK、DAPI Treg, immune checkpoint killer T cells, total macrophages, Depleted T cells, NK cells, immune checkpoints BB04 PD-1、PD-L1、CD68、 CD8、CD163、pan-CK、DAPI M1 macrophages, M2 macrophages, killer T cells, immune checkpoints BB05 CD3、CD4、CD20、 CD56、FOXP3、pan-CK、DAPI depleted T cells, immune checkpoints total T cells, helper T cells. B cells, NK cells, Treg BB06 CD4、CD8、GZMB、 Ki67、FOXP3、pan-CK、DAPI depleted T cells, immune checkpoints total T cells, helper T cells. B cells, NK cells, Treg BB07 CD11c、CD20、CD68、 CD163、CD3、pan-CK、DAPI tertiary lymphoid structures, B cells, T cells, and Germinal center activated B cells, follicular dendritic cells Tertiary lymphoid structures, B cells, killer T cells BB08 CD20、CD21、CD23、 CD3、Ki67、pan-CK、DAPI tertiary lymphoid structures, B cells, T cells, and Germinal center activated B cells, follicular dendritic cells BB09 CD20、CD23、CD8、 PD-1、PD-L1、pan-CK、DAPI Tertiary lymphoid structures, B cells, killer T cells follicular dendritic cells, depleted T cells, immune checkpoints BB10 PD-1、CD8、CD68、 CD56、FOXP3、pan-CK、DAPI killer T cells, total macrophages, NK cells, Treg, immune checkpoints  

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Multi-color pathology data analysis
Product description Using spectral signal recognition technology, the overlapping multi-color signals could be accurately split, and with inForm intelligent quantitative pathology analysis software and professional bioinformatics analysis team, qualitative, conformal and quantitative analysis of tissues, cells and proteins in the images could be performed, and finally the data results are visually displayed in the form of visual figures, providing customers with standardized and customized data analysis to meet different types of needs.

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Proteomics testing service

 Wuhan National Human Genetic Resources Library has two proteomics technology platforms, one is the ultra-micro precise proteomics technology platform based on Olink Neighborhood Extension Technology (PEA), which focuses on targeted protein detection, can take into account high throughput, high specificity, high sensitivity and wide dynamic range, and is mainly applicable to humans and mice; The other is an unbiased proteomics technology platform based on mass spectrometry (LC-MS), which has no species limitation. In addition to conventional identification and quantification, it can also assist in protein interaction and post-translational modification research.

Olink Proteomics Services
Proteomics services by LC/MS
Olink Precision Proteomics Analysis Service
Product Description With the accomplishment of human whole genome sequencing, medical research has moved from genomics to the post-genomic era. The research value of proteins as functional executors of life activities is gaining more and more attention. Protein biomarkers can serve as important features in pathophysiology, bridge the gap between genome and phenotype, and have a profound impact on improving future healthcare, especially in precision medicine. However, the progress has been stunted by the lack of technologies that can provide high specificity, sensitivity, and throughput. The emergence of Olink technology, however, can break through these obstacles and accelerate the development of proteomics in the field of precision medicine. Technology principle. Olink protein detection platform is based on the neighbor extension technology Proximity Extension Assay (PEA) for protein detection. For each protein to be detected, a pair of antibodies is designed at two adjacent antigenic epitopes, which are coupled to a specific DNA single strand with paired bases at the end of the single strand. When the pair of antibodies binds to the target protein, the two adjacent DNA single strands are hybridized and complemented and extended in an enzymatic reaction, which then forms a double-stranded template. The detection of the double-stranded template using microfluidic qPCR or NGS enables the quantitative analysis of the protein. This technique cleverly converts protein quantification to DNA quantification, solving the problem that proteins cannot be amplified, thus enabling the detection of trace amounts of proteins. Technical advantages. 1. High specificity: High specificity by using two antibodies to recognize one protein, labeling and full validation by Barcode. 2. Wide dynamic range: The entire panel spans 10 log values, allowing simultaneous consideration of proteins of different abundance. 3. High sensitivity: The detection sensitivity is as low as fg/mL level, which can detect thousands of disease-related low abundance proteins (especially low abundance biomarkers in plasma and serum samples) at the histological level. 4. High detection throughput: 96/384/3072panel, each panel can detect 88 samples at a time. By combining qPCR and NGS high throughput detection platform, it can produce millions of protein data per week. 5. Micro sample: 48-384 protein markers can be detected simultaneously in 1μL plasma, and more than 3000 protein markers can be detected in 8μL plasma. 6. High reproducibility: strict quality control, 99.8% of proteins without cross-reactivity, mature detection platform, high data quality, good reproducibility, suitable for large data analysis requirements, and the reproducibility requirements of clinical translational applications. Application areas. Biomarker research: disease risk assessment, disease typing, disease prediction, disease diagnosis and prognosis. Drug discovery: drug target discovery, clinical treatment effect evaluation, basic research.

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Protein qualitative analysis services
a. Protein full-spectrum analysis Protein full-spectrum analysis can also be called mass spectrometry shotgun analysis. which refers to the component analysis.The Objects of study are complete tissues, body fluids or their extracts, with the aim of identifying as many peptides and protein molecules as possible. The basic principle of protein mass spectrometry identification is to digest proteins into peptide mixtures by proteases, and then ionize them by electron bombardment or other means to form charged ions with different mass-to-charge ratios, and then separate the peptide ions with specific mass-to-charge ratios by a mass analyzer. The protein was identified by comparing the actual spectrogram with the primary and secondary mass spectra of protein digested by protease in theory. Schematic diagram of protein full spectrum analysis   Instruments used: Orbitrap Eclipse, Orbitrap Exploris 480, Q Exactive plus; AB SCIEX: TripleTOF 5600 plus. Technical advantages: Comprehensive identification of a large number of proteins in whole cells, tissues or complex mixed samples, with 200-5000 proteins can be identified. Application areas: Identification of all expressed proteins in tissues, cells or organelles in a given physiological state. Sample requirements: Sample Sample requirements Protein solution Total protein > 500 μg, concentration > 0.1 μg/μL; Buffer solution without detergent NP40, Triton X-100, etc.   Lead time（Project Cycle） Progress  Time period Sample collection/Quality control 2-3 working days Sample processing  4-5 working days Mass spectrometry detection 2-3 working days Data analysis 2-3 working days Total  No more than 15 working days Note: For more than 5 samples, 3 working days will be added for each additional sample. If there is an urgent need, please contact us for negotiation.   b. Protein Identification LC-MS/MS technology was used to identify proteins for moderately complex samples such as single strips (i.e., SDS-PAGE samples), IP, Co-IP, and Pull-down purification solutions. 凝胶电泳条带鉴定路线图   Pull-down experiment roadmap   Instruments: Orbitrap Eclipse, Orbitrap Exploris 480, Q Exactive Plus . Technical advantages: high sensitivity, high accuracy. Suitable for moderately complex protein samples, such as IP, Co-IP . Application areas: disease marker research, mechanism of action research, plant resistance research, drug target research, specific functional protein identification.   Sample requirements: Sample Sample requirements SDS-PAGE bands   Komas staining, silver staining (mass spectrometry compatible) bands are clearly visible Protein solution Total protein >5μg, concentration >0.1μg/μL; buffer without detergent NP40, Triton X-100, etc.   Lead time（Project Cycle） Progress Time period Sample collection/Quality control 1-2 working days Sample processing  2-3 working days Mass spectrometry detection 2-3 working days Data analysis      2-3 working days Report generation 1-2 working days Total  No more than 10 working days Note: For more than 10 samples, 1 working days will be added for each additional sample. If there is an urgent need, please contact us for negotiation.

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Protein Quantification Service
a.TMT™ Technology TMT™ (Tandem Mass Tag™) technology is an in vitro peptide labeling technology developed by Thermo Scientific, which is designed for the identification and quantification of proteins in different samples by tandem mass spectrometry (MS). This technique uses multiple isotope labels to label the amino groups of peptides, and after LC-MS/MS analysis, the relative amounts of proteins in 2-16 groups of different samples can be compared simultaneously. TMT™技术示意图   Instruments used:Orbitrap Eclipse, Orbitrap Exploris 480,Q Exactive Plus. Technical advantages: Simultaneous analysis of protein differences in 2-16 samples of different conditions. Suitable for a wide range of sample types: cytoplasmic proteins, membrane proteins, nuclear proteins, extracellular proteins, etc. Compared with protein quantification by gel separation, TMT mass spectrometry can detect more types of protein molecules, such as low-abundance proteins, strongly acidic (base) proteins, <10KD or >100KD protein molecules. Application areas: disease marker screening, mechanism of action research, plant resistance research, drug action target research, special functional protein screening.   Sample requirements Sample Type      Sample requirements (per group of samples) Protein extract   Concentration >1μg/μL, total protein >300μg Cell sample   Cell volume >10^7 Tissue samples   Animal, microbial tissue wet weight >10mg; plant fresh tissue >100mg Body fluid sample  Blood volume>500μL   　Lead time： Progress  Time period Sample collection/Quality control 1-2 working days Sample processing  5-6 working days Mass spectrometry test 3-4 working days Data analysis 2-3 working days Total  No more than 20 working days Note: For more than 10 samples, 1 working days will be added for each additional sample. If there is an urgent need, please contact us for negotiation.   b.Label-free Label-free quantification is the mass spectrometric analysis of enzymatic protein peptides by liquid chromatography-mass spectrometry (LC-MS). Label-free定量蛋白组学实验流程   Instruments used: THERMO: Orbitrap Eclipse, Orbitrap Exploris 480,Q Exactive Plus.   Technical advantages: no labeling of proteins, low cost and low volume of samples required. Application areas: disease marker screening, mechanism of action research, plant resistance research, drug target research, special function protein screening.   c.SWATH/DIA SWATH/DIA is a new panoramic scanning mode for mass spectrometry. Compared with the traditional shot-gun technique, SWATH/DIA acquisition mode is able to scan all precusor peptide in the scan interval at ultra-high speed and perform secondary fragmentation, thus obtaining complete peptide information. With SWATH /DIA acquisition mode, complete quantitative and qualitative results can be obtained in a single experiment, without method optimization. SWATH/DIA mode overcomes the problem of shot-gun scan randomness, resulting in better data reproducibility. SWATH/DIA is particularly suitable for enrichment of purified samples, e.g. SWATH/DIA quantification after IP to detect proteins that interact with the target protein. SWATH技术示意图   Instruments used: THERMO: Orbitrap Eclipse, Orbitrap Exploris 480, Q Exactive plus; AB SCIEX: TripleTOF 5600 plus Technical advantages High sensitivity, which has the data acquisition mode of MRM, combined with a high-resolution mass spectrometry system with comparable sensitivity to MRM. Quantitative correlation between duplicate samples can reach above 0.99. Quantitative accuracy almost comparable to MRM technique. the quantification range can span 4 orders of magnitude The quantitative results are very good. Application areas: mechanism and regulatory mechanism research, screening of disease markers, screening of drug use and prognostic markers, molecular typing of diseases.     Sample requirements： Sample Type      Sample requirements (per group of samples) Protein extract   Concentration >1μg/μL, total protein >300μg Cell sample   Cell volume >107 Tissue samples   Animal, microbial tissue wet weight >10mg; plant fresh tissue >100mg Body fluid sample Blood volume>500μL   Lead time: Progress Time period Sample collection/Quality control    1-2 working day Sample processing  5-6 working days Mass spectrometry test 3-4 working days Data analysis 2-3 working days Total  No more than 20 working days Note: For  more than 4 samples, 5 working day will be added fo

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Protein modification analysis service
The research work of proteomics not only focuses on the changes of protein expression levels in different cell growth periods or under disease conditions, but also many vital life processes are not only controlled by the relative abundance of proteins, but also regulated by spatiotemporal and spatiotemporal distribution of reversible post-translational modifications. Therefore, revealing the rules of post-translational modifications is an important prerequisite for analyzing the complex and diverse biological functions of proteins. The molecular weight of protein will change when it is modified after translation. The molecular weight of protein or polypeptide can be accurately determined by mass spectrometry. At the same time, the content of post-translational modified protein in the sample is low and the dynamic range is wide, so it is necessary to enrich the modified protein or peptide segment before mass spectrometry detection.

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Small molecule metabolite detection services

Wuhan National Human Genetic Resources Bank is committed to the targeted and non-targeted mass spectrometry technology detection of small molecular metabolites and nucleic acid mass spectrometry technology detection services. The main technical services include: structural determination of small molecular compounds, determination of biomacromolecule mass, nucleic acid mass spectrometry, discovery of biomarkers, including metabolomics and lipomics analysis.

Metabolomics service
Nucleic acid mass spectrometry detection service
Targeted metabolomics
Targeted metabolomics analysis is the targeted and specific detection and analysis of the specified metabolites in the list by using standard samples as controls. The key points of targeted metabolomics analysis are high accuracy, high throughput and high reliability. Although this type of analysis is more routine than metabolic profiling, it can be used for very large numbers of samples. Targeted metabolomics can be applied to. (1) Validating new findings and hypotheses proposed by metabolomics experiments; (2) Conducting hypothesis-based exploratory experiments and studying metabolic models for specific metabolites. Basic Research Process The process of targeted metabolomics research can be divided into: sample collection, standard curve creation, metabolite extraction and isolation, on-line assay, and data analysis.   Sample types Serum - Plasma - Urine - Cerebrospinal fluid - Saliva - Feces - Microorganisms - Cells - Plants - Tissues - Others   　 Targeted metabonomics services and cycles are shown in the following table: 靶向代谢组学服务和周期 平台 仪器型号 代谢物 化合物数目 检测周期 检测价格 GC-MS/MS Aglient 7890A-5975C或8890-7010B 短链脂肪酸 11种短链脂肪酸 约15个工作日 请询价 游离脂肪酸 37种游离脂肪酸 UHPLC-QqQ MS SCIEX QTRAP 4500或Triple Quad 4500 维生素 12种水溶性 约20个工作日 请询价 氨基酸 32种氨基酸 神经递质 27种神经递质 植物激素 10种植物激素 胆汁酸 48种胆汁酸 辅酶 18种辅酶 抗生素 105种抗生素   11种短链脂肪酸绝对定性定量分析 序号 中文名称 英文名称 分子式 CAS号 相对分子质量 1 乙酸 acetic acid C2H4O2 64-19-7 60.05 2 丙酸 propionic acid C3H6O2 79-09-4 74.08 3 异丁酸 isobutyric acid C4H8O2 79-31-2 88.11 4 丁酸 n-butyric acid C4H8O2 107-92-6 88.11 5 异戊酸 isovaleric acid C5H10O2 503-74-2 102.13 6 戊酸 n-valeric acid C5H10O2 109-52-4 102.13 7 4-甲基戊酸 4-methylvaleric acid C6H12O2 646-07-1 116.16 8 正己酸 hexanoic acid C6H12O2 142-62-1 116.15 9 2-甲基己酸 2-methylhexanoic acid C7H14O2 4536-23-6 130.18 10 正庚酸 n-heptylic acid C7H14O2 111-14-8 130.18 11 正辛酸 n-octanoic acid C8H16O2 124-07-2 144.21   37种游离脂肪酸绝对定性定量分析 编号 中文名称 英文名称 简写 1 丁酸 butyric acid C4:0 2 己酸 caproic acid C6:0 3 辛酸 caprylic acid C8:0 4 癸酸 capric acid C10:0 5 十一烷酸 undecanoic acid C11:0 6 月桂酸 lauric acid C12:0 7 十三烷酸 tridecanoic acid C13:0 8 十四酸 myristic acid C14:0 9 十四碳烯酸 myristoleic acid C14:1n5 10 十五烷酸 pentadecanoic acid C15:0 11 顺-10-十五碳烯酸 cis-10-pentadecenoic acid C15:1n5 12 棕榈酸 palmitic acid C16:0 13 棕榈油酸 palmitoleic acid C16:1n7 14 十七烷酸 heptadecanoic acid C17:0 15 顺-10-十七烯酸 cis-10-heptadecenoic acid C17:1n7 16 硬脂酸 stearic acid C18:0 17 反油酸 elaidic acid C18:1n9t 18 油酸 oleic acid C18:1n9c 19 反亚油酸 linolelaidic acid C18:2n6t 20 亚油酸 linoleic acid C18:2n6c 21 γ-亚麻酸 r-linolenic acid C18:3n6 22 α-亚麻酸 α-linolenic acid C1

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Non-targeted metabolomics
Non-targeted metabolomics (i.e. discovery metabolomics) is a non-hypothetical approach that aims to obtain as many metabolites as possible from a single analysis. The main purpose is to compare the metabolomes (all metabolites of a given organism) of the control and experimental groups to find out the differences in their metabolites and to explore the metabolic pathways between the differential metabolites. The analysis generally includes: a. Metabolic profiling (also called differential expression analysis): looking for metabolites of interest with statistically significant abundance changes in a set of experimental and control samples. b. Identification: After performing metabolic profiling, the chemical structure of these metabolites is determined. c. Interpretation: The final step in the research process, explaining the association between the metabolites found and the biological process or biological state. Research Process The non-targeted metabolomics research process can be mainly divided into experimental design, sample collection and processing, metabolite extraction and concentration, sample detection, data analysis, metabolite identification, and finally biological interpretation. Because the metabolome changes extremely fast, the metabolite species are various, the concentration varies greatly, the chemical properties are different, and the data information is huge, each step may have a large impact on the final results, so it is crucial to realize the standardized operation of metabolomics   Technology platform The main platforms for non-targeted metabolomics are nuclear magnetic resonance (NMR), gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS), each of which has a certain bias, so researchers need to choose the appropriate platform according to the purpose of the study and the type of samples. The purpose is to achieve a more complete and comprehensive study. Table Comparison of major assay platforms for non-targeted Major Assay Platforms Advantages Disadvantages NMR 1. High throughput 2. Universality 3. Good objectivity and reproducibility 4. Fast (2~3 min/sample) 5. Simple pretreatment, no derivatization and separation required 6. Non-invasive 7. Can detect most organic compounds in the sample 8. Long instrument life 1. Detection dynamic range is narrow 2. Lower sensitivity and resolution than MS 3. The sample amount is relatively large (0.1 ~ 0.5 mL) 4. Expensive instrumentation and maintenance costs GC-MS 1. High throughput 2. High precision, sensitivity and reproducibility 3. Has a reference standard spectral database, easy to characterize 4. The sample amount is moderate (0.1~0.2 mL) 5. Can detect most organic and some inorganic molecules in the sample   1. Requires derivatization 2. Requires separation 3. Slow analysis speed (20~40 min/sample) 4. It is difficult to identify new compounds 5. Not suitable for the analysis of difficult volatile, thermally unstable substances LC-MS 1. High throughput 2. High separation rate and sensitivity 3. Wide dynamic range of detection 4. Simple sample processing, no need for derivatization 5. Small sample size (10~100 μL) 6. Applicable to thermally unstable, non-volatile, not easily derivatized and large molecular weight substances  1. Slow analysis speed (15~40 min/sample) 2. Lack of standard spectral database for reference 3. Difficult to identify new compounds 4. Higher cost 5. Short instrument life   Data analysis methods The data obtained from non-targeted metabolomics are complex and multidimensional, and the chemometric methods need to be fully applied to deeply explore the information in them. The data are generally pre-processed first, and the specific process includes normalization, data transformation, centralization, and normalization steps. Only after pre-processing, simplification and dimensionality reduction of the data are realized, and reliable data models are established. The pattern recognition methods applied to metabolomics data analysis mainly include unsupervised learning methods and supervised learning methods. The unsupervised learning methods mainly include: principal component analysis (PCA), nonlinear mapping, cluster analysis, etc. Supervised learning methods are mainly based on partial least squares (PLS), neural network improvement methods, least squares-discriminant analysis (PLS-DA), orthogonal least squares (OPLS), etc., among which PCA and PLS-DA are commonly used pattern recognition methods in metabolomics.   Non-targeted metabolomics data processing process   Sample types Serum - Plasma - Urine - Cerebrospinal fluid - Saliva - Feces - Microorganisms - Cells - Plants - Tissues – Other   Lead time

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Nucleic acid mass spectrometry detection service
MassARRAY is based on Matrix Assisted Laser Desorption /Ionization-Time of Flight (MALDI-TOF) technology, and is the global original nucleic acid mass spectrometry platform for high precision DNA qualitative analysis. This technology platform Perfectly integrates the high sensitivity of PCR technology, the high throughput of chip technology,the high accuracy of mass Spectrometry technology and the powerful function of computer Intelligent analysis, which is the best tool for large sample size Verification of a large number of candidate SNP and DNA methylation markers discovered by sequencing and chip technology.The MassARRAY system provides a unique solution for targeted gene detection using limited samples. It can meet the assay design, validation and performance needs of genomics laboratories.   Technology Principle The MassARRAY time-of-flight mass spectrometry detection system consists of a matrix-assisted laser desorption ionization (MALDI) source, a time-of-flight mass analyzer (TOF), and a template preparation plate.The principle of MALDI is to irradiate a co-crystalline film formed by the sample and matrix with a laser, and the matrix absorbs energy from the laser and transmits it to the biomolecules, causing them to ionize.The principle of TOF is Biomolecules are ionized and accelerated through the flight tube by an electric field, and the mass-to-charge ratio of the ions is proportional to the ion's time of flight. The system is extremely accurate and fast, allowing multiple samples to be tested at one time. Multiplex PCR amplification is first performed, followed by single-base extension with modified ddNTP, enabling the mass spectrometry system to accurately identify differences in individual bases and report the frequency of each base according to peak area, and thus accurately analyze the frequency of allele distribution in the sample. Application scope Detection of variant types :. a.SNPs/SNVs (point mutations) b.Indels (insertion/deletion mutations) c.CNVs (copy number variants) d.Gene Fusions e.Methylations Main application areas a.Suitable for genome-wide SNP association studies and subsequent large sample size validation b. Susceptibility gene analysis and gene localization for complex diseases c.High sensitivity analysis of somatic mutations d. Population genetics studies e. Pharmacogenomics (drug development and individual drug use) f. Other: pathogenic microbial detection, health management, biological sample identification, etc.   Available tests Skin type genetic test Advanced tumor susceptibility gene test for men(six items) Metabolic genetic test Advanced tumor susceptibility gene test for women (seven tests) Weight and health management gene test Deafness gene test Scientific exercise gene test    Folic acid gene test Breast cancer susceptibility gene test Gene test for hypertension Basic five tumor susceptibility gene test Epilepsy gene test Cardiovascular disease gene test              Project advantages a. Multiplex reactions, fast detection cycle Single reactions up to 10 to 60 assays and only simple PCR and extension reagents are required, no fluorescent probes are needed and the project lead time is within 5 working days. b.High accuracy and sensitivity Typing accuracy >99.7%, which is the gold standard for SNP detection; detection sensitivity up to 0.1% for low frequency alleles and rare mutations. c.Flexible throughput The standard 96 chips are available to meet the needs of different detection volumes, and the samples can be tested on the machine without making up samples. d. High sample compatibility Each reaction well requires only 5-10ng of genomic DNA, and supports biological samples from various sources: blood, blood collection cards, oral swabs, saliva, semen, fresh tissue, puncture tissue and FFPE samples, seeds, roots, leaves, etc. Even highly degraded samples can be tested. e. Support different types of marker detection for a wide range of applications It can detect SNP, point mutation, In/Del, CNV, gene fusion and methylation and many other variant types, which can be applied to genetic diseases, solid tumors and liquid biopsies, pharmacogenomics and many other fields.

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