搜索
技术平台

High throughput cell sorting

Product Description
Case Analysis
Result display
Sample delivery
Q&A

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

 

Signed article: RNF186/EPHB2 Axis is Essential in Regulating TNF Signaling for Colorectal Tumorigenesis in Colorectal Epithelial Cells

Sample type: stable transitional cell line

Application techniques: Flow cytometry, etc.

Study Overview: The team found that EPHB2 is required for TNF-induced colorectal epithelial cell signaling activation and pro-inflammatory cytokine production. Upon TNF stimulation, EPHB2 is ubiquitinated by its E3 ligase RNF186. The ubiquitinated EPHB2 then recruits and further phosphorylates the nine tyrosine sites of TAB2, prompting TAB2 to bind TAK1, thereby activating the NF-kB signaling pathway and producing pro-tumor cytokines such as TNF. In RNF186 knockout mice, defects in the TNF signaling pathway in colorectal epithelial cells, compared to wild-type control mice, were colitis-driven colorectal cancer model phenotypes such as tumor number and size were significantly reduced.

Furthermore, and more surprisingly, the EPHB2 gene mutant (D862N) found in a colorectal cancer family was a functionally acquired mutation that promoted activation of the TNF signaling pathway compared to wild-type EPHB2. As RNF186 increases EPHB2-D862N ubiquitination and further enhances affinity for TAB2, leading to more intense activation of NF-kB signaling pathway and TNF production during tumorigenesis, this study also reveals the mechanism by which EPHB2 mutations lead to colon carcinogenesis.

96-well plate sorting of knockdown GFP-positive cell lines for monoclonal screening 

(1) Conventional sorting and back testing  

Conventional GFP sorting

 

(2) Multicolor sorting

(3) Develop and optimize sorting scheme
Q: How to effectively obtain the sorting of tissue-like lymphocytes?
A: It is suggested that Ficoll separation or magnetic bead separation can be used before upflow separation.

 

 

(4) Sorting helps single cell sequencing, SMART-Seq2

 

 

Single cell sequencing one-stop service 

 

 

           

                                                                                                          Living cell sorting                                                         D45 target cell sorting

Sample Type     

Sample Requirements 

Notes

Cells Cells in culture

Fill the culture bottle with cell culture solution, tighten the cap and seal with sealing film   

Please keep 4℃ for in-city delivery (in case of ice bag, physical isolation is required to avoid freezing) and deliver to our laboratory within 2 hours; if mailing samples, only frozen cells can be mailed on dry ice

Cell suspension

After digestion with trypsin into a single cell suspension (or suspension without trypsin) and washing with PBS, count live cells, make sure the viability is over 80%, and adjust the cell concentration to (5-10)×105 cells/mL. 

Cryopreserved cells

Cell concentration of (5-10) × 105 cells/mL, viability >80%     

 

Dry ice delivery

Blood Whole blood

Anticoagulated whole blood (volume needs to be communicated according to the specific type of test) to confirm that the blood is non-coagulated and non-hemolyzed 

1. if intracellular cytokines need to be detected, it is recommended to use heparin anticoagulation tubes to store the anticoagulated blood and avoid using EDTA anticoagulation tubes.

2. avoid violent collisions and rapid temperature change during the transportation of blood samples, keep them at 4℃ (in case of ice bags, physical isolation is required to avoid freezing) and deliver them to our laboratory within 24h.

3. If the sample cannot be tested immediately and needs to be frozen, the whole blood sample cannot be frozen, and PBMC needs to be separated before freezing; the frozen sample needs to be transported on dry ice.

PBMC

Lysate or lymphatic isolate to obtain PBMC, freeze

Tissue samples (spleen, lymph nodes, lung, subcutaneous tumors, etc.)   

Fresh tissue should be no less than 0.5 cm × 0.5 cm × 0.5cm in size, necrotic tissue needs to be removed and blood needs to be cleaned; preserve the sample with tissue preservation solution and keep the sample in a 4°C environment (in case of ice bags, it needs to be physically isolated to avoid tissue freezing) and send  to our laboratory within 24h

1. fresh tissue samples are taken in situ and preserved with preservation solution as soon as possible within 1h.

2. If the sample cannot be tested within 48h, it needs to be frozen and stored, and the tissue should not be frozen directly, it needs to be processed into single cell suspension and then frozen, and transported on dry ice

Follow us

Link: