Quantitative gDNA TCR/BCR Sequencing with RepSeq IQ™
Quantitative gDNA TCR/BCR Sequencing with RepSeq IQ™
Services
Allow us to prepare your samples for sequencing with our cell thawing, staining, sorting, plating and nucleic acid extraction options for bulk repertoire and single cell sequencing projects. We ensure efficient and cost-effective services from sample processing to sequencing for projects of all scales.
Ensure viable DNA or RNA from cells or tissue. For projects that simply require RNA from whole blood, we recommend blood collection into PaxGene tubes (or similar).
Send us cells that are fully prepared to be plated for iPair services by iRepertoire’s experienced FACS team.
Cryopreserved cells require thawing and processing prior to sorting and downstream extraction. Our sample processing service guarantees workable samples for downstream analysis.
Our cell staining team is highly proficient at isolating rare cell populations via tetramer and antibody staining and sorting via FACs. We’ve successfully stained and sorted cell subsets at as low as 0.001% availability. We can provide flow cell sorting for bulk repertoire services, as well as single cell isolation and plating in conjunction with our iPair service.
If you plan to submit FFPE samples to iRepertoire for extraction, the preferred sample submission type would be a minimum of 30 µm curls containing at least 30% tumor tissue.
This can be submitted either as 25 µm curls (two minimum) or 10 µm curls (3 minimum) stored in properly labeled 1.5 mL Eppendorf tubes. FFPE slides are also acceptable and should be at least 10 µm in thickness (3 slides minimum). FFPE extraction is performed on a Covaris ME220 with acoustic sonication, which helps to increase the yield of intact RNA.
We can also accept solid tissue, either snap frozen tissue or stored on RNAlater (QIAgen). For solid tissue submissions, we can accept anywhere from 5 mg to 30 mg of whole tissue. Note: The maximum capacity of most commercial extraction kits varies by tissue type, but generally 30 mg is the maximum.
If cells need to be sorted downstream (e.g., isolated T-cell or B-cell populations), we request cryopreserved cells at a concentration of 5-10*106 cells/mL of cryopreservation media with a >75% viability when thawed. The recommended cryopreservation media is CryoStor CS5 or CS10 (BioLife Solutions).
If cells only require nucleic acid extraction, we recommend cells be stored either on Qiagen’s RLT lysis buffer or Qiagen RNAprotect Cell Reagent (please follow this link to a learning center article with more detailed information). For isolated cells, we prefer a submission of 10,000+ cells, with a minimum submission of 250 cells. As a note, the maximum cell input for a single RNA extraction is 1*107 cells.
RNA samples should be stored in nuclease-free water and shipped in 1.5 mL micro-centrifuge tubes.
For RNA submission concentrations and quantities, please see either our RepSeq or RepSeq+ page (depending on your service needs). Please note that the minimum recommended volumes are enough for one repeat. Technical replicates are standard for RepSeq+ barring requests to retain precious samples.
If the listed quantities of RNA are difficult to obtain, please know that we are experienced in achieving amplification from RNA input amounts as low as 200 pg. Quality and RNA intactness are much more important parameters than the RNA input amount.