Specific cell types can be purified from peripheral blood or bone marrow using a negative or positive selection strategy. Examples of cell types that CTCL has regularly purified for studies are CLL B cells, CD34+ cells, CD66+ cells and CD138+ cells. Isolated cell purity is assessed by flow cytometry.
Flow Cytometry & Data Acquisition
PBMC or total nucleated cells are isolated from blood and stained with antibodies to identify and quantitate specific cell types. CTCL will help develop and validate specific antibody panels for investigator's studies. CTCL uses the instrumentation provided by the Flow Cytometry Core Lab to acquire data and can help with or perform data analysis with FlowJo software. Cost of flow cytometry assays increases with the number of antibodies required in the panel.
Plasma, Serum & PBMC Isolation
Plasma is isolated from peripheral blood collected in an anti-coagulant-treated vacutainer tube such as an EDTA or Na Citrate-treated tube. Serum is isolated from peripheral blood collected in a vacutainer tube containing no anti-coagulant (red cap). Blood tubes are centrifuged at 1500 RCF for 10 min at 4°C. Investigators can specify custom processing requirements (ie. speed, temperature, time, and aliquot volumes) for downstream applications. Samples can be stored at the CTCL for future studies or shipped to other institutions.
Peripheral Blood Mononuclear Cells (PBMC) or Bone Marrow nucleated cells are isolated from blood or bone marrow collected in an anti-coagulant treated vacutainer tube such as an EDTA, Na Citrate, heparin-treated tube. Mononuclear cells are collected by ficoll gradient or total nucleated cells can be collected using a red cell lysis treatment. Samples can be snap frozen as a cell pellet or cryopreserved in cell freeze media.
Sample Storage & Shipping
The CTCL has both -80°C freezer and liquid nitrogen storage space for long-term sample biobanking. Freezers are regularly monitored and maintained following a CTCL Standard Operating Procedure. NIST temperature validation is performed biannually. Investigators will be charged quarterly fee on the inventoried number of boxes being used.
CTCL can remove, aliquot, and ship or distribute samples to collaborators upon the request of the investigator. There is a charge for this service dependent on the number of samples.
Protein Biomarker Quantification
The quantity or concentration of proteins such as growth factors and cytokines expressed in tissues like human plasma and serum are quantified using commercially available ELISA assay kits or multiplex mesoscale kits. CTCL customers must provide at least 0.5 ml of sample.
- ELISA assays are available from may commercial sources. Samples can be run in batches of 40. Customers will be charged per plate.
- Mesoscale multiplex assays are available from Meso Scale Diagnostics, LLC. 40 samples can be analyzed on one 96-well mesoscale plate which will quantitate two to ten analytes per plate, depending on the Mesoscale assay kit used. Customers will be charged per plate.
Nucleic Acid Purification
DNA can be isolated from snap frozen cell pellets, tissues, saliva, and buccal mouthwash using Qiagen Puregene kit. DNA is quantified using Quibit Fluorimeter. Other quantification and qualification assays can be performed upon request.
RNA is isolated from a cell lysate or tissue using Qiagen RNAeasy kit. RNA is quantified using Qubit Fluorimeter and quality assessed using Agilent Bioanalyzer. Other quantification and qualification assays can be performed upon request.
Using commercial available probe/primer sets or designing custom-made primers, CTCL can analyze RNA and DNA samples by PCR, qPCR, or SNP assays to quantitate gene expression or determine sample genotype.
If you have a project idea but don’t know where to start, or if you have questions about what kind(s) of samples to collect (Plasma, Serum, Peripheral Blood Mononuclear Cells), how to collect them or how much to collect, the CTCL is available to help. Please contact us with your inquiries.
We can help investigators design correlative studies for clinical trials, as well as help with budgeting. Visit the Getting Started section for more information. Please refer to our online pricelist for estimates. Customized pricing is available upon request.