Services

The iPSC Core Facility provides three service packages designed to cater to all steps of human iPS cell line derivation, including, establishment of primary cell cultures from biopsies, reprogramming of primary somatic cells, and analyses of the resulting cells lines for ensuring high quality. The set of derivation services are designed to provide optimal solutions for specific types of cells and at a range of costs. Notably, we provide a feeder-free human iPS cell derivation service for fibroblasts, PBMCs and lymphoblastoid cells. Please for reprogramming of other cell types and pricing for different methods.

Packages for production of human iPS cell lines as a service

Primary line derivationiPSC derivationCharacterization
Service Packages 1 (a-b)Service Package 2 (a-b)Service Package 3 (a-b)

 

 

Service Package 1a: Derivation of primary fibroblast line from skin biopsy

  • Sample submission: Skin biopsy
  • Duration: The derivation service for primary fibroblasts spans 30 days.
  • Health certificate outlining that the donor is HIV-, HBV-, HCV- has to be provided.
  • 2-4mm diameter fresh biopsy is provided by the client.
  • Skin fibroblasts are expanded over 30 days to approximately 25x10E6 cells total, and 3 frozen vials of cells are stored for reprogramming, and/or sent to clients.

Service Package 1b: Isolation of peripheral blood mononuclear cells (PBMCs)

  • Sample submission: Fresh whole blood
  • Duration: The isolation service for PBMCs spans 1 days.
  • Health certificate outlining that the donor is HIV-, HBV-, HCV- has to be provided.
  • Minimum 10 ml of the whole blood in Heparin or EDTA collection tube is provided by the client.
  • PBMCs are isolated using Ficoll-Paque density gradient centrifugation.
  • From 10 ml whole blood 22-26 million cells can be isolated.

Service Package 2a: Reprogramming of fibroblasts using mRNA

  • Sample submission: Fresh skin biopsy or cryopreserved fibroblast cell lines
  • Target cell type: Adult and neonatal dermal fibroblasts
  • Duration: The service requires 2-3 months for completion.
  • Health certificate outlining that the donor or cryopreserved fibroblasts are HIV-, HBV-, HCV- has to be provided.
  • The primary fibroblasts must be mycoplasma negative.
  • Cultures of primary cells are expanded to 5x10E6 cells.
  • Cells are plated on Matrigel matrix and transfected daily with mRNA cocktail.
  • 10 or more colonies are picked between day 10 and 15.
  • 3 clones exhibiting best morphologies and growth rates are further expanded for 5-6 passages and cryopreserved (2-5 vials per line over 3 passages).
  • Clones are shipped to clients as frozen vials.

The reprogramming method is also suitable for immortalized fibroblasts.

Service Package 2b: Reprogramming of whole blood using mRNA

  • Sample submission: Fresh whole blood or cryopreserved EPCs
  • Target cell type: Endothelial Progenitor Cells (EPCs)
  • Duration: The service required 3-4 months for completion.
  • Health certificate outlining that the donor or cryopreserved EPCs are HIV-, HBV-, HCV- has to be provided.
  • In the case of whole blood submission, the isolation of PBMCs and derivation of EPCs precedes the reprogramming proces. The EPCs derivation process requires 18-24 days.
  • Cells are plated on iMatrix matrix and transfected daily with mRNA cocktail.
  • 10 or more colonies are picked between day 15 and 20.
  • 3 clones exhibiting best morphologies and growth rates are further expanded for 10 passages and cryopreserved (2-5 vials per line over 3 passages).
  • Clones are shipped to clients as frozen vials.

The reprogramming method is also suitable for lymphoblastoid cell lines (LCLs).

Service Package 3a: Basic characterisation of hiPSC

  • Target: Characterisation of newly established or cryopreserved pluripotent cell lines.
  • Duration: The service requires 2 months for completion.
  • Pluripotency testing: Immunocytochemistry  of Oct4, Sox2, Lin28 and Nanog.
  • RNA extraction and qRT-PCR analysis of pluripotency factors.
  • Potency testing: Direct differentiation into three germ layers derivate, RNA extraction, and qRT-PCR analysis of differentiation markers.

Service Package 3b: human iPS cell G-banding karyotyping | pilot phase

  • Duration: The service requires 2 months for completion.
  • iPSC are expanded into 10 cm dishes.
  • Fixation of cells.
  • Metaphase preparation.
  • G-banding stain done in house.
  • 20 metaphases are included in the report provided to customer.

Service Package 4a: Training

Basic training (adjusted individually to the level of the student) focuses on practical experience including methodology and major techniques used in working with hPSCs such as maintenance, preservation and characterization.


Additional advanced individual training possible upon request. Topics might include differentiation, derivation and genetic manipulation.

Additional Information

Information regarding service package 2 (a-c)

It is possible to provide the iPSC Core Facility with established fibroblast lines for reprogramming:

1. The fibroblast should be at low passage (passage 8 or lower).

2. The culture should exhibit rapid proliferation rate and not show signs of senescence. 

3. Primary cells will be processed for reprogramming unless the cultures exhibit signs of senescence or abnormal morphology. We will consult clients if the fibroblasts exhibit poor morphology, and it will be the responsibility of the client to pay half the reprogramming fees if reprogramming fails after consultation with the clients.

 Information regarding service package 2 (a-f)

1. Reprogramming may (infrequently) fail and the underlying reasons may be difficult to pin down. In this case the Core Facility will attempt different methods and modifications of the protocols to improve reprogramming (e.g. by knocking down p53, enhancement by chemicals, etc.). If the cultures fail to reprogram after two rounds of attempts using different methods of reprogramming the client will be charged only half of the cost.

2. For most protocols establishment of iPS cell lines can be done in the presence of MEFs or on feeder-free matrix as requested by the client.

3. Some of the reprogramming experiments performed with HMGU collaborators will be performed free of charge. Please inquire. 

4. The typical maximum number of clones that we offer to characterize is five. After characterization we shall deliver 3 vials of 3 best clones and the remaining of the uncharacterized iPS cell bulk line.

5. After reprogramming we shall deliver back to our collaborators all frozen vials of parental fibroblasts PBMCs cultures used for the reprogramming. The Core Facility will store backup frozen vials of derived iPS cells till the lines are properly established by the clients’ labs, typically within a year from delivery time.

6. HMGU collaborators that request re-derivation of iPS cell lines that were lost due to improper culturing will be subjected to the reprogramming fees.