Derivation Services

To place an order, please use the service request form. The HSCI iPS Core offers the following derivation and characterization services (rates differ according to Harvard affiliation and profit/non-profit status):

Rates for Harvard and HSCI Faculty+ Quantity Discounts
Derivation service modules Base Rate (0-4 samples) Discount (for 5-9 samples  Discount (for 10-49 samples) Discount (for 50+ samples)
Module 1: PBMC isolation and freezing from blood samples $201 $193 $185 $159
Module 2a: Reprogramming fibroblasts using Sendai virus method $5,120 (2-5 colonies) $4,923 (2-5 colonies) $4,727 (2-5 colonies) $4,139 (2-5 colonies)
Module 2b: Reprogramming blood (T cells or erythroblasts) using the Sendai virus method $5,608 (2-5 colonies) $4,875 (2-5 colonies) $4,681 (2-5 colonies) $4,100 (2-5 colonies)
Module 2c: Reprogramming fibroblasts using the modified mRNA/microRNA combination $5,283 (2-5 colonies) $5,079 (2-5 colonies) $4,874 (2-5 colonies) $4,261 (2-5 colonies)
Module 2d: Reprogramming somatic cells using the episomal vector technology $4,939 (2-5 colonies) $4,752 (2-5 colonies) $4,564 (2-5 colonies) $4,003 (2-5 colonies)
Characterization service modules  
Module 3: ICC, EB formation, gene expression $1,441/line $1,379/line $1,317/line $1,131/line
Module 4: Karyotyping $630/line N/A
Module 5: DNA fingerprinting $336/line N/A
Module 6: Teratoma formation $1,696/line N/A
Module 7: ddPCR for BCL2l1 $244/line $232/line $221/line $187/line
Module 8: Sendai virus elimination testing $236/line $225/line $214/line $181/line

+Note: "Harvard and HSCI Faculty" includes two groups of users: (1) All Harvard faculty and researchers using HU 33-digit charging code, and (2) faculty with HSCI appointments.

Rates for non-profit institutions Quantity Discounts
Derivation service modules Base Rate (0-4 samples) Discount (for 5-9 samples) Discount (for 10-49 samples) Discount (for 50+ samples)
Module 1: PBMC isolation and freezing from blood samples $223 $214 $206 $181
Module 2a: Reprogramming fibroblasts using Sendai virus method $5,884 (2-5 colonies) $5,688 (2-5 colonies) $5,491 (2-5 colonies) $4,903 (2-5 colonies)
Module 2b: Reprogramming blood (T cells or erythroblasts) using the Sendai virus method $5,832 (2-5 colonies) $5,639 (2-5 colonies) $5,445 (2-5 colonies) $4,864 (2-5 colonies)
Module 2c: Reprogramming fibroblasts using the modified mRNA/microRNA combination $6,047 (2-5 colonies) $5,843 (2-5 colonies) $5,639 (2-5 colonies) $5,025 (2-5 colonies)
Module 2d: Reprogramming somatic cells using the episomal vector technology $5,703 (2-5 colonies) $5,516 (2-5 colonies) $5,329 (2-5 colonies) $4,767 (2-5 colonies)
Characterization service modules  
Module 3: ICC, EB formation, gene expression $1,569/line $1,507/line $1,445/line $1,258/line
Module 4: Karyotyping $683/line N/A
Module 5: DNA fingerprinting $389/line N/A
Module 6: Teratoma formation $1,707/line N/A
Module 7: ddPCR for BCL2l1 $254/line $243/line $232/line $198/line
Module 8: Sendai virus elimination testing $247/line $236/line $225/line $192/line
Rates for for-profit institutions Quantity Discounts
Derivation service modules Base Rate (0-4 samples) Discount (for 5-9 samples) Discount (for 10-49 samples) Discount (for 50+ samples)
Module 1: PBMC isolation and freezing from blood samples $359 $341 $323 $269
Module 2a: Reprogramming fibroblasts using Sendai virus method $9,936 (2-5 colonies) $9,936 (2-5 colonies) $8,942 (2-5 colonies) $7,452 (2-5 colonies)
Module 2b: Reprogramming blood (T cells or erythroblasts) using the Sendai virus method $9,849 (2-5 colonies) $9,356 (2-5 colonies) $8,864 (2-5 colonies) $7.387 (2-5 colonies)
Module 2c: Reprogramming fibroblasts using the modified mRNA/microRNA combination $10,212 (2-5 colonies) $9,702 (2-5 colonies) $9,191 (2-5 colonies) $7,659 (2-5 colonies)
Module 2d: Reprogramming somatic cells using the episomal vector technology $9,630 (2-5 colonies) $9,148 (2-5 colonies) $8,667 (2-5 colonies) $7,222 (2-5 colonies)
Characterization service modules  
Module 3: ICC, EB formation, gene expression $3,600/line $3,420/line $3,240/line $2,700/line
Module 4: Karyotyping $1,154/line N/A
Module 5: DNA fingerprinting $658/line N/A
Module 6: Teratoma formation $2,885/line N/A
Module 7: ddPCR for BCL2l1 $430/line $408/line $387/line $322/line
Module 8: Sendai virus elimination testing $417/line $396/line $375/line $313/line

 

Process Guideline

Derivation services

Module 1: PBMC isolation and freezing.

The HSCI iPS core provides the isolation and freezing of PBMC from blood sample in preparation for their reprogramming. Four to eight ml of blood in EDTA tube (purple cap) should be delivered to the core within 24 hours of the blood draw. The PBMC are isolated by a density gradient centrifugation method using Ficoll and then frozen down at 1-3 x10^6 cells/cryovial.

In the unlikely event that we fail to isolate PBMC from properly collected blood samples due to errors by the Core, there will be no charge for this attempt. We will ask the investigator to provide a new blood sample to complete the service.

Module 2a*: Reprogramming fibroblasts using the Sendai virus method

This service, which takes 2-3 months to complete, includes:

  • Fibroblast expansion, mycoplasma testing, freezing vials and plating for transduction.
  • Transduction of 1 well (200,000 cells/well) with Sendai virus.
  • Splitting onto two 10cm dishes with irradiated MEFs. Feeding every day (one dish with VPA, one dish without)
  • Picking colonies a month later
  • Expansion of colonies
  • Mycoplasma testing
  • Freezing (2-6 vials per line)

Module 2b: Reprogramming blood (T cells or erythroblasts) using the Sendai virus method

This service, which takes 2-3 months to complete, includes:

  • Peripheral blood mononuclear cells (PBMC) isolation form one tube of blood (4 ml minimum)
  • T cells or erythroblasts expansion for few days
  • Mycoplasma testing for cell culture and plating for transduction.
  • Transduction of 1 well (150,000 cells/well) with Sendai virus.
  • Splitting onto one 10cm dish with irradiated MEFs. Feeding every day.
  • Picking colonies a month later
  • Expansion of colonies
  • Mycoplasma testing
  • Freezing (2-6 vials per line)

Module 2c*: Reprogramming fibroblasts using the modified mRNA/microRNA method

This service, which takes 2-3 months to complete, includes:

  • Fibroblast expansion, mycoplasma testing, freezing 3-5 vials
  • Plating fibroblasts into 1 well (50,000 cells/well) on matrigel
  • Preparation of Nuffs conditioned medium
  • Transfection with microRNA on day 1
  • Transfection with mRNA on day 2-3-4
  • Transfection with microRNA/mRNA on day 5
  • Transfection with mRNA every day until day 12
  • Picking 2-5 colonies around day 15 to 20
  • Expansion of colonies
  • Mycoplasma testing
  • Freezing (2-6 vials per line)

Module 2d*: Reprogramming somatic cells using the episomal vector technology

This service, which takes 2-3 months to complete, includes:

  • Somatic cell expansion, mycoplasma testing, freezing 1-3 vials
  • Nucleofecting 500,000 cells with the Epi5 vectors.
  • Splitting onto one 10 cm dish with irradiated MEFs. Feeding every day.
  • Picking 2-5 colonies a month later
  • Expansion of colonies
  • Mycoplasma testing
  • Freezing (2-6 vials per lines)

*Information regarding modules 2a, 2c, and 2d:

  • The fibroblasts can be sent frozen or brought to the core as live culture. The investigator is required to test the fibroblasts for mycoplasma before submission and only negative samples will be accepted. The fibroblasts should be at a low passage (ideally less than 10) and they should be proliferating. In our experience, senescent cultures are not as amenable to reprogramming. If the lack of proliferation or abnormal morphologies is observed, we will discuss with the investigator and either proceed with obtaining new fibroblast samples or attempt to transduce. Failure in such situations will result in the investigator bearing the cost of the attempt.
  • In some situations, for no apparent reason, we do not obtain any colonies from the first round of reprogramming. We will then try one more round using modified strategies (e.g. p53 knock-down, addition of chemical compounds). If the second round is unsuccessful, we will not repeat the reprogramming. The investigator will be charged for half of the price of module 2a, 2c or 2d for two failed attempts.
  • We target 5 colonies per sample for picking. In our experience, 5 colonies have been sufficient to generate 3 karyotypically normal iPS cell lines. The iPS lines can be delivered to the investigator in frozen vials and/or live culture. If the characterization is done at the core, we will deliver 5 frozen vials for the characterized iPS lines and 2 frozen vials for the extra lines. If only module 2 is done at the core, 2 frozen vials will be delivered for each iPS line. No vials will be kept at the core. Unused frozen fibroblasts/PBMC will be delivered to the investigator at the same time.
  • Normally, the iPS colonies are picked and expanded on MEFs. While the derivation will continue to be done on MEFs, we offer feeder-free colony-picking and expansion at no extra charge.

Characterization services

Our current characterization services are only offered for iPS lines derived at the Core. Once iPS lines are established, we will discuss with you which module(s) of characterization you will choose to have the Core perform. The workflow is optimized for proceeding without having to restart the cultures.

Module 3: ICC, EB formation, and Gene Expression

This service, which takes about 1 month to complete, includes:

  • Alkaline phosphatase assay
  • ICC for Oct4, Nanog, TRA-1-60, SSEA3, SSEA4
  • RNA extraction, cDNA synthesis and qRT-PCR for endogenous Oct4, Nanog, endogenous Sox2, Rex1, hTERT, Dnmt3b
  • EB formation (15 days differentiation), RNA extraction, cDNA synthesis and qRT-PCR for 2 mesoderm markers, 2 ectoderm markers and 2 endoderm markers

The data will be sent to the investigator in PowerPoint by e-mail.

Module 4: Karyotyping

This service, which takes about 1 month to complete, includes:

  • Expanding the iPS line to a T25 flask
  • Mycoplasma testing
  • Shipping the line to Cell Line Genetics for karyotyping

The reports will be sent to the investigator by e-mail two to three weeks later.

Module 5: DNA fingerprinting

This service, which takes about 1 month to complete, includes:

  • DNA extraction from fibroblasts
  • Feeder free culture of iPS line and DNA extraction
  • Shipping the DNA to Cell Line Genetics for DNA fingerprinting
  • Short tandem repeat polymorphisms (STRs) for 15 loci plus Amelogenin

The reports will be sent to the investigator by e-mail two weeks later.

Module 6: Teratoma formation

This service, which takes about 2-3 months to complete, includes:

  • Expansion of iPS line
  • Collection of cells and delivery to the genome modification facility
  • Injection into kidney capsule (3 immuno-suppressed mice)
  • Teratoma removal after 8-11 weeks
  • Histology work
  • Analysis