University of California, Riverside

Stem Cell Core Facility



Cell Culture Facility


1. Planning Stage

  • Principle investigators and researchers who are planning to conduct work on human embryonic and/or induced pluripotent stem cells in the Core should contact the Academic Coordinator,Dr. Iva Afrikanova, at ivka.afrikanova@ucr.edu, or (951)827-4954 for an appointment.

  • The Core currently allows the propagation of stem cells of human origin and mouse embryonic stem cells in culture suites.

2. Regulatory Review Requirements for Human Stem Cells

  • The maintenance and use of HUMAN PLURIPOTENT CELLS (embryonic, induced or lines intended to be induced) requires review by the following regulatory committees (as mandated by federal and state agencies):

Committee

Covered activities

Why?

Protocol

Application

Stem Cell Research Oversight Committee

SCRO

All handling/ use of human pluripotent cells

Ensures state mandated review of ethical concerns related to human stem cells

Stem Cell Use Authorization (SCUA)

Contact IBC@ucr.edu

Institutional Biosafety Committee

IBC

All handling/ use of human derived materials

Ensures appropriate safety procedures for handling human cells

Biological Use Authorization (BUA)

http://or.ucr.edu/OrPortal/index.aspx (UCR faculty only) or contact IBC@ucr.edu

Human Research Review Board

HRRB

All research involving humans or materials derived from humans

Ensures that the rights of the original donors of human materials are maintained

UCR HRRB protocol

HRRB review of human pluripotent cell line use is accomplished through the SCUA (the HRRB reviews relevant SCUA sections)

Institutional Animal Care and Use Committee

IACUC

Only research involving the use of live non-human vertebrate animals

Ensures appropriate animal care. Needed only if mouse embryonic fibroblasts (MEF) are to be harvested or if experiments will involve vertebrate animals

Animal Use Protocol (AUP)

http://or.ucr.edu/Home/Forms.aspx?T=2 or contact IACUC@ucr.edu

  • Because it may take several months to complete the above reviews, it is recommended that researchers who have not obtained the above approvals complete the necessary paperwork and begin the approval process prior to the time they anticipate beginning a stem cell project.

  • PIs with existing BUAs must amend their BUA to include the Core Facility as a location.

  • To help investigators complete the above requirements and obtain MTAs (see next section), a checklist for completing paperwork and obtaining proper clearances is provided in the appendix of this document.

3. Material Transfer Agreement (MTA)

  • The Office of Technology and Commercialization reviews and approves Material Transfer Agreements (MTA). PIs are required to obtain their own MTAs, signed by authorized representatives, prior to using materials in the Core. Please contact UCR's Material Transfer Officer at MTA@ucr.edu to facilitate this process. (See http://www.ora.ucr.edu/IP/MTA.aspx for further information and guidance.)

  • Receipt, use, and storage of human pluripotent stem cells must comply with the MTA of the provider. Please be aware that different providers may have different MTA requirements. All PIs are able to maintain HESCs and HIPSCs at the Stem Cell Core, subject to the following MTA requirements:

  1. PIs are required to obtain their own MTA to grow HESCs and HIPSCs.

  2. The MTA should indicate that these cells will be used in their lab and in the Core. PIs will then be able to bring cells into the Core, and in some instances, may be able to buy them from the Core.

4. Cell screening

  • Any live cell cultures that are brought into the Core will require screening for Mycoplasma and must be verified to be clear of Mycoplasma, Acholeplasma, Ureaplasma and other cell culture contamination (bacteria, fungi).

  • The Core will provide screening for Mycoplasma. While eventually there will be an established recharge rate for this service, initially the investigator will need to provide the reagents needed for Mycoplasma screening and the Core personnel will perform the testing.

  • Once a line is established to be free from contamination, it can be expanded, frozen down, and used in the Core. Should samples of the line be returned to the PI's laboratory, they would need to be rescreened before being brought back into the Core.

  • PIs may choose to use cells provided by the Core, which have already gone through this screening process.  

5. Stem Cell Core Cell Culture Requirements

  • Sterile and aseptic cell culturing techniques will be practiced at all times.

  • Only individuals who have been trained for cell culturing will be approved to use the sterile culture suites.

  • Users of the culture facilities should not consume yeast based beverages 24 hours prior to working in the culture labs. Individuals who bake at home should not do so for 3 days prior to using the culture facilities.

  • Lab coats, face masks, and booties must be worn at all times in the tissue culture rooms.

  • All culture work in the Core must be done with reagents (media, growth factors, plasticware, etc) supplied by the Core. This is necessary to minimize contamination in the sterile areas.

  • Any cells that show signs of contamination will be discarded.

  • Individuals who do not comply with the sterile operating procedures of the Core will not be allowed to perform experiments in the Core.

6. Tissue Culture Suites

There are three tissue culture rooms in the culture suite at the Stem Cell Core. Tissue Culture Room A houses one Class II biosafety tissue culture cabinet and one laboratory fume hood. Tissue Culture Room B and C each house two Class II biosafety tissue culture cabinets.

  • The culture suites foyer is a semi-sterile area.
  • Only individuals with approved projects in the culture area are admitted to this section of the Core.
  • Upon entering the culture suite, it is necessary to step on a blue sticky pad and put on a lab coat, which will be provided by the Core.
  • The tissue culture rooms are sterile areas.
  • When entering a culture room, it is necessary to step on a blue sticky mat and put on booties. The lab coat and booties must be worn at all times in the tissue culture rooms.
Tissue Culture Room A

Tissue Culture Room A (TC Room A, Prep Room) is assigned for maintenance of the Stem Cell Core's own human embryonic stem cell (HESC) and human induced pluripotent stem cell (HIPSC) lines. This room is also used for culturing mouse embryonic stem cells lines. It is also assigned for the extraction and preparation of mouse embryonic tissues such as fibroblasts and for the production of transgenic cell types.

  • This room houses two incubators. One is for use with human cells and the other for use with mouse embryonic stem cells and mouse embryonic fibroblasts.
  • A fluorescence stereoscopic (dissecting) microscope is available in TC Room A for use in dissecting mice and for in vivo work.
  • Experiments involving the use of retroviruses, lentiviruses, and/or adenoviruses can be conducted in the class II biosafety tissue culture cabinet in TC Room A only.
  • Magnetic activated cell sorter (MACS, Miltenyi Biotech) for live cell sorting with direct or indirect fluorescence using antibody markers is available for use with human and mouse stem cells in this room on a fee for usage basis.
Tissue Culture Room B

Tissue Culture Room B (TC Room B) is used to maintain other human stem cell types (e.g., non-pluripotent) as well as to culture and prepare cells for incubation in the BioStation CT. Both normal and low oxygen incubators are available.

  • BioStation users will need to go through training before scheduling a run in the BioStation.
  • BioStation users should consult with the Academic Coordinator to determine when their experiment will fit into the schedule.
  • Investigators are allowed to bring other human primary cell cultures and human stem cell types (for example, adult, fetal, neural and hematopoietic stem cells) to the Core provided they go through Mycoplasma screening. A tissue culture biosafety cabinet in TC Room B will be assigned for use with cells that are not pluripotent.
Tissue Culture Room C

Tissue Culture Room C (TC Room C) will be used exclusively for culturing pluripotent human stem cells. Both normal and low oxygen incubators are available.

  • This room has an inverted Nikon fluorescence microscope and camera.

7. Tissue Culture Equipment Room

  • The Core houses a -80oC freezer, a liquid nitrogen tank, and an x-ray irradiator in the Equipment Room.
  • Each PI will be assigned a specific shelf area in the -80oC freezer.
  • For cell cryopreservation, researchers will be allocated a space in the cryogenic liquid nitrogen tank. The Academic Coordinator or Lab Assistant will be responsible for freezing and retrieving frozen cells and documenting the cell freezing log.
  • Any animal cells can be irradiated with the x-ray irradiator provided they are brought to the Core in 15 ml sealed tubes (cells can not be brought to the irradiator in dishes or flasks). Tubes must remain sealed while they are in the Core. Cells collected and irradiated in tubes do not require Mycoplasma screening prior.

More Information 

General Campus Information

University of California, Riverside
900 University Ave.
Riverside, CA 92521
Tel: (951) 827-1012

Facility Information

Stem Cell Core Facility
Next to 501A Keen Hall

in association with:
College of Natural and Agricultural Sciences
College Building North, floors 2 & 3

Tel: (951) 827-6555
Fax: (951) 827-4190

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