Cell Culture Unit






 

1. Cell Culture Unit. Introduction

The use of in vitro cell culture systems is today an essential technology platform in the development of many scientific investigations. During the last decades, this method has become an indispensable tool in an attempt to unravel the molecular and cellular mechanisms of any biological process.

According to this relevance, GENYO offers an adequate infrastructure devoted to cell culture. The Unit is equipped with three experimental areas and one cryopreservation area installed in various locations of the building. The first one fulfill the most strict requirements for ensuring aseptic conditions thanks to independent air-conditioning, as well as strict access protocols through a safety room. It is divided into two areas, a positive pressure room aimed at the standard culture of cell lines, and a negative pressure room aimed at cell experiments with special biological safety conditions.

The other two experimental areas are suited for cell lines culture with less strict conditions, with equipment for adequate manipulation of embryonic stem cells. The Cryo preservation area is specially designed to locate cryo freezers and liquid nitrogen supply, with the strict compliance of the safety protocols.

1.1 Aim

The main objective of the Cell Culture Unit is to provide support to research in GENYO, and to facilitate the task of researchers of the Center, as well as to provide assistance and services in this work area.

The Unit provides users with the possibility to cultivate and experiment with all cell types of a different nature, both primary culture and established cell lines. It also offers to users the possibility to perform a wide variety of techniques for the study of cellular and molecular characterization of cell culture, since it is supported by the Flow Cytometry, Molecular and Imaging Microscopy, and Genomics and Genotyping Units, which are essential in the interpretation of the results obtained in the cell assays.

The Unit is not restricted to provide the equipment necessary for the maintenance, experimentation and cryopreservation in liquid nitrogen of cell culture, but it also offers users scientific and technical assistance, ,support, and services such as detection of mycoplasma, and the adequate treatment in case of contamination, as well as the extraction and isolation of primary cultures, transfection assays, cell line maintenance , cell bank, and the optimal supply of common reagents for cell culture.

2. Areas of activities

Cell culture experiments require adequate conditions for their optimal development, such as aseptic environment, strict monitoring equipment, and the constant evaluation of cell cultures to avoid contamination with pathogenic agents as well as their correct cryopreservation. These areas of activities are essential to provide a quality service that the Unit is intended to offer to researchers of GENYO.

2.1 Asepsis

The Unit is equipped with laminar flow cabinets with different specifications able to guarantee sterility of the work area. Also, all the incubators are equipped with the US AutoFlow with a filtration system able to minimize potential cross-contamination problems within the chamber.

In order to guarantee optimal aseptic conditions required in several experimental processes, the area located in the first floor is equipped with a triple absolute air filtration with overpressure system, avoiding unfiltered air intake from outside.

2.2 Monitoring

The technical staff carries out strict control of the correct functioning and maintenance of all the facilities and equipment of the Unit, through specific protocols. Pressure and temperature parameters of the incubators, laminar flow cabinets, fridge and freezers temperature, as well as other relevant parameters are daily controlled.

2.3 Detection of contaminants

Contamination of cell cultures with bacteria, fungi, virus, and yeasts represents a major problem in cell culture. These contaminants may lead to the death of the cell culture or the alteration of its properties. Whereas these microorganisms are easily detected during routine cell culture by observation under microscope, one class of bacteria known as mycoplasma regularly evades detection, therefore it is important to perform periodical controls to determine its possible presence in the cell culture.

Among the large variety of methods available for the detection, the technical staff uses the most rapid and sensitive methods for identifying false-positives or other ambiguities.

2.4 Cryopreservation

The use of adequate storage methods is essential to minimize the accumulation of genetic alterations in established cell lines, as well as to avoid senescence and transformation of primary cell culture, or the accidental loss of cell culture due to death or contamination of the sample, Today, the best method is the preservation in liquid nitrogen. The Unit is equipped with a cryopreservation room, specially fitted with cryo freezers connected to a general supply system that guarantees the maintenance of the optimal conditions for cryopreservation.

3. Equipment

The Unit is equipped with:

Laminar flow cabinets::

  • Class II, type A vertical laminar-flow biological cabinets, which keep the work area free of contamination and guarantee the highest protection of the operator and environment.
  • Horizontal laminar flow cabinets, which guarantee the maximal protection of the product from contamination.

Incubators:

This equipment includes a CO2 centralized system that ensures constant and automated supply of it, as well as nitrogen gas.

Centrifuges:

Microscopios:

Refrigerators and freezers:

Which ensure the safe storage of reagents and solutions

Cryofreezers:

  • Programmable cryo freezers

Which ensure the correct preservation and viability of cells stored for long periods of time.

Other equipment available:

These facilities are essential for the good functioning of the Unit.

4. Services

  • We provide scientific advice to users of the Unit.
  • The Unit provides management of the full use of the facilities and maintenance of the infrastructure to obtain optimal performance.
  • Supervision to comply with the working rules of the Unit in order to avoid cell culture contamination, by requiring the users good working habits of the facilities of the Unit.
  • Periodic and exhaustive control of the facilities and equipment, to ensure the supply of consumable items and common reagents.
  • Control of critical parameters for cell culture, such as temperature and CO2 supply to incubators, and levels of liquid nitrogen in the cryo preservation tanks.
  • Design of a cleaning, disinfection and/or sterilization of the instruments used in the cell culture rooms.
  • Performance of assays for detecting mycoplasma, contamination and adequate treatment of contaminated cell culture.
  • Extraction and isolation of primary cell culture, infection and transfection of cell lines.
  • Maintenance and experiment of cells for groups without personnel specialized in cell culture.

5. Useful Links

American Type Culture Collection (ATCC) (USA)

Cell Culture Protocols Online

European Collection of Cell Lines (ECACC)

Fresh Human Tumor Cell Culture Atlas

Health Protection Agency Culture Collection (UK)

International Society for Cellular Therapy (ISCT)

IPD – ESTDAB Database

Red de Terapia Celular

Red Temática de Investigación Cooperativa en Cáncer (RTICC)

6. Staff Contact

The Unit provides a skilled staff with extensive scientific and technical experience. Dr. Juan Carlos Rodríguez-Manzaneque, is the Scientific Manager of the Genomic Oncology Area of GENYO. He has large experience in the field of cell biology and has authored many works, in particular those on endothelial biology and angiogenesis. The use of cell systems has been a constant in his scientific career, particularly specialized during the last years in 3D cell growth models in matrices of different characteristics in an attempt to mimic physiological and pathological contexts.

Mr. Víctor García Cabrera is the Technical Manager: He has a large experience in cell culture as well as in molecular biology techniques and maintenance of cell culture rooms. He is also responsible for the management and coordination of the functioning Unit under the supervision of the Scientific Manager.

Contact
Víctor García Cabrera, Technical Manager.
unidad.cultivos@genyo.es

Application:

If you have never been a user associated with this Service, you have to implement this Application Form to register as user, and send it to: victor.garcia@genyo.es.

The Cell Culture Unit provides the infrastructure and technical support necessary, as well as the appropriate advice about the best conditions required to each project, based on the means available. Once registered in the Unit, and depending on the type of study you want to perform, you will be assigned to the most adequate working area suited to your needs.