Requirments for primary human hepatocyte

5.1 Source materials and ancillary materials

The collection of human biological source material (source material) shall comply with domestically recognized ethics and local laws and regulations.

Written and valid informed consent shall be signed by the donor. The content of informed consent shall include but not be limited to the research purpose, potential research and clinical diagnosis application under appropriate conditions feedback on unexpected discoveries potential commercial value and other issues affiliated to the research. Mechanisms for protecting the personal data and privacy of donors shall be established.

The cell research and production organizations shall establish and implement cell donor evaluation standards, and establish collection methods, transportation standards, handover standards and storage standards to ensure the safety of donors and cells. The source material and the derivative cells shall be accompanied with detailed documentation on acquisition methods, source and relevant clinical information, including but not limited to the donor's general information, past medical history and family history. The past medical history and family history shall be collected in detail for the information related to genetic diseases. The information of the donor's ABO blood type and the human leukocyte antigen (HLA) type I and type II classification shall be collected.

The origin of cells shall be traceable by referring to the relevant informed consent and/or their genome and functional data.

Ancillary materials such as culture medium and growth factors shall meet the corresponding quality control requirements. The ancillary materials shall be inspected and laboratory tested if necessary.

When using animal serum, they shall be demonstrated to be of low risk of viral contamination and donor testing performed where appropriate. Serum from animals in geographical regions with known prion epidemics (e.g. bovine spongiform encephalopathy) shall be prohibited.

If human blood components are used in the culture medium, including but not limited to albumin, transferrin and various cytokines, the source, batch number and quality verification reports shall be provided. State-approved meterials shall be used where feasible.

The source material donors should be screened for HIV, HAV, HBV, HCV, HTLV and TP, and the results should be recorded.

The isolation and acquisition of cells shall be performed in a primary (or secondary) biosafety laboratory.

5.2 Primary quality attributes 5.2.1 Cell morphology

Primary human hepatocyte cultured in 2D conditions shall exhibit a polygonal morphology with a diameter of 20–30 µm under the optical microscope. Cells can have single, dual or multiple nuclei that are large and round in shape. Tight junctions and Bile duct can be found between cells in the adherent cultures.

5.2.2 Cell viability

The cell viability shall be ≥70% after resuscitation.

5.2.3 Cell markers

The expression of ALB and HNF4A shall be ≥90% of the cell population.

5.2.4 Albumin secretion

The Albumin secretion shall be ≥800 ng/(106 cells × 24 h).

5.2.5 Drug metabolism function

The representative drug metabolism ability of PHHs (e.g. CYP3A4, testosterone as the substrate) indicated by the intrinsic clearance rate shall be ≥100 μl/(h × 106 cells).

5.2.6 Bile secretion (applicable to adherent cells only)

The BEI index (d8-TCA as the substrate) shall be ≥30%.

5.2.7 Microorganisms

Primary human hepatocytes shall be negative for bacteria, fungi, mycoplasma, HIV, HAV, HBV, HCV, HTLV, EBV, HCMV and TP.

5.3 Process control 5.3.1 Cell isolation

During the cell isolation process, cross-contamination and confusion between donors, tissues and cells shall be avoided and a risk management strategy shall be developed.

During the process of culture and expansion after cell isolation, the passage and the cells name shall be clearly specified and the operation date, culture conditions, operators' names or initials shall be indicated.

5.3.2 Cell isolation process and cell identification

The equipment, the culture system and the operation steps used for cell isolation shall be clarified. The standard operating procedure for cell isolation shall be established for reproducibility.

The isolated cells shall have the defined morphological, molecular and functional characteristics listed in 5.2.

5.3.3 Cryopreservation

Cryopreserved PHHs shall be clearly labelled with the cell name, the culture conditions, the passage number, the operators' names or initials and the cryopreservation date. Cryopreserved cells shall have the same unique identification used during the processes of collection, isolation and culturing.

The cryopreservation procedure shall follow the known principles of cryopreservation of mammalian cells and shall be recorded according to the relevant regulations.

5.3.4 Resuscitation

The resuscitation process shall be as short as possible to ensure the optimal viability and maintenance of biological functions of PHHs.

The cell information, including but not limited to the cell name, the batch name, the passage number, the culture conditions, operators' names or initials and resuscitation date, shall be documented and recorded.

5.3.5 Identification of cell STR

The short tandem repeat (STR) signature of PHHs shall be consistent with that of the donor.

留言 (0)

沒有登入
gif