The QC team is responsible for testing the following types of samples:
- Raw Materials
- Pre-GMP Starting Materials
- In-Process Samples
- Final Product testing
- Stability testing
All samples for QC testing or long-term sample storage are recorded and allocated a unique sample number. Internal tests are carried out according to defined SOPs and the results are documented in the associated Forms. In some instances results are recorded in Laboratory Notebooks as appropriate. All results are then checked, reviewed and approved by the authorized person.
The following test methods are available at the CBF and are performed internally:
- Visual appearance
- Sterility check
- Endotoxin by the chromogenic kinetic method
- Enzyme-linked immunosorbent assay (ELISA)
- Infectious titre by immunostaining
- Virus particle number by spectrophotometric analysis
- Viral particle concentration by real-time quantitative PCR (This is for information only as it is not yet fully validated)
- Vector insert presence and sequence verification by polymerase chain reaction (PCR) and Sanger sequencing
- Sequencing by the massively parallel sequencing technology known as next-generation sequencing (NGS). This assay is for information only as it is not yet fully validated.
- Restriction enzyme analysis of viral genome
- Protein concentration by absorbance at 280 nm
- Gel electrophoresis and Western blotting
Analytical Assay Validation
Validation and Verification of Analytical Methods is covered by internal CBF SOPs. Wherever possible, assays used within the CBF use spiked samples and standards to confirm the suitability of assays and validity of the results derived therefrom. Assays used in GMP analysis are typically developed during the development stage of a project and therefore full validation of an assay may not be possible for the early phase trials. Risk assessment is used when considering assay qualification in order to assess product quality and minimise volunteer / patient risk from clinical-grade products manufactured at the CBF. The QC is also responsible for managing a number of tests that are performed externally by Contract Research Organization (CRO). Some of the tests performed listed below, depending on the stage of the purification process.
- Microbial Identification by gram staining and MALDI-ToF
- Adventitious Virus Detection, such as 28 day in vitro assay for the detection of virus contaminants, with blind passage; Cytotoxicity study on three indicator cell lines; Detection of Toxicity or Test Article Breakthrough (Post Neutralisation) in Suckling Mice and/or Embryonated Eggs; In Vivo Assay for Extraneous Agents in Viral Vaccines for Human Use according to European Pharmacopoeia using Suckling Mice and Embryonated eggs (Ph.Eur.2.6.16)
- Replication Competent Adenovirus assay using A549 detector cells.
- Microbial Contaminants, such as Test for Mycoplasma spp (Including Mycoplasmastasis Assay) (Eu Pharm Current Edition; Section 2.6.7 Mycoplasmas/USP - NF Mycoplasma Tests, USP <63> Current Edition)
- Retrovirus Detection by assaying for oReverse Transcriptase (RT) by Real Time Fluorescent Product Enhanced Reverse Transcriptase (F-PERT) assay
- Species Specific Virus Detection such as detection of Adeno Associated Virus (AAV) by Real Time Polymerase Chain Reaction (PCR)
- Residual Process Related Impurity Tests such as Detection and Quantitation of HEK 293 HCP by ELISA; Detection and Quantitation of Bovine Serum Albumin (BSA) Contaminants by ELISA; Detection and Quantitation of Benzonase Endonuclease by ELISA; Quantitation of Residual Human Embryonic Kidney (HEK) 293 host cell DNA by Real Time Polymerase Chain Reaction (PCR); and detection of Residual CsCl by Mass Spectrometry-HPLC
- Sterility Test and Qualification of Test Article by Direct Inoculation Method (Eu Pharm Current Edition; Section 2.6.1 Sterility and USP Current Edition <71> Sterility Tests)
- Osmolality testing
- Extractable volume