Choice of Virus
Spiking experiments can be performed using either relevant viruses (e.g. HIV or HAV for the validation of human plasma) or model viruses (the bovine viral diarrhoea virus (BVDV) is a model for the hepatitis C virus, for example). The viruses to be used are chosen according to the initial risk; for each given situation, the choice of viruses must be adapted to suit the product and the manufacturing process, and must take into account the following parameters:
- the level of risk associated with the starting material.
- emerging viruses.
- the need to cover a broad spectrum of viruses in terms of size, class and resistance to physical & chemical treatments.
Since 1987, Texcell has benefited from the Institute Pasteur's state-of-the-art expertise in virology by carrying out virus validation studies with well-characterised strains.
Today, we offer well-documented and validated titration methods for each virus. The detection limits and the cell passages used for titration are specified in our in-house qualification files.
The availability of more than 25 viruses allows Texcell to cover a broad range of resistances and structural & genomic types and to offer you the most specific choice of viruses according to your product and the corresponding regulatory requirements.
List of available viruses
| Virus | Family | Origin | size (nm) | Genom | Envelope | resistance |
| Adenovirus type 5 | Adenoviridae | human | 70-90 | DNA | non-env | very high |
| Baculovirus | Baculoviridae | Insect | 45-100 | DNA | Enveloped | medium |
| Bovine viral diarrhea virus | Flaviviridae | Bovine | 45-55 | RNA | Enveloped | low |
| Canin parvovirus | Parvoviridae | Canine | 18-26 | DNA | non-env | very high |
| Encephalomyocarditis virus | Picornaviridae | Murine | 28-30 | RNA | non-env | medium |
| Equine rhinovirus | Picornaviridae | Equine | 28-30 | RNA | non-env | medium |
| hepatitis A virus | Picornaviridae | human | 28-30 | RNA | non-env | high |
| Herpes simplex virus type 1 | HerpesViridae | human | 150-200 | DNA | Enveloped | medium |
| Human Immunodeficiency virus 1 | Retroviridae | human | 80-100 | RNA | Enveloped | low |
| Human rhinovirus | Picornaviridae | human | 28-30 | RNA | non-env | medium |
| Influenza virus H3N8 | Orthomyxoviridae | Equine | 80-120 | RNA | Enveloped | medium |
| Minute virus of mice | Parvoviridae | Murine | 18-26 | DNA | non-env | very high |
| Moloney Murine Leukemia virus (EcoT.) | Retroviridae | Murine | 80-100 | RNA | Enveloped | low |
| Murine Leukemia virus(4070A)(amphoT) | Retroviridae | Murine | 80-100 | RNA | Enveloped | low |
| Parainfluenza 3 virus | Paramyxoviridae | human | 150-300 | RNA | Enveloped | low |
| Poliovirus Sabin 1 | Picornaviridae | human | 28-30 | RNA | non-env | medium |
| Poliovirus Saukett | Picornaviridae | human | 28-30 | RNA | non-env | medium |
| Porcine parvovirus | Parvoviridae | Porcine | 18-26 | DNA | non-env | very high |
| Pseudorabies (Aujeszky) | HerpesViridae | Porcine | 150-200 | DNA | Enveloped | medium |
| reovirus type 3 | Reoviridae | human | 60-80 | RNA | non-env | medium |
| Simian virus 40 | Polyomaviridae | Simian | 45-100 | DNA | non-env | very high |
| Sindbis virus | Togaviridae | human | 40-60 | RNA | Enveloped | low |
| Theiler virus | Picornaviridae | Murine | 28-30 | RNA | non-env | medium |
| Transmissible gastroenteritis virus | coronaviridae | Porcine | 80-120 | RNA | Enveloped | medium |
| Vaccine virus | Poxviridae | human | 170-260 | DNA | Enveloped | high |
| Vesiculo stomatitis virus | Rhabdoviridae | Porcine | 45-100 | RNA | Enveloped | low |
Preparation of virus stocks
The virus stocks that we produce for spiking experiments are well characterised and optimised for each kind of step to be evaluated:
- particular care is paid to the protein concentration of the viral stocks used when validating nanofiltration steps: it has to be as low as possible in order to avoid membrane clogging.
- aggregates must be removed prior to use of a virus stock for the validation of a nanofiltration step.
- for strong inactivation steps (NaOH 1N, autoclaving, etc.), we select the viral stocks that have the highest titres - allowing achievement of a high LRV (provided that no infectious particles are recovered following the treatment).
Titration methods
Several methods can be used to determine the viral titres of samples, depending on the virus in question: plaque assays (detection of plaque forming units, PFUs), focus assays (detection of foci forming units, FFUs) or tissue culture infective dose assays (TCID50). The virus stocks used in spiking experiments have a titre of over 107 PFU/mL, FFU/mL or TCID50/mL. For some viruses, much higher titres are regularly obtained. As stated in paragraph 5.9 of CPMP/BWP/268/95, Version 2, 29 February 1996, "the volume of the virus spiked will be less than 10%", the final theoretical concentration of virus in the contaminated starting material will be: {stock titre} x {spike dilution}. The statistical evaluation of the titre is fully documented in our standard operating procedure, available from Texcell on request.
