Virus Counter
Encyclopedia
The Virus Counter is an instrument for rapid quantification of virus
es in liquid samples. It is a specialized flow cytometer
that uses high-sensitivity fluorescence detection to give a direct measurement of the concentration of virus particles in a fraction of the time required for traditional plaque assays.
The Single Nanometric Particle Enumerator (SNaPE) was developed in 2001 and evaluated using respiratory viruses. The SNaPE research instrument was based on the principle of fluorescence detection from single stained nucleic acids aggregates. While the quantification results from this instrument correlated well with expected virus concentration, the values tended to be significantly higher than those obtained by standard plaque titer methods typically used for virus quantification
. The measured concentrations were similar in magnitude to qPCR results, both being substantially higher than plaque titer values. While this early, simple version of the instrument provided proof of concept using several viruses from commercial and collaborator sources, it was not ideal for commercial application.
The next step of the Virus Counter’s evolution also took place in the Rowlen’s lab at UC-Boulder as Carrie Stoffel’s Ph.D. project. By 2004 Carrie had added a second detection channel to the SNaPE instrument and improved data analysis substantially to increase specificity. The principle of detection was based on staining proteins and nucleic acids with fluorescent dyes within a liquid sample, where the protein stain emits in the red region of the visible spectrum and the nucleic acid stain emits in the yellow region of the visible spectrum. As stained particles pass through the probe region and interact with a laser, emissions are detected and recorded for analysis. This dual channel system established the concept of measuring intact virions through detection of co-localized protein and nucleic acids. Results from baculovirus
analyzed with this dual-channel virus counter system were lower in magnitude than the SNaPE results, being more similar in magnitude and correlated with infectious assay results. This system was validated using several viruses from commercial sources as well as real-world samples from collaborators.
A commercial version of the Virus Counter is now available from InDevR
Inc. The instrument is representative of the next generation of “personal” flow cytometers designed exclusively for a niche application, in this case virus quantification. Similar to virus concentration determined by transmission electron microscopy, the Virus Counter provides a measurement of the total number of virus particles per mL. The Virus Counter has been successfully demonstrated to readily quantify a diverse range of viruses, including influenza
, adenovirus, dengue, baculovirus
, respiratory syncytial
, parainfluenza, rubella
, cytomegalovirus
, and herpes simplex
virus.
The ability to rapidly quantify viruses has high impact in:
Virus
A virus is a small infectious agent that can replicate only inside the living cells of organisms. Viruses infect all types of organisms, from animals and plants to bacteria and archaea...
es in liquid samples. It is a specialized flow cytometer
Flow cytometry
Flow cytometry is a technique for counting and examining microscopic particles, such as cells and chromosomes, by suspending them in a stream of fluid and passing them by an electronic detection apparatus. It allows simultaneous multiparametric analysis of the physical and/or chemical...
that uses high-sensitivity fluorescence detection to give a direct measurement of the concentration of virus particles in a fraction of the time required for traditional plaque assays.
History and Development
The Virus Counter’s roots can be traced back to work done in Professor Kathy Rowlen’s lab at the University of Colorado in Boulder as part of the Ph.D. thesis project of Matthew Ferris.The Single Nanometric Particle Enumerator (SNaPE) was developed in 2001 and evaluated using respiratory viruses. The SNaPE research instrument was based on the principle of fluorescence detection from single stained nucleic acids aggregates. While the quantification results from this instrument correlated well with expected virus concentration, the values tended to be significantly higher than those obtained by standard plaque titer methods typically used for virus quantification
Virus Quantification
Virus quantification involves counting the number of viruses in a specific volume to determine the virus concentration. It is utilized in both research and development in commercial and academic laboratories as well as production situations where the quantity of virus at various steps is an...
. The measured concentrations were similar in magnitude to qPCR results, both being substantially higher than plaque titer values. While this early, simple version of the instrument provided proof of concept using several viruses from commercial and collaborator sources, it was not ideal for commercial application.
The next step of the Virus Counter’s evolution also took place in the Rowlen’s lab at UC-Boulder as Carrie Stoffel’s Ph.D. project. By 2004 Carrie had added a second detection channel to the SNaPE instrument and improved data analysis substantially to increase specificity. The principle of detection was based on staining proteins and nucleic acids with fluorescent dyes within a liquid sample, where the protein stain emits in the red region of the visible spectrum and the nucleic acid stain emits in the yellow region of the visible spectrum. As stained particles pass through the probe region and interact with a laser, emissions are detected and recorded for analysis. This dual channel system established the concept of measuring intact virions through detection of co-localized protein and nucleic acids. Results from baculovirus
Baculovirus
The baculoviruses are a family of large rod-shaped viruses that can be divided to two genera: nucleopolyhedroviruses and granuloviruses . While GVs contain only one nucleocapsid per envelope, NPVs contain either single or multiple nucleocapsids per envelope. The enveloped virions are further...
analyzed with this dual-channel virus counter system were lower in magnitude than the SNaPE results, being more similar in magnitude and correlated with infectious assay results. This system was validated using several viruses from commercial sources as well as real-world samples from collaborators.
A commercial version of the Virus Counter is now available from InDevR
InDevR
InDevR is a biotechnology company that develops advanced life science instrumentation and assays for analysis of viruses and other microorganisms, with product focus on Virus Quantification and pathogen detection / identification. InDevR Inc. is a privately held, woman-owned small business located...
Inc. The instrument is representative of the next generation of “personal” flow cytometers designed exclusively for a niche application, in this case virus quantification. Similar to virus concentration determined by transmission electron microscopy, the Virus Counter provides a measurement of the total number of virus particles per mL. The Virus Counter has been successfully demonstrated to readily quantify a diverse range of viruses, including influenza
Influenza
Influenza, commonly referred to as the flu, is an infectious disease caused by RNA viruses of the family Orthomyxoviridae , that affects birds and mammals...
, adenovirus, dengue, baculovirus
Baculovirus
The baculoviruses are a family of large rod-shaped viruses that can be divided to two genera: nucleopolyhedroviruses and granuloviruses . While GVs contain only one nucleocapsid per envelope, NPVs contain either single or multiple nucleocapsids per envelope. The enveloped virions are further...
, respiratory syncytial
Human respiratory syncytial virus
Human respiratory syncytial virus is a virus that causes respiratory tract infections. It is the major cause of lower respiratory tract infections and hospital visits during infancy and childhood. A prophylactic medication exists for preterm birth infants and infants with a congenital heart...
, parainfluenza, rubella
Rubella
Rubella, commonly known as German measles, is a disease caused by the rubella virus. The name "rubella" is derived from the Latin, meaning little red. Rubella is also known as German measles because the disease was first described by German physicians in the mid-eighteenth century. This disease is...
, cytomegalovirus
Cytomegalovirus
Cytomegalovirus is a viral genus of the viral group known as Herpesviridae or herpesviruses. It is typically abbreviated as CMV: The species that infects humans is commonly known as human CMV or human herpesvirus-5 , and is the most studied of all cytomegaloviruses...
, and herpes simplex
Herpes simplex
Herpes simplex is a viral disease caused by both Herpes simplex virus type 1 and type 2 . Infection with the herpes virus is categorized into one of several distinct disorders based on the site of infection. Oral herpes, the visible symptoms of which are colloquially called cold sores or fever...
virus.
The ability to rapidly quantify viruses has high impact in:
- Vaccine Research, Development, and Manufacturing
- Antiviral Research
- Viral VectorViral vectorViral vectors are a tool commonly used by molecular biologists to deliver genetic material into cells. This process can be performed inside a living organism or in cell culture . Viruses have evolved specialized molecular mechanisms to efficiently transport their genomes inside the cells they infect...
Research and Protein Expression
- Bioprocess Optimization
- Viral Antigen Characterization
- Viral Diagnostic Standard Characterization
- Viral-Based Gene TherapyGene therapyGene therapy is the insertion, alteration, or removal of genes within an individual's cells and biological tissues to treat disease. It is a technique for correcting defective genes that are responsible for disease development...
Research