Virus Purification And Isolation Using Viraffinity™ For Anti-HIV Drug Development Research

Oct. 7, 2012 - PRLog -- MONMOUTH JUNCTION, NJ.– Authors Abdurahman et al published an article in the journal Antimicrobial Agents and Chemotherapy titled, Activity of the Small Modified Amino Acid α-Hydroxy Glycineamide on In Vitro and In Vivo Human Immunodeficiency Virus Type 1 Capsid Assembly and Infectivity which cites Viraffinity™ from Biotech Support Group for virus purification. Researchers studied the role of alpha-hydroxy-glycineamide (alpha-HGA), a metabolite with antiviral specificity towards human immunodeficiency virus type 1 (HIV-1) virion. Alpha-hydroxy-glycineamide (alpha-HGA) affects reverse transcriptase or protease inhibitor resistant HIV-1 virion isolates to replicate and prevents the HIV-1 capsid protein from forming tubular or core structures in vitro and in vivo. Scientists developed an assay for virus production which infected HIV-LAI or HeLa-tat cells with HIV-1 clone pNL4-3 in culture medium having alpha-HGA and ACH-2 cells that were stimulated to produce virus.

Virus particles treated with alpha-HGA and untreated with alpha-HGA  in the supernatant  were purified using Viraffinity™ from Biotech Support Group. Viraffinity™ is a unique water-insoluble elastomeric polyelectrolyte that has been engineered for the capture and recovery of viruses. Upon adding Viraffinity™ to the sample, mixing and centrifuging removes cell debris from the culture supernatants. It’s simple protocol is developed especially for high yield recovery of virions, viral proteins or viral nucleic acids.  The centrifuged pellet contains polyelectrolyte-bound viruses that can then be recovered using a moderately alkaline pH solution. Next researchers analyzed the Viraffinity™ purified alpha-HGA treated and untreated HIV-1 clone pNL4-3 or stimulated ACH-2 cell cultures infected HeLA-tat cells and the denatured whole-cell lysates by SDS-polyacrylamide gel electrophoresis.

Presumably Viraffinity™ does not affect whole infectious non-enveloped virus, virions, viral components upon purification and maintains whole infectious non-enveloped virus, virions, viral components function. Major viral proteins were analyzed by western blot and the western blot analysis of lysates showed no effect on the processing of capsid proteins. The purified viruses and lysate samples were transferred to a nitrocellulose membrane and detected with HIV-1 positive human sera. Recombinant proteins of the C-terminal p24 domain were incubated, trypsin digested and the peptide mixtures were analyzed by matrix assisted laser desorption ionization-mass spectrometry (MALDI-MS). Alpha-HGA bound to the N- and C-terminal domains of the HIV-1 capsid protein and affected the ability of the HIV-1 capsid protein to assembly. Development and analysis of lead compounds with anti-HIV specificity and novel mechanisms of antiviral action is possible with Viraffinity™ because it purifies whole infectious non-enveloped virus & non-infectious enveloped virus, isolates antigenic virions, enveloped and non-enveloped, enriches for viral nucleic acids and prepares viral samples for subsequent detection and analysis.

Please contact Biotech Support Group for more information on Viraffinity™'s application.

About Biotech Support Group LLC
Biotech Support Group LLC is a leading provider of genomics and proteomics sample preparation products and enrichment reagent kits as well as integrated biotechnology services for life sciences research, biomarker and drug discovery. Based in New Jersey, it’s principal products include:  AlbuVoid™ for albumin depletion, Cleanascite™ for lipid adsorption and clarification, NuGel™ for passivated silica-based affinity chromatography, and ProCipitate™ & ProPrep™ for nucleic acid isolation. Biotech Support Group is the leading developer of sample preparation products for separating and purifying hemoglobin from blood samples. HemoVoid™ is  a hemoglobin depletion reagent kit from red blood cells and HemogloBind™ is a hemoglobin capture reagent from hemolyzed serum. Currently, Biotech Support Group LLC and ProFACT Proteomics Inc., are collaborating on the development of a proteomics platform used in functional profiling for proteomic analysis and a separations method for generating sub-proteomes used in biomarker and functional proteomic prospecting. For more information, go to: www.biotechsupportgroup.com

CONTACT:
Dr.Swapan Roy
Biotech Support Group
1 Deer Park Drive, Suite M,
Monmouth Junction, NJ 08852, USA              
732-274-2866
sales@biotechsupportgroup.com

Patents
Composition and utility patents for Viraffinity™ and related technologies are covered under U.S. Patent Numbers 5,294,681, 5,453,493, 5,658,779 and other patents pending.

European Patent Application EP2305809 Methods for nucleic acid isolation and kits using a microfluidic device and concentration step

References

Hanta Virus
Hai-Tao Yu, Hong Jiang, Ye Zhang, Xue-Ping Nan, Yu Li, Wei Wang, Wei Jiang, Dong-Qiang Yang, Wen-Jing Su, Jiu-Ping Wang, Ping-Zhong Wang, and Xue-Fan Bai.Hantaan Virus Triggers TLR4-Dependent Innate Immune Responses. Viral Immunology.2012;

Mihalic, K. A., et al, Development of a Chemiluminescent Western Blot for Detecting Hantaan-Specific Antibodies, poster American Society of Tropical Medicine and Hygiene Meeting, October 1997.

Human Mouth Virus
Metagenomic Analysis of the Human Mouth Virus Population and Characterisation of Two Lytic Viruses, Al-Jarbou, Ahmed, Theses, Dept. of Infection. Immunity and Inflammation.2009

Lepidoptera: Gelechiidae
An isometric virus of the potato tuber moth Tecia solanivora (Povolny) (Lepidoptera: Gelechiidae) has a tri-segmented RNA genome Jean-Louis Zeddam et. al Journal of Invertebrate Pathology, Volume 99, Issue 2, October 2008, Pages 204-211

Human Immunodeficiency Virus Type 1
Mutation in the loop C-terminal to the cyclophilin A binding site of HIV-1 capsid protein disrupts proper virus assembly and infectivity Samir Abdurahman , Stefan Höglund , Anders Höglund and Anders Vahlne. Retrovirology 2007.

Abdurahman S, Végvári A, Youssefi M, Levi M, Höglund S, Andersson E, Horal P, Svennerholm B, Balzarini J, Vahlne A. Activity of the small modified amino acid alpha-hydroxy glycineamide on in vitro and in vivo human immunodeficiency virus type 1 capsid assembly and infectivity. Antimicrobial Agents and Chemotherapy. 2008 Oct;52(10):3737-44.

Poliovirus type 1 (PV1), Hepatitis A virus (HAV), Norwalk virus

Detection Methods for Human Enteric Viruses in Representative Foods. Leggitt, Paris R.1; Jaykus, Lee-Ann, Journal of Food Protection®, Volume 63, Number 12, December 2000 , pp. 1738-1744(7).

Polio Virus Type 1 and Calcivirus

Rapid Detection of Foodborne Viruses from Minimally Processed Foods.Microbial Detection Methods 2000

Polio Virus
Ting, W.T. E., E. M. Nielson, and C.C. Tseng. 1997. The use of Viraffinity matrix to concentrate waterborne polioviruses for RT-PCR detection. Abstr. Q-169. p.484. InAbstracts of the 97th General Meeting of the American Society for Microbiology 1997, American Society for Microbiology, Washington, D.C.

Bacteriophage Lambda DNA
Hitti, J., et al, Fast and Convenient Purification of Bacteriophage Lambda DNA with Viraffinity Matrix, poster Cold Spring Harbor Conference on Genome Mapping & Sequencing, May 1997.

Suggested References:

Gamble, T. R., S. Yoo, F. F. Vajdos, U. K. von Schwedler, D. K. Worthylake, H. Wang, J. P. McCutcheon, W. I. Sundquist, and C. P. Hill. 1997. Structure of the carboxyl-terminal dimerization domain of the HIV-1 capsid protein. Science 278:849-853

Tang, C., E. Loeliger, I. Kinde, S. Kyere, K. Mayo, E. Barklis, Y. Sun, M. Huang, and M. F. Summers. 2003. Antiviral inhibition of the HIV-1 capsid protein. Journal of Molecular Biology. 327:1013-1020

Boden, D., A. Hurley, L. Zhang, Y. Cao, Y. Guo, E. Jones, J. Tsay, J. Ip, C. Farthing, K. Limoli, N. Parkin, and M. Markowitz. 1999. HIV-1 drug resistance in newly infected individuals. JAMA 282:1135-1141