Clone: J5
Background: Over the past decade our double-stranded RNA (dsRNA)antibodies have been used extensively to detect and characterise plant and animal viruses with dsRNA genomes or intermediates. In addition, the anti-dsRNA antibodies can be used as a diagnostic tool to detect pathogens, including detection in paraffin-embedded fixed tissue samples (Richardson et al. 2010).The J5 IgG2b antibody recognizes dsRNA with very similar affinity and specificity to our J2 antibody (see Schonborn et al., 1991), but has a different isotype – thus allowing more flexibility for the simultaneous detection of dsRNA with other markers, particularly in immunofluorescence microscopy, and has been used to detect replicative intermediates of the fish virus Infectious Pancreatic Necrosis Virus (IPNV) (Levican-Asenjo et al., 2019) or of ECMV in Vero cells.The J5 antibody can detect all tested forms of dsRNA, including poly(A):poly(U), poly(I):poly(C) and dsRNA from viruses such as Dengue Virus, Encephalomyocarditis Virus, Vaccinia Virus, Reovirus or Cucumber Mosaic Virus. Similarly to our other antibodies dsRNA-binding of J5 is sequence-independent, as long as the length of the dsRNA exceeds 40nt. The antibody does not react with ssRNA, ssDNA or dsDNA. J5 has been tested successfully in nucleic acid ELISA, immunoblotting and immunofluorescence microscopy.
Purification Method: Affinity chromatography on Protein A-agarose.
Concentration: Concentration after reconstitution: 1.00 mg/ml as determined by A280 nm (A280 nm = 1.47 corresponds to 1 mg/ml antibod
Source: Female DBA/2 mice were injected intraperitonially with a mixture of 50 ug L-dsRNA and 75 ug methylated bovine serum albumin, emulsified in complete Freund's adjuvant. After several boosts spleen cells were fused with Sp2/0-Agl4 myeloma cells to generate the hybridoma clone.
Purity: Gel electrophoretically pure IgG antibody.
Formulation: The lyophilised sample should be reconstituted with 200 µl sterile distilled water. The mAb will then be in PBS without any stabilisers or preservatives at a concentration of 1 mgr/ml. As a result of the lyophilisation procedure, the reconstituted antibody may contain small amounts of denatured protein in the form of aggregates that may interfere with some applications such as immunohistochemistry (e.g. by giving high backgrounds). We therefore highly recommend centrifuging (microcentrifuge) the reconstituted antibody before use and using the supernatant.
References: 1) Schönborn, J., Oberstrass, J., Breyel, E., Tittgen, J., Schumacher, J. and Lukacs, N. (1991) Monoclonal antibodies to double-stranded RNA as probes of RNA structure in crude nucleic acid extracts. Nucleic Acids Res.19, 2993-3000. 2) Lukacs, N. (1994) Detection of virus infection in plants and differentiation between coexisting viruses by monoclonal antibodies to double-stranded RNA. J. Virol. Methods 47, 255-272. 3) Lukacs, N. (1997) Detection of sense:antisense duplexes by structurespecific anti-RNA antibodies. In: Antisense Technology. A Practical Approach, C. Lichtenstein and W. Nellen (eds), pp. 281-295. IRL Press, Oxford.4) S. J. Richardson, A. Willcox, D. A. Hilton, S. Tauriainen, H. Hyoty, A. J. Bone, A. K. Foulis, N. G. Morgan. Use of antisera directed against dsRNA to detect viral infections in formalin-fixed paraffin-embedded tissue. J Clin Virol. (2010) 49(3); 180-5. doi: 10.1016/j.jcv.2010.07.015.5) Levicán-Asenjo J, Soto-Rifo R, Aguayo F, Gaggero A, Leon O. Salmon cells SHK-1 internalize infectious pancreatic necrosis virus by macropinocytosis. J Fish Dis. 2019 Jul;42(7):1035-1046. doi: 10.1111/jfd.13009.
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