Mouse anti double-stranded RNA (J5)

Mouse monoclonal antibody J5 can be used for ELISA, dsRNA-immunoblotting, immunoaffinity chromatography and in certain systems also for immunohistochemistry (see references). Please note that nucleic acid separation prior to dsRNA-immunoblotting must be carried out by polyacrylamide gel electrophoresis, because the sensitivity of detection is considerably lower after blotting from agarose gels.

Not for use for clinical purposes. For in vitro use only.

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.

Concentration after reconstitution: 1.00 mg/ml as determined by A280 nm (A280 nm = 1.47 corresponds to 1 mg/ml antibod

Microbiology

The lyophilised sample should be reconstituted with 500 ?l 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.

Mouse monoclonal antibody J5 recognises double-stranded RNA (dsRNA) provided that the length of the helix is greater than or equal to 40 bp. dsRNA-recognition is independent of the sequence and nucleotide composition of the antigen. All naturally occurring dsRNAs investigated up to now (40-50 species) as well as poly(I).poly(C) and poly(A).poly(U) have been recognised by J5, although in some assays its affinity to poly(I).poly(C) is about 10 times lower than that to other dsRNA antigens

Affinity chromatography on Protein A-agarose.

Gel electrophoretically pure IgG antibody.

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.

Mouse anti dsRNA