Immunocapture

A variety of methods have been used to either physically isolate and / or detect dsRNA using our antibodies. These include ELISA1,2 (for quantitative determination of dsRNA), sandwich ELISA1, dot-blots2,3,4 (for quantitation, specificity testing), immunoprecipitation5,6 and immunoblotting1,7 (for detection and characterisation of dsRNA separated by electrophoresis in polyacrylamide gels).Képre kattintva az ablak bezáródik.

 



J2 antibody is used in dot blots to detect dsRNA from Leishmania RNA virus (LRV) in the Leishmania parasite L. guyanensis. A serial dilution of 1000 to 10 parasites from LRV-positive and negative control strains - Lg4147LRVhigh and LG4147LRVneg respectively - is shown. Picture taken from Zangger et al. (2013) PLoS Negl Trop Dis 7:e2006.


Due to the high affinity and sequence-independent binding of our antibodies, they have also been used  to detect dsRNA contaminants in RNA produced for therapeutic purposes, primarily using dot-blots3. Promising developments in gene therapy suggest such tests may provide a straightforward strategy for quality-control of in vitro produced mRNA molecules. The publications detailing such experiments can be found in our database of publications, searching for "non-viral applications" or "contaminating dsRNA detection".

 

1Schönborn J, Oberstrass J, Breyel E, Tittgen J, Schumacher J, 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

2Zangger H, Ronet C, Desponds C, Kuhlmann FM, Robinson J, Hartley MA, Prevel F, Castiglioni P, Pratlong F, Bastien P, Müller N, Parmentier L, Saravia NG, Beverley SM, Fasel N. (2013) Detection of Leishmania Virus in Leishmania parasites. PLoS Negl Trop Dis. 7:e2006.

3Bauhofer O, Summerfield A, McCullough KC, Ruggli N. (2005) Role of double-stranded RNA and Npro of classical swine fever virus in the activation of monocyte-derived dendritic cells. Virology 343:93-105.

4Kariko K, Muramatsu H, Ludwig J, Weissman D. (2011) Generating the optimal mRNA for therapy: HPLC purification eliminates immune activation and improves translation of nucleoside-modified, protein-encoding mRNA. Nucleic Acids Res 39:e142

5Kaneko H, Dridi S, Tarallo V, Gelfand BD, Fowler BJ, Cho WG, Kleinman ME, Ponicsan SL, Hauswirth WW, Chiodo VA, Kariko K, Yoo JW, Lee DK, Hadziahmetovic M, Song Y, Misra S, Chaudhuri G, Buaas FW, Braun RE, Hinton DR, Zhang Q, Grossniklaus HE, Provis JM, Madigan MC, Milam AH, Justice NL, Albuquerque RJ, Blandford AD, Bogdanovich S, Hirano Y, Witta J, Fuchs E, Littman DR, Ambati BK, Rudin CM, Chong MM, Provost P, Kugel JF, Goodrich JA, Dunaief JL, Baffi JZ, Ambati J. (2011) DICER1 deficit induces Alu RNA toxicity in age-related macular degeneration. Nature 471:325-330

6Cruz C, Housley J. (2014)  Endogenous RNA interference is driven by copy number. eLife 2014;3:e01581

7Lukacs 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