Anti-FLAG tag antibody, clone 40G6.I9 [HRP]

RNA interference offers therapeutic opportunities for the clinical targeting of otherwise undruggable oncogenes. However RNAi can have off-target effects that considerably increase treatment risks. To manage these side effects and allow an easy subtraction of their activity in healthy tissues, we present here the TAG-RNAi approach where cells that are not designated targets do not have the mRNA tag. Using TAG-RNAi we first established the off-target signatures of three different siRNAs specific to the Cyclin D1 oncogene by RNA-sequencing of cultured cancer cells expressing a FLAG-HA-tagged-Cyclin D1. Then, by symmetrical allografts of tagged-cancer cells and untagged controls on the left and right flanks of model mice, we demonstrate that TAG-RNAi is a reliable approach to study the functional impact of any oncogene without off-target bias. Finally we show, as examples, that mutation-specific TAG-RNAi can be applied to downregulate two oncogenic mutants, KRAS-G12V or BRAF-V600E, while sparing the expression of the wild-type proteins. TAG-RNAi will thus avoid the traditional off-target limitations of RNAi in future experimental approaches.

Objective. This study aimed to clarify the clinical features, the serum level of autoantibodies, and cytokine of myositis patients with anti-EJ antibody, which targets glycyl tRNA-synthetase (GlyRS). Methods. Sera of 236 Chinese patients with myositis were screened for anti-EJ by a novel immunoprecipitation assay of flag-tagged GlyRS. Anti-EJ positive patients are evaluated for the clinical features and cytokine profile. Results. The sera from 4 of 236 adult myositis patients were found to carry the anti-EJ using established novel immunoprecipitation assay and immunoblotting. The prevalence of anti-EJ in our cohorts is about 1.7%. The decline of anti-EJ level was detected in two patients during disease remission. Interstitial lung disease and muscle weakness, but not skin involvement, are common clinical features of anti-EJ positive patients. Moreover, using a cytokine profile analyses, we found that the serum levels of IP-10, IL-6, MCP-1, and VEGF were significantly elevated in patients with anti-EJ and gradually decreased during disease remission of two patients, whereas IL-8 level was obviously reduced in these patients. Conclusion. The novel immunoprecipitation assay is suitable to detect and monitor the levels of anti-EJ autoantibody. The serum levels of anti-EJ, IP-10, IL-6, MCP-1, and VEGF may be related to disease activity in myositis patients with anti-EJ antibodies.