anti probdnf  (Bioss)

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Long non-coding SINEUP RNA enhancing BDNF translation prevents dendritic atrophy following an Aβ-peptide challenge

doi: 10.1007/s00018-026-06175-4

Figure Lengend Snippet: SINEUP strategy, specificity and efficacy in targeting BDNF protein upregulation. A-B . Rat BDNF gene structure with schematic representation of BDNF transcripts characterised by multiple alternative splicing of 5’ and 3’UTRs (grey boxes), a coding sequence (CDS, white boxes) and two poly-adenylation sites (polyA). B. SINEUP-PAN and SINEUP-Ex1 annealing sites on BDNF transcripts used in this study. C-H. HEK293T cells are co-transfected with different plasmids to demonstrate the efficacy of SINEUP-PAN or SINEUP-Ex1 on the indicated BDNF transcripts with respect to control (GFP). Plasmids were transfected in 1:3 ratio. Tubulin (55 kDa) is indicative of gel loading and is used for normalization in the Western blot analysis. Histograms quantify average data from n = 4 experiments. The molecular weight for the BDNF-GFP band of 64–65 KDa correspond to the predicted size for a chimeric protein made of the proBDNF form (32–34 KDa) and the GFP (27–30 KDa). C . Effect of SINEUP-PAN on protein levels encoded by the transfected chimeric Ex1-BDNF-GFP transcript with respect to control represented by the plasmid encoding the reporter protein GFP alone. p = 0.0005. D . Effect of SINEUP-Ex1 on protein levels encoded by the transfected chimeric Ex1-BDNF-GFP transcript. p = 0.0299. E-F . Effect of SINEUP-PAN on protein levels encoded by the transfected chimeric Ex2B-BDNF-GFP (E) and chimeric Ex2C-BDNF-GFP (F) transcripts. n = 3. G-H . Effect of SINEUP-Ex1 on chimeric Ex2B-BDNF-GFP (G) and chimeric Ex2C-BDNF-GFP (H) transcripts with respect to control (GFP). All the quantitative results shown are produced from a repetition of n = 3 experiments and the Western blot images show the greatest effect obtained and therefore, are not representative of the average results

Article Snippet: The membrane was blocked by 5% non-fat milk in TBST- solution (1 h, RT) and then, the HEK293T cells were incubated with the rabbit polyclonal anti proBDNF antibody (Alomone, Cat. # ANT-006, dilution 1:1000, over-night at 4 °C) while the SH-SY5Y cells were incubated also with the mouse monoclonal anti-BDNF recognizing both BDNF forms (Sigma Cat. #B5050, dilution 1:1000, over-night at 4 °C). and mouse anti alpha tubulin (Sigma, Cat. # T6074, dilution 1:3000) over-night at 4 °C.

Techniques: Alternative Splicing, Sequencing, Transfection, Control, Western Blot, Molecular Weight, Plasmid Preparation, Produced