Anti-TRIM16 Antibody Picoband®

Western blot analysis of TRIM16 using anti-TRIM16 antibody (A09514-4).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30ug of sample under reducing conditions.
Lane 1: rat testis tissue lysates,
Lane 2: rat C6 whole cell lysates,
Lane 3: mouse NIH/3T3 whole cell lysates,
Lane 4: mouse Neuro-2a whole cell lysates.
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-TRIM16 antigen affinity purified polyclonal antibody (Catalog # A09514-4) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for TRIM16 at approximately 63KD. The expected band size for TRIM16 is at 63KD.

LSPs induce autophagic cell death via the Gal3-Trim16 signaling axis. A) Cell viability of LLC cells after treatment with LSPs (32.77 µg mL−1) for 24 h or different cell death pathway inhibitors. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). B,C) Bio-TEM images (B) and quantification of autophagosomes and autolysosomes (C) of LLC cells treated with LSPs (32.77 µg mL−1), LSPs plus laser ablation, or Autophinib. Red arrows indicate autolysosomes, and blue arrows indicate autophagosomes. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). D) Volcano plot of differentially expressed genes in LLC cells under LSPs versus PBS treatment. E) KEGG pathway enrichment analysis of downregulated signaling pathways after LSPs treatment. F) Heatmap of representative genes related to apoptosis and autophagy in PBS- and LSPs-treated cells. G) Co-IP analysis of Gal3-Trim16 interaction in PBS, LSPs, MSPs, SSPs, and LSP plus laser ablation groups. H,I) Immunofluorescence staining of Gal3 (green), Trim16 (red), and Hoechst (blue) (H) and quantification by Manders’ coefficient of Gal3-Trim16 colocalization efficiency (I) in LLC cells treated with LSPs, LSPs plus laser ablation, GB1107, or Autophinib. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). J,K) Immunofluorescence staining of Trim16 (green), ULK1 (red), and LC3 (cyan) (J) and quantification by Manders’ coefficient of Trim16-ULK1 colocalization, as well as ULK1-LC3 colocalization efficiency (K) in LLC cells treated with LSPs, MSPs, SSPs, LSPs plus laser ablation, GB1107, Trim16-siRNA or Autophinib. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****).
Index in PubMed under a CC BY license. PMID: 41082270

LSPs exhibit potent anticancer efficacy in vivo, which can be terminated by pulsed laser ablation. A) Schematic of the in vivo treatment protocol involving intratumoral injection of LSPs, LSPs plus laser ablation, or Autophinib administration. B) Representative tumor images from mice treated with PBS, LSPs, LSPs plus laser ablation, or Autophinib for 14 days. C,D) Tumor volume curves (C) and individual tumor growth trajectories (D) of mice treated with PBS, LSPs, LSPs plus laser ablation, or Autophinib for 14 days. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). E,F) LC3 immunofluorescence (IF, top) and immunohistochemistry (IHC, bottom) staining of tumor sections from mice treated with PBS, LSPs, LSPs plus laser ablation, or Autophinib for 14 days (E), and corresponding quantification of LC3 integrated density and positive area (F). Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). G) Bio-TEM images and quantification of autophagosomes and autolysosomes of LLC tumor tissues treated with LSPs, LSPs plus laser ablation, or Autophinib. Red arrows indicate autolysosomes, and blue arrows indicate autophagosomes. Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****). H) Quantification by Manders’ coefficient of Gal3-Trim16 and Trim16-LC3 colocalization efficiency in tumor tissue sections treated with LSPs, LSPs plus laser ablation, or Autophinib based on immunofluorescence staining of Gal3 (red), Trim16 (green), and LC3 (purple). Data are presented as mean ± s.d. Statistical significance: p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****).
Index in PubMed under a CC BY license. PMID: 41082270

IHC analysis of TRIM16 using anti-TRIM16 antibody (A09514-4).
TRIM16 was detected in paraffin-embedded section of mouse intestine tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2μg/ml rabbit anti-TRIM16 Antibody (A09514-4) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

IHC analysis of TRIM16 using anti-TRIM16 antibody (A09514-4).
TRIM16 was detected in paraffin-embedded section of rat intestine tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2μg/ml rabbit anti-TRIM16 Antibody (A09514-4) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

IF analysis of TRIM16 using anti-TRIM16 antibody (A09514-4).
TRIM16 was detected in immunocytochemical section of HEPA1-6 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5μg/mL rabbit anti-TRIM16 Antibody (A09514-4) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.