anti cd9 polyclonal primary antibodies  (Bioss)

Journal: Journal of Orthopaedic Translation

Article Title: Functionally improved mesenchymal stem cells via nanosecond pulsed electric fields for better treatment of osteoarthritis

doi: 10.1016/j.jot.2024.03.006

Figure Lengend Snippet: nsPEFs enhanced the release of EVs from MSCs and EVs-nsPEFs enhanced viability, proliferation and migration of chondrocytes and reversed the change of cartilage markers in OA-like chondrocytes. (A) Morphology of EVs and EVs-nsPEFs under TEM. Scale bars, 200 nm. (B) The number of EVs in random field (n = 6) was counted. (C) EVs size distributions and particle concentration analyzed by NTA. (D) Analysis changes of particle concentration between EVs and EVs-nsPEFs. (E) Analysis changes of the protein concentration between EVs and EVs-nsPEFs using BCA method. (F) WB results for the EVs markers CD9, Alix, TSG101 and the negative marker Calnexin. (G) Representative bioluminescence images for Red dye Dil-labeled EVs and EVs-nsPEFs in chondrocytes at 6 h, 12 h and 24 h. Scale bars, 50 μm. (H) CCK8 assay to determine the viability of chondrocytes co-cultured with EVs and EVs-nsPEFs (n = 6). (I) Representative images of EDU staining of chondrocytes co-cultured with EVs and EVs-nsPEFs. Scale bars, 200 μm. (J) Representative images of transwell migration assay of chondrocytes co-cultured with EVs and EVs-nsPEFs. Scale bars, 100 μm. (K) Quantitative analysis of positive cells by EDU staining (n = 4). (L) Quantitative analysis of migrated cells by transwell assay (n = 4). (M) Schematic diagram of experimental design. (N) Representative images of immunofluorescence assay to detect the expression of Collagen II and MMP13 in chondrocytes in control (Group Normal), OA-like chondrocytes (Group IL-1β), OA-like chondrocytes co-cultured with EVs (Group EVs) or EVs-nsPEFs (Group EVs-nsPEFs). Scale bars, 50 μm. Quantitative analysis of immunofluorescence assay to detect the expression of (O) Collagen II and (P) MMP13 in chondrocytes (n = 4). * P < 0.05, ** P < 0.01, **** P < 0.0001. Bar labeled with different lowercase letters indicate a significant difference.

Article Snippet: After blocking with 5% bovine serum albumin (BSA) blocking solution for 1 h at room temperature, membranes were incubated with anti-CD9 primary antibody (1: 1000; Affinity, AF5139, China), ALIX (1: 2000; Proteintech, 12422-1 -AP, USA), TSG101 (1:500; Affinity, DF8427, China) and Calnexin (1:1000; Affinity, AF5362, China) at 4 °C overnight, respectively.

Techniques: Migration, Concentration Assay, Protein Concentration, Marker, Labeling, CCK-8 Assay, Cell Culture, Staining, Transwell Migration Assay, Transwell Assay, Immunofluorescence, Expressing, Control