mouse anti probdnf  (Santa Cruz Biotechnology)

Journal: Cell Death and Differentiation

Article Title: Localized release of muscle-generated BDNF regulates the initial formation of postsynaptic apparatus at neuromuscular synapses

doi: 10.1038/s41418-024-01404-4

Figure Lengend Snippet: Representative images ( a ) and quantification ( b ) showing the spatial localization of endogenous BDNF, proBDNF, TrkB, and pTrkB receptors at aneural AChR clusters in cultured Xenopus muscles. Arrows and arrowheads indicate localized signals at the perforated and AChR-rich regions of aneural clusters, respectively. c Representative images showing the spatial association (arrows) between BDNF and PLS at perforated aneural AChR clusters. The yellow lines indicate the region-of-interest for generating line profiles. Line profile plots showing the relative fluorescence intensities of AChR (red), BDNF (green), and PLS core and cortex markers (magenta). The colored boxes indicate the spatial enrichment of markers with their respective fluorescence signals above the cutoff intensity of 50%. d Maximal projection of Airyscan confocal z-stack images showing the spatial enrichment of BDNF (top panels) or proBDNF (bottom panels) at PLS core regions within aneural AChR clusters. The orthogonal xz views show the differential distributions of AChR, cortactin, and BDNF/proBDNF. Scale bars represent 5 µm, unless stated otherwise.

Article Snippet: The primary antibodies used in this study include BDNF antibody (1:1000; Alomone Labs, ANT-010; RRID: AB_2039756), proBDNF antibody (1:1500; Alomone Labs, ANT-006; RRID: AB_2039758), TrkB antibody (1:200; Santa Cruz Biotechnology, sc-119; RRID: AB_632559), phospho-TrkB (Tyr705) antibody (1:200; Thermo Fisher Scientific, PA5-38077; RRID: AB_2554680), cortactin antibody (1:1200; EMD Millipore, 05-180; RRID: AB_309647), and vinculin antibody (1:100; DSHB, VN 3-24; RRID: AB_2214518).

Techniques: Cell Culture, Muscles, Fluorescence

Journal: Cell Death and Differentiation

Article Title: Localized release of muscle-generated BDNF regulates the initial formation of postsynaptic apparatus at neuromuscular synapses

doi: 10.1038/s41418-024-01404-4

Figure Lengend Snippet: Western blot analysis ( a ) and quantification ( b ) showing the effective knockdown of both endogenous BDNF and proBDNF expression by antisense BDNF MO. Anti-α-tubulin was used as a loading control. c–f Representative images ( c ) showing the reduced intensity of aneural AChR clusters, BDNF, and proBDNF signals in BDNF MO muscle cells. Insets: Fluorescent dextran signals indicate the presence of the MO in muscle cells. Quantification showing the effects of BDNF knockdown on the formation of aneural AChR clusters ( d ) and their normalized fluorescence intensity ( e ), as well as the normalized intensity of BDNF and proBDNF ( f ) at the perforated regions of aneural AChR clusters. g – i Representative images ( g ) showing the effects of BB-94 or furin inhibitor on the localization of BDNF and proBDNF at perforated aneural AChR clusters. Quantification showing the effects of BB-94 or furin inhibitor on the normalized intensity of BDNF ( h ) and proBDNF ( i ) at the perforated regions of aneural AChR clusters. Scale bars represent 5 μm. 8-bit pseudo-color images highlight the relative fluorescence intensity. Data are means ± SEM. The numbers indicated in the bar regions represent the total numbers of blots ( b ) or muscle cells ( d – f , h , i ) measured from three independent experiments. *, **, ***, **** represent p ≤ 0.05, 0.01, 0.001, and 0.0001, respectively (Student’s t test ( b , d – f ) or one-way ANOVA with Tukey’s multiple comparisons test ( h , i )).

Article Snippet: The primary antibodies used in this study include BDNF antibody (1:1000; Alomone Labs, ANT-010; RRID: AB_2039756), proBDNF antibody (1:1500; Alomone Labs, ANT-006; RRID: AB_2039758), TrkB antibody (1:200; Santa Cruz Biotechnology, sc-119; RRID: AB_632559), phospho-TrkB (Tyr705) antibody (1:200; Thermo Fisher Scientific, PA5-38077; RRID: AB_2554680), cortactin antibody (1:1200; EMD Millipore, 05-180; RRID: AB_309647), and vinculin antibody (1:100; DSHB, VN 3-24; RRID: AB_2214518).

Techniques: Western Blot, Knockdown, Expressing, Control, Fluorescence

Journal: Cell Death and Differentiation

Article Title: Localized release of muscle-generated BDNF regulates the initial formation of postsynaptic apparatus at neuromuscular synapses

doi: 10.1038/s41418-024-01404-4

Figure Lengend Snippet: a In nerve-muscle cocultures, we speculate that PLSs direct the vesicular trafficking of not only MT1-MMP, but also BDNF and its precursor proBDNF for localized release at AChR clusters. The intracellular endoprotease furin mediates the proteolytic conversion of proBDNF to mBDNF for activating TrkB receptors in muscles. Knockdown of muscle BDNF expression affects aneural AChR cluster formation; therefore, less AChR molecules are redistributed and recruited for the assembly of nerve-induced synaptic AChR clusters. b At developing NMJs in vivo, MBKO mouse embryos exhibit reduced AChR prepatterning in the diaphragm muscles and reduced axonal branching and arborization in the phrenic nerves at E13.5. The defects in AChR prepatterning reduce the number of AChR molecules being redistributed to the postsynaptic apparatus, as reflected by the reduced intensity of synaptic AChR clusters in MBKO mice, at E18.5.

Article Snippet: The primary antibodies used in this study include BDNF antibody (1:1000; Alomone Labs, ANT-010; RRID: AB_2039756), proBDNF antibody (1:1500; Alomone Labs, ANT-006; RRID: AB_2039758), TrkB antibody (1:200; Santa Cruz Biotechnology, sc-119; RRID: AB_632559), phospho-TrkB (Tyr705) antibody (1:200; Thermo Fisher Scientific, PA5-38077; RRID: AB_2554680), cortactin antibody (1:1200; EMD Millipore, 05-180; RRID: AB_309647), and vinculin antibody (1:100; DSHB, VN 3-24; RRID: AB_2214518).

Techniques: Muscles, Knockdown, Expressing, In Vivo

Journal: Brain Sciences

Article Title: Ouabain Counteracts Retinal Ganglion Cell Death Through Modulation of BDNF and IL-1 Signaling Pathways

doi: 10.3390/brainsci15020123

Figure Lengend Snippet: Primary and secondary antibodies for Western blotting.

Article Snippet: Mouse anti-proBDNF , Santa Cruz Biotechnology (Dallas, TX, USA) , 1:1000.

Techniques: Western Blot