anti lamp1 antibody  (Bioss)

Journal: EMBO Molecular Medicine

Article Title: MHC-I upregulation by macbecin II in the solid tumors potentiates the effect of active immunotherapy

doi: 10.1038/s44321-025-00213-7

Figure Lengend Snippet: ( A ) Flow cytometry analysis for INF-γ-CD8 + T cells in the four groups. ( B ) Granzyme B expression was examined by immunohistochemistry (left) and quantified (right) in the four groups. The staining intensity was analyzed by the unpaired two-tailed Student’s t-test ( n = 4/group, biological replicates). ( C ) LAMP1 expression was examined through immunohistochemistry (left) and quantified (right) in the four groups. The staining intensity was measured and analyzed by the one-way ANOVA with a Tukey post-hoc test ( n = 4/group, biological replicates). ( D , E ) MHC-I expression was examined in EPCAM+ tumor cells and CD11c+ dendritic cells in the dissociated tumor by FACS. Statistical analysis was performed using the one-way ANOVA with a Tukey post-hoc test ( n = 3/group, biological replicates). ( F , G ) Animal weight ( F ) and AST activity ( G ) in the blood of mice were measured at the end-point. Data are represented as mean +/− SEM ( n = 5/group, biological replicates). ( H ) B16-Ova cells (500 cells/well, n = 3/group, biological replicates, were seeded in a 96-well plate and treated with a combination of macbecin II (0.1 µM) and OT-1 T cells (E:T ratio 5:1) with or without ectopic MHC-I expression for 48 h. The T cells were activated with IL2-ep13nsEV that were isolated from B16-Ova lysate-pulsed BMDCs (E:T ratio 5:1). After the incubation, cells were washed with ice-cold PBS to remove dead cells. The live cells were fixed with methanol at room temperature for 15 min and stained with crystal violet. The dye was dissolved in 10% acetic acid and measured at 590 nm. ( I ) E0771 cells (500 cells/well, n = 3/group, biological replicates) were seeded in a 96-well plate and treated with a combination of T cells (E:T ratio 5:1) and macbecin II (0.1 µM) with or without ectopic MHC-I expression for 48 h. The T cells were isolated from the spleen of E0771-bearing syngeneic mouse and they were activated with IL2-ep13nsEV that were isolated from E0771 lysate-pulsed BMDCs. After the incubation, cells were processed as described in ( H ). Statistical significance was determined by the one-way ANOVA with a Tukey post-hoc test. ( J ) MHC-I was ectopically expressed in B16-F1 cells, and the MHC-I expression was confirmed by FACS. The result was analyzed by the unpaired two-tailed Student’s t-test ( n = 3/group, biological replicates). ( K ) Cell viability was examined by MTS assay for B16-F1 with or without MHC-I over expression (OE) ( n = 3/group, biological replicates). ( L , M ) Animal weight ( L ) and AST activity ( M ) were measured at the end-point ( n = 5/group, biological replicates). Data are presented as mean +/− SEM.

Article Snippet: LAMP1 antibody , Bioss , BS-1970R.

Techniques: Flow Cytometry, Expressing, Immunohistochemistry, Staining, Two Tailed Test, Activity Assay, Isolation, Incubation, MTS Assay, Over Expression

Journal: EMBO Molecular Medicine

Article Title: MHC-I upregulation by macbecin II in the solid tumors potentiates the effect of active immunotherapy

doi: 10.1038/s44321-025-00213-7

Figure Lengend Snippet: Reagents and tools table

Article Snippet: LAMP1 antibody , Bioss , BS-1970R.

Techniques: Recombinant, Over Expression, Plasmid Preparation, Sequencing, Proliferation Assay, Red Blood Cell Lysis, Staining, Activation Assay, Isolation, Cell Isolation, cDNA Synthesis, Bradford Protein Assay, AST Assay, Enzyme-linked Immunosorbent Assay, Software

Journal: EMBO Molecular Medicine

Article Title: MHC-I upregulation by macbecin II in the solid tumors potentiates the effect of active immunotherapy

doi: 10.1038/s44321-025-00213-7

Figure Lengend Snippet: Reagents and tools table

Article Snippet: LAMP1 antibody , Bioss , BS-1970R.

Techniques: Recombinant, Over Expression, Plasmid Preparation, Sequencing, Proliferation Assay, Red Blood Cell Lysis, Staining, Activation Assay, Isolation, Cell Isolation, cDNA Synthesis, Bradford Protein Assay, AST Assay, Enzyme-linked Immunosorbent Assay, Software

Journal: Journal of Neuroinflammation

Article Title: Glutamine metabolism modulates microglial NLRP3 inflammasome activity through mitophagy in Alzheimer’s disease

doi: 10.1186/s12974-024-03254-w

Figure Lengend Snippet: Reduced glutamine metabolism enhances microglial mitophagy and inhibits NLRP3 inflammasome activation. ( A ) Western blots of pro- and mature IL-1β and caspase-1 in lysates and supernatants of LoAMs in the presence or absence of the anti-oxidant and free radical scavenger NAC ( n = 3). ND, not detected. ( B ) Western blots and densitometry quantification of the mitophagy protein PINK1, parkin, and p62 in LoAMs in the presence or absence of BPTES ( n = 3). ( C ) Autophagosome formation indicated by puncta and quantitated by confocal microscopy in LoAMs in the presence or absence of BPTES or GLS1 siRNA. Scale bar: 10 μm ( n = 6). ( D ) Confocal microscopy detection and quantitation of co-localization of the mitochondrial protein TOM20 (green), lysosomal protein LAMP1 (red), and DAPI (blue) in LoAMs in the presence or absence of BPTES or GLS1 siRNA. Scale bar: 10 μm ( n = 6). ( E ) Confocal microscopy detection and quantitation of co-staining of MitoTracker ang LysoTracker in LoAMs in the presence or absence of BPTES or GLS1 siRNA. Scale bar: 10 μm ( n = 6). ( F ) WB analysis of pro- and mature IL-1β and caspase-1 in lysates and supernatants of LoAMs in the presence or absence of mitophagy stimulator NMN ( n = 3). n represents the number of biological replicates. Data are presented as the mean ± SEM. Statistical significance was determined by one-way ANOVA. * p < 0.05; ** p < 0.01; *** p < 0.001

Article Snippet: After fixed with 4% PFA, permeabilized using 0.5% triton X-100 and blocked in 5% BSA, the coverslips were then stained with mouse monoclonal anti-TOM20 (1:200; 66777-1-Ig, Proteintech) and rabbit polyclonal anti-LAMP1 (1:100; bs-1970R, Bioss) or goat polyclonal anti-ASC (1:50; ab175449, Abcam) and rabbit polyclonal anti-NLRP3 (1:500; 19771-1-AP, Proteintech) overnight at 4℃.

Techniques: Activation Assay, Western Blot, Confocal Microscopy, Quantitation Assay, Staining

Journal: Journal of Neuroinflammation

Article Title: Glutamine metabolism modulates microglial NLRP3 inflammasome activity through mitophagy in Alzheimer’s disease

doi: 10.1186/s12974-024-03254-w

Figure Lengend Snippet: Reduced glutamine metabolism enhances mitophagy in LPS-primed oAβ 1−42 -treated primary microglia via AMPK/mTORC1 signaling. (A and B) WB analysis and densitometry-based quantification of raptor and p-AMPK in LoAMs in the presence or absence of BPTES ( A ) and GLS1 siRNA ( B ) ( n = 3). ( C ) Autophagosome formation indicated by puncta and quantitated by confocal microscopy in LoAMs with or without rapamycin. Scale bar: 10 μm ( n = 6). ( D ) Confocal microscopy detection and quantitation of co-localization of the mitochondrial protein TOM20 (green), lysosomal protein LAMP1 (red), and DAPI (blue) in LoAMs in the presence or absence of rapamycin. Scale bar: 10 μm ( n = 6). ( E ) Confocal microscopy detection and quantitation of co-staining of MitoTracker ang LysoTracker in LoAMs in the presence or absence of rapamycin. Scale bar: 10 μm ( n = 6). ( F ) Western blot analysis of pro- and mature IL-1β and caspase-1 in lysates and supernatants of LoAMs in the presence or absence of rapamycin ( n = 3). ND, not detected. n represents the number of biological replicates. Data are presented as the mean ± SEM. Statistical significance was determined by one-way ANOVA. * p < 0.05; ** p < 0.01; *** p < 0.001; NS, not significant

Article Snippet: After fixed with 4% PFA, permeabilized using 0.5% triton X-100 and blocked in 5% BSA, the coverslips were then stained with mouse monoclonal anti-TOM20 (1:200; 66777-1-Ig, Proteintech) and rabbit polyclonal anti-LAMP1 (1:100; bs-1970R, Bioss) or goat polyclonal anti-ASC (1:50; ab175449, Abcam) and rabbit polyclonal anti-NLRP3 (1:500; 19771-1-AP, Proteintech) overnight at 4℃.

Techniques: Confocal Microscopy, Quantitation Assay, Staining, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Transcranial Magneto-Acoustic Stimulation Protects Synaptic Rehabilitation from Amyloid-Beta Plaques via Regulation of Microglial Functions

doi: 10.3390/ijms25094651

Figure Lengend Snippet: TUS and TMAS treatments attenuated amyloid burdens and amyloid-induced neurotoxicity. ( A ) Coronal sections of sham-, TUS-, or TMAS-treated 5xFAD mice were stained with DAPI (blue) for nuclei, anti-Aβ (red) for Aβ plaques, and Iba1 (green) for microglia. Representative images of the hippocampal regions are shown. Original magnification ×20; scale bar: 100 μm. ( B ) Statistical analysis of the difference in the Aβ fluorescence areas between sham, TUS, and TMAS groups (6 mice from each group were used for analysis). ( C ) Quantitation of the number of plaque-associated microglia in A (6 mice from each group were used for analysis). ( D ) Quantitation of the area of Lamp1-positive dystrophic neurites in E (6 mice from each group were used for analysis). ( E ) Coronal sections from sham-, TUS-, or TMAS-treated 5xFAD mice were stained with DAPI (blue) for nuclei, anti-Aβ (red) for Aβ, and Lamp1 (green) for dystrophic neurites. Representative images of the hippocampus regions are shown. Original magnification ×40; scale bar: 50 μm. Zoom-in images on the right have a scale bar equal to 20 μm. All data in the results are expressed as mean ± SEM. * p < 0.05; *** p < 0.001.

Article Snippet: The primary antibodies used were as follows: anti-Iba1 (Wako; Cat# 016-20001, RRID: AB_839506, 1:500, Osaka, Japan), anti-Iba1 (Santa Cruz Biotechnology; Cat# sc-32725, RRID: AB_667733, 1:500, Santa Cruz, CA, USA), anti-Aβ (Abcam; Cat# ab32136, RRID: AB_2289606, 1/500, Cambridge, UK), anti-PSD95 (Cell Signaling Technology; Cat# 3450S, RRID: AB_2292883, 1:2000, Danvers, MA, USA), anti-CD11c (Biolegend; Cat# 117304, RRID: AB_313773, 1:1000, Santa Cruz, CA, USA), anti-Lamp1 (BIOSS; Cat# bs-1970R, 1:100, Beijing, China), and anti-CD68 (Bio-Rad; Cat# MCA1957, RRID: AB_322219, 1:50, Heraklion, CA, USA).

Techniques: Staining, Fluorescence, Quantitation Assay