mouse monoclonal anti aif1 iba1  (Elabscience Biotechnology)

Journal: International Journal of Molecular Sciences

Article Title: Pinealectomy-Induced Neuroinflammation Varies with Age in Rats

doi: 10.3390/ijms26168093

Figure Lengend Snippet: The cellular distribution of pAkt1 in the cornu ammonis area 1 (CA1) of the dHipp was examined in 3-month-old rats that underwent a sham operation. Representative confocal micrographs depict pAkt1 (green) in neuronal and glial cells, with nuclei counterstained with Hoechst (blue). Double labeling with the neuronal marker NeuN (red) reveals intense pAkt1 immunoreactivity in NeuN-positive pyramidal neurons within the CA1 pyramidal cell layer. This produces prominent somatic/perisomatic labeling and extensive pAkt1-NeuN overlap in the merged image ( A1 – A4 ). ( B1 – B4 ) Co-staining with the astrocytic marker glial fibrillary acidic protein (GFAP; red) reveals an absence of the pAkt1 signal in GFAP-positive astrocytes across the stratum radiatum and stratum oriens. Merged images demonstrate no colocalization. ( C1 – C4 ) Co-staining with the microglial marker ionized calcium-binding adapter molecule 1 (Iba1; red) likewise reveals no pAkt1 signal in Iba1-positive microglia, with no co-localization in the merged channels. Scale bar: 20 μm.

Article Snippet: The primary antibodies for immunofluorescence were mouse monoclonal [1B7] antibody to NeuN (1:3000, Antibodies.com, St. Louis, MO, USA, A85405), mouse monoclonal anti-GFAP antibody (1:400, Elabscience, E-AB-70205), mouse monoclonal anti-AIF1 (Iba1) antibody (1:400, Elabscience, E-AB-70353), rabbit polyclonal anti-Phospho-AKT1 (Thr308) (1:200, Affinity Biosciences, Cincinnati, OH, USA, AF0832) and rabbit polyclonal anti-NF kappaB p100/p52 (1:400, Affinity Biosciences, AF6373).

Techniques: Labeling, Marker, Staining, Binding Assay

Journal: International Journal of Molecular Sciences

Article Title: Pinealectomy-Induced Neuroinflammation Varies with Age in Rats

doi: 10.3390/ijms26168093

Figure Lengend Snippet: The cell-type-specific localization of NF-κB in the hippocampal formation of 3-month-old sham-operated rats was revealed by triple-label immunofluorescence. Representative micrographs from the pyramidal cell layer (stratum pyramidale) in CA1 show Hoechst (blue), NF-κB (green), and cell-type markers (red). Merged panels illustrate colocalization ( A1 – A4 ). ( A1 – A4 ) show NF-κB with the neuronal marker NeuN. NF-κB labeling is prominent in CA1 pyramidal neurons and frequently overlaps with NeuN, appearing yellow/orange in the merged image. Perinuclear and intranuclear puncta are evident, consistent with NF-κB distribution across the cytoplasm and nucleus. ( B1 – B4 ) NF-κB with the astrocytic marker GFAP: GFAP-positive processes are abundant around the stratum pyramidale and extend into the stratum radiatum. However, they exhibit only sparse spatial overlap with the NF-κB channel. ( C1 – C4 ) show NF-κB with the microglial marker Iba1. Ramified Iba1-positive profiles are present, but they show minimal colocalization with NF-κB within the stratum pyramidale. The images shown are from 3-month-old rats that underwent a sham operation, and the exposure settings were identical across channels. The same qualitative cellular pattern was observed in other experimental groups. Scale bar = 20 μm.

Article Snippet: The primary antibodies for immunofluorescence were mouse monoclonal [1B7] antibody to NeuN (1:3000, Antibodies.com, St. Louis, MO, USA, A85405), mouse monoclonal anti-GFAP antibody (1:400, Elabscience, E-AB-70205), mouse monoclonal anti-AIF1 (Iba1) antibody (1:400, Elabscience, E-AB-70353), rabbit polyclonal anti-Phospho-AKT1 (Thr308) (1:200, Affinity Biosciences, Cincinnati, OH, USA, AF0832) and rabbit polyclonal anti-NF kappaB p100/p52 (1:400, Affinity Biosciences, AF6373).

Techniques: Immunofluorescence, Marker, Labeling

Journal: International Journal of Molecular Sciences

Article Title: Pinealectomy-Induced Neuroinflammation Varies with Age in Rats

doi: 10.3390/ijms26168093

Figure Lengend Snippet: Representative photomicrographs demonstrating age-related and pinealectomy-induced alterations in microglial cells within the dHipp of male rats. Coronal sections from 3-, 14-, and 18-month-old sham-operated (sham) and pinealectomized (pin) rats were immunostained for Iba1, a marker of microglia. Panels ( A1 – F1 ) illustrate low-magnification overviews, identifying specific hippocampal subregions analyzed in detail: MoDG, GrDG, and PoDG layers of the DG, along with hippocampal subfields CA3c, CA3b, CA3a, and CA1. Panels ( A2 – F6 ) represent higher magnification images, clearly depicting microglial morphology and distribution across the respective hippocampal areas under different experimental conditions. Scale bars = 500 µm ( A1 , B1 , C1 , D1 , E1 , F1 ), 100 μm ( A2 – A6 , B2 – B6 , C2 – C6 , D2 – D6 , E2 – E6 , F2 – F6 ).

Article Snippet: The primary antibodies for immunofluorescence were mouse monoclonal [1B7] antibody to NeuN (1:3000, Antibodies.com, St. Louis, MO, USA, A85405), mouse monoclonal anti-GFAP antibody (1:400, Elabscience, E-AB-70205), mouse monoclonal anti-AIF1 (Iba1) antibody (1:400, Elabscience, E-AB-70353), rabbit polyclonal anti-Phospho-AKT1 (Thr308) (1:200, Affinity Biosciences, Cincinnati, OH, USA, AF0832) and rabbit polyclonal anti-NF kappaB p100/p52 (1:400, Affinity Biosciences, AF6373).

Techniques: Marker

Journal: International Journal of Molecular Sciences

Article Title: Pinealectomy-Induced Neuroinflammation Varies with Age in Rats

doi: 10.3390/ijms26168093

Figure Lengend Snippet: Effect of pinealectomy on Iba1 expression in the dHipp, including the DG, CA3c, CA3b, CA3a and CA1 regions. Transformation index of Iba1 + microglia across hippocampal subfields ( A ) MoDG, ( B ) GrDG, ( C ) CA3c, ( D ) CA3b, ( E ) CA3a, and ( F ) CA1. Bars show mean ± SEM for sham and pinealectomized (pin) rats at 3 months (red), 14 months (blue), and 18 months (green); number of animals (n = 5–6) per group. TI was computed on individually segmented Iba1-positive microglia (single-cell morphometry) as TI = P 2 /(4πA), where p is perimeter and A is cell area (higher values indicate more ramified morphology). MoDG ( A ): * p = 0.021, 3-month-old pin rats vs. matched sham controls. *** p < 0.001, 14-month-old pin rats vs. matched sham controls. GrDG: ( B ) * p = 0.011, 3-month-old pin rats vs. matched sham controls. CA3c ( C ): ** p = 0.003, 3- and 18-month-old pin rats vs. matched sham controls. CA3b ( D ): * p = 0.017, 18-month-old sham vs. 3-month-old sham rats; * p = 0.032, 3-month-old pin rats vs. matched sham controls; * p = 0.016, 18-month-old pin rats vs. matched sham rats. CA3a ( D ): ** p = 0.006, 3-month-old pin rats vs. matched sham control; ** p = 0.003, 14-momth-old pin rats vs. matched controls. CA1 ( E ) * p = 0.0108, 3-month-old pin rats vs. matched sham control; *** p < 0.001, 14-momth-old pin rats vs. matched controls.

Article Snippet: The primary antibodies for immunofluorescence were mouse monoclonal [1B7] antibody to NeuN (1:3000, Antibodies.com, St. Louis, MO, USA, A85405), mouse monoclonal anti-GFAP antibody (1:400, Elabscience, E-AB-70205), mouse monoclonal anti-AIF1 (Iba1) antibody (1:400, Elabscience, E-AB-70353), rabbit polyclonal anti-Phospho-AKT1 (Thr308) (1:200, Affinity Biosciences, Cincinnati, OH, USA, AF0832) and rabbit polyclonal anti-NF kappaB p100/p52 (1:400, Affinity Biosciences, AF6373).

Techniques: Expressing, Transformation Assay, Control

Journal: bioRxiv

Article Title: Transglutaminase 2 function in glioblastoma tumor efferocytosis

doi: 10.1101/2024.08.29.610293

Figure Lengend Snippet: A. THP-1 cells growing on coverslips were differentiated into macrophages and exposed to apoptotic cells that were labelled with pHrodo. Cells were fixed at the indicated times and analyzed for pHrodo fluorescence by microscopy. Numbers of positive cells were quantitated using Fiji software. Data shown are the mean and standard deviation from three biological replicates. For each biological replicate, 3-4 random fields of view were analyzed. * indicates P < 0.05 by a two-sided t test in comparisons of the untreated 1 h time point data with either the ONO-7475 or the NC9 1 h time point data. B. Efferocytosis in human glioblastoma organoids. Nuclei are labelled with DAPI (blue); macrophages/microglia are detected with Iba1 immunofluorescence (green); apoptotic cells are detected by fluorescent TUNEL assay (red). The small white square indicates the site of the cell shown in close up in the bottom images. Bottom images show a macrophage/microglial cell efferocytosing multiple apoptotic cells. Left panel shows DAPI only; middle sample shows DAPI and TUNEL; right panel shows DAPI, TUNEL and Iba1. Red arrows indicate examples of apoptotic cells with nuclear condensation. C. Quantitation of apoptotic cells in organoids that were either untreated (control) or treated with 5 nM ONO-7475 for 48 h or 10 µM NC9 for 24 h. Apoptotic cells were counted using image analysis and were normalized to organoid surface area, as described in Materials and Methods. Circles show apoptotic cells counts from two or three sections from one organoid per condition. Mean and standard deviation are shown in the adjacent closed circle with error bars. * indicates a P value < 0.05 using a one-tailed Student’s t-test. n.s. not significant. D. Organoids from a second patient were isolated using either 5% O2 or 20% O2 conditions. Once established, organoids were either untreated or treated with 10 µM NC9 for 48 h. Quantitation of apoptotic cells was performed as in C. Each open circle shows the apoptotic cell count from one organoid. Mean and standard deviation are shown in the adjacent closed circle with error bars. * indicates a P value < 0.05 using a two-tailed Student’s t-test. n.s. not significant

Article Snippet: Anti-Iba1/AIF1 mouse monoclonal antibody (used for immunofluorescence in xenografts) and anti-CD68 mouse monoclonal antibody (used for immunofluorescence in human tissue samples) were from Millipore/Sigma (cat. #s MABN92 and AMAB90873, respectively).

Techniques: Fluorescence, Microscopy, Software, Standard Deviation, Immunofluorescence, TUNEL Assay, Quantitation Assay, Control, One-tailed Test, Isolation, Cell Counting, Two Tailed Test

Journal: Autophagy

Article Title: Activation of PPARA-mediated autophagy reduces Alzheimer disease-like pathology and cognitive decline in a murine model

doi: 10.1080/15548627.2019.1596488

Figure Lengend Snippet: Primary antibodies used in this study.

Article Snippet: Primary antibodies Source Catalog no. Western blot Immunofluorescence Immunohistochemistry Rabbit monoclonal anti-MAP1LC3B/LC3B Cell Signaling Technology 3868 1:1000 - - Rabbit monoclonal anti-MAP1LC3B/LC3B Abcam ab64781 - 1:500 - Mouse anti-β-amyloid,17–24(4G8) BioLegend 800701 - 1:500 1:500 Rabbit polyclonal anti-SQSTM1 Elabscience EAP3350 1:1000 - - Rabbit polyclonal anti-LAMP1 Abcam ab24170 1:1000 1:300 - Rabbit monoclonal anti-BECN1 Cell Signaling Technology 3495 1:1000 - - Rabbit monoclonal FKBP5 Cell Signaling Technology 12210 1:1000 - - Mouse monoclonal anti-DLG4 Cell Signaling Technology 36233 1:1000 - - Rabbit polyclonal anti-DLG4 Abcam ab18258 - - 1:200 Rabbit monoclonal anti-GFAP Cell Signaling Technology 12389 1:1000 1:300 - Mouse monoclonal anti-GFAP Millipore MAB360 - 1:300 - Rabbit monoclonal anti-AIF1 Abcam ab178680 1:1000 1:300 - Mouse monoclonal anti-AIF1 Millipore MABN92 - 1:300 - Rabbit polyclonal anti-TFEB Elabscience EAP2314 1:1000 - - Rabbit polyclonal anti-PPARA Elabscience ESAP13084 1:500 - - Mouse monoclonal anti-SYP Millipore MAB5258-50UG 1:10000 - - Mouse monoclonal anti-CD68 Abcam ab201973 1:1000 - - Rabbit monoclonal anti-β-amyloid/Aβ/total Aβ Cell Signaling Technology 8243 1:1000 - - Rabbit monoclonal anti-β-amyloid (1-42 Specific)/Aβ42 Cell Signaling Technology 14974 1:1000 - - Rabbit anti-β-amyloid (1-40 Specific)/Aβ40 Cell Signaling Technology 12990 1:1000 - - Mouse monoclonal anti-GAPDH Proteintech 60004–1-Ig 1:10000 - - Mouse monoclonal anti-ACTB Beijing Zhong Shan-Golden Bridge Biological Technology CO., LTD TA-09 1:10000 - - Open in a separate window Primary antibodies used in this study.

Techniques: Western Blot, Immunofluorescence, Immunohistochemistry