mouse anti pro spc  (Santa Cruz Biotechnology)

Journal: bioRxiv

Article Title: Type II alveolar cells with constitutive expression of MHCII and limited antigen presentation capacity contribute to improved respiratory viral disease outcomes

doi: 10.1101/2021.03.18.435984

Figure Lengend Snippet: a,b, General gating strategy for cell sorting of murine AT2s using MHCII as a positive marker ( a ) or without using MHCII as a positive marker ( b ). The right column contour plots are the same as the final pseudocolor plots in the corresponding gating strategies, and they are shown to highlight the final AT2 population defined from each strategy. c, Comparison of the two populations of AT2s identified by the two gating strategies in terms of MHCII expression (contour plot, left) or pro-SPC expression (histogram, right).

Article Snippet: Tissue was then processed, paraffin-embedded, sectioned, and stained with DAPI, and the following antibodies: anti-mouse pro-SPC (AB3786, EMD Millipore), anti-mouse MHCII (107601, Biolegend), and anti-mouse E-cadherin (ab76319, Abcam).

Techniques: FACS, Marker, Expressing

Journal: bioRxiv

Article Title: Type II alveolar cells with constitutive expression of MHCII and limited antigen presentation capacity contribute to improved respiratory viral disease outcomes

doi: 10.1101/2021.03.18.435984

Figure Lengend Snippet: a , MHCII expression by AT2s from naïve SPC-Cre-ERT2 +/- (SPC Cre ), H2-Ab1 fl/fl (Ab1 fl/fl ), and SPC-Cre-ERT2 +/- × H2-Ab1 fl/fl (SPC ΔAb1 ), lungs measured by flow cytometry; histograms represent n>50 mice per strain total from >10 independent experiments. b , Immunofluorescence detection of MHCII, SPC, and E-Cadherin in Ab1 fl/fl , SPC ΔAb1 , and MHCII -/- lungs; scale bar depicts 20 μm, representative images of n=1-2 mice per strain. c,d, Percent and MFI of each cell type expressing MHCII protein for lung AT2s, B cells, alveolar macrophages, CD103 + DCs, and spleen B cells and CD8 + DCs, from naïve SPC Cre , Ab1 fl/fl , and SPC ΔAb1 mice, measured by ex vivo flow cytometry analysis; bars show n=4 mice per strain from 1 experiment, representative of 2 similar independent experiments for the Ab1 fl/fl and SPC ΔAb1 strains (n=8 mice total per strain). e, H&E staining of naïve 12 week old Ab1 fl/fl and SPC ΔAb1 lungs, images representative of n=5 mice per strain. Scale bar depicts 200 μm. f-i, Absolute number of lung CD4 + and CD8 + T cells ( f ), and frequency of lung CD4 + and CD8 + T cells expressing CD44, CD62L ( g ), Foxp3 ( h ), and CD69, CD11a ( i ), as indicated, from naïve 12 week old Ab1 fl/fl and SPC ΔAb1 mice measured by ex vivo flow cytometry analysis; n=9-10 mice per strain from 1 experiment, representative of 2 independent experiments. c-d, f-i , Data shown are mean plus SEM, analyzed by Mann-Whitney test ( f [CD4s]) or unpaired two-tailed Student’s t -test ( f [CD8s], g-i ). Full statistical test results are in .

Article Snippet: Tissue was then processed, paraffin-embedded, sectioned, and stained with DAPI, and the following antibodies: anti-mouse pro-SPC (AB3786, EMD Millipore), anti-mouse MHCII (107601, Biolegend), and anti-mouse E-cadherin (ab76319, Abcam).

Techniques: Expressing, Flow Cytometry, Immunofluorescence, Ex Vivo, Staining, MANN-WHITNEY, Two Tailed Test

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Muc5b Enhances Murine Honeycomb-like Cyst Formation

doi: 10.1165/rcmb.2019-0138LE

Figure Lengend Snippet: Wild-type mice develop Muc5b-filled honeycomb cysts (HCs) in response to lung injury. Antibodies used for immunohistochemistry included goat anti-mouse Muc5b (11765, 1:1000; Everest), rabbit anti-mouse pro-SPC (ab90716, 1:100; Abcam), and chicken anti-mouse Krt5 (Poly9059, 1:1000; BioLegend). Appropriate fluorescent secondary antibodies were obtained from Jackson ImmunoResearch, reconstituted at 1 mg/ml, and used at 1:200. (A) Muc5b protein is increased in response to productive influenza infection (28 d postinfection) or repeated bleomycin treatment (*P < 0.05 and ***P < 0.005). (B) Influenza A H1N1 (H1N1) infection does not lead to significant fibrosis compared with UV-irradiated control virus as measured by hydroxyproline, whereas repeated bleomycin injury leads to significant increases in hydroxyproline compared with saline-treated control (***P < 0.005). (C) H1N1 generates considerable airspace remodeling resembling honeycombing (Krt5 stained DAB) 75 days after infection (top left). Similar to what is observed in humans, H1N1 HCs are characterized by Krt5-basal cells surrounding dense mucociliary structures filled with Muc5b+ colloid and generally exclude type II pneumocytes (top right). HCs also develop in mice subjected to repeated bleomycin injury (bottom left). Repeat bleomycin injury–derived HCs are generally indistinguishable from H1N1-derived HCs (bottom right). Honeycomb structures are highlighted by red (DAB panels) and white (IF panels) asterisks. (D) Cysts in mice subjected to single-dose bleomycin persist in the absence of significant fibrosis. (E) In general, resolution of H1N1 injury involves more robust honeycombing than repeat or single-dose bleomycin.

Article Snippet: Antibodies used for immunohistochemistry included goat anti-mouse Muc5b (11765, 1:1000; Everest), rabbit anti-mouse pro-SPC (ab90716, 1:100; Abcam), and chicken anti-mouse Krt5 (Poly9059, 1:1000; BioLegend).

Techniques: Immunohistochemistry, Infection, Irradiation, Staining, Derivative Assay

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Muc5b Enhances Murine Honeycomb-like Cyst Formation

doi: 10.1165/rcmb.2019-0138LE

Figure Lengend Snippet: Durable murine honeycombing varies with expression of Muc5b in the bleomycin, but not H1N1, models. HC were morphometrically quantified via unbiased stereology; initially a 350 micron grid was used to quantify the volume fraction of the cysts. Cystic areas were then randomly sampled using a 100 micron grid to determine cyst size. Lung volume was calculated using the Cavalieri method. (A) No significant differences were observed in cyst volume or number based on genotype among mice infected with H1N1. (B) Bleomycin-injured mice expressing Muc5b under control of the SPC promoter had a significant increase in cyst volume (P < 0.05). The overall number of cysts is independent of genotype (*P < 0.05, **P < 0.01, and ***P < 0.005). KO = knockout; WT = wild-type (see text for additional strain details).

Article Snippet: Antibodies used for immunohistochemistry included goat anti-mouse Muc5b (11765, 1:1000; Everest), rabbit anti-mouse pro-SPC (ab90716, 1:100; Abcam), and chicken anti-mouse Krt5 (Poly9059, 1:1000; BioLegend).

Techniques: Expressing, Infection, Knock-Out