Journal: Scientific Reports

Article Title: Lrig1 -expressing epidermal progenitors require SCD1 to maintain the dermal papilla niche

doi: 10.1038/s41598-023-30411-7

Figure Lengend Snippet: Phenotypic changes following Scd1 deletion in Lrig1 -expressing epidermal hair follicle cells. ( a ) RNA in-situ hybridisation of Lrig1 on P19 WT dorsal skin. Polr2A (housekeeping gene) was used as positive control and DapB (Bacterial gene) as negative control. ( b ) RNA in-situ hybridisation of Scd1 , Lrig1 and Duplex Lrig1 and Scd1 on P19 WT Telogen hair follicle. ( c ) RNA in-situ hybridisation of Scd1 , Lrig1 and Duplex in P45 Control Lrig1 + / + ; Scd1 fl/fl anagen bulb. ( d ) RNA in-situ hybridisation of Scd1 , Lrig1 and Duplex in P45 Mutant Lrig1-Cre ERT2/ + ; Scd1 fl/fl anagen bulb. Purple dotted line denotes boundary of DP cells. Black arrowhead denotes loss of Scd1 transcript expression (scale bar 20 μm). ( e ) Representative image of shaved Lrig1 Control mouse whole body hair cycle progression (n = 10 mice), post tamoxifen injection at P19. ( f ) H&E stained sections of Lrig1 Control skin at P21, P40, P48 and P52. ( g ) Representative image of shaved Lrig1 Mutant mouse whole body hair cycle progression (n = 13 mice), post tamoxifen injection at P19. ( h ) H&E stained sections of Lrig1 Mutant skin at P21, P40, P48 and P52. Blue arrowhead indicates presence of cellular infiltrates. Black arrowhead indicates exposed club-like hair shaft and missing DP (scale bar 50 μm).

Article Snippet: The following antibodies were used: Rat monoclonal anti-Ki67 (catalogue no. 14-5698-82, eBioscience), Rabbit polyclonal anti-mouse Scd1(M38) (catalogue no. #2438, Cell signalling), and Goat anti-Rabbit conjugated to Alexa Fluor A568 (catalogue no. A11036, Life Technologies).

Techniques: Expressing, In Situ, Hybridization, Positive Control, Negative Control, Mutagenesis, Injection, Staining

Journal: Scientific Reports

Article Title: Lrig1 -expressing epidermal progenitors require SCD1 to maintain the dermal papilla niche

doi: 10.1038/s41598-023-30411-7

Figure Lengend Snippet: Altered expression of dermal papilla and hair matrix markers following Scd1 deletion. ( a – c ) Representative images of alkaline phosphatase (AP) stain of Lrig1 Control and Mutant hair follicles at P42, P45 and P48. Red arrowhead indicates the presence of AP +ve DP in anagen and catagen hair follicles. ( d ) Quantification of AP +ve DP in Lrig1 Control and Mutant hair follicles at P42, P45, P48 and P52 (n = 2 mice, 100 hair follicles per mouse) Student's t-test applied, *p < 0.05 and **p < 0.001. ( e , f ) RNA in-situ hybridisation of Igfbp3 in DP cells at P48 and P52. Black arrowhead indicates the presence of Igfbp3 +ve DP in hair follicles. ( g , h ) RNA in-situ hybridisation of APCDD1 in DP cells at P48 and P52. Black arrowhead indicates the presence of APCDD1 +ve DP in hair follicles. ( i – k ) Ki67 antibody stained sections of Lrig1 control and Mutant skins at P45, P48 and P52. White arrowhead indicates Ki67 +ve matrix. ( l ) Percentage of Ki67 +ve cells in Lrig1 Control and Mutant matrix progenitors at P45, P48 and P52 (n = 3 mice, 12 hair follicles analysed per mouse). Student’s t-test applied, *p < 0.05 and **p < 0.001. Yellow opaque line denotes analysed area of expression. ( m – o ) RNA in-situ hybridisation of Msx2 in matrix cells, IRS and pre-cortex of Lrig1 Control and Mutant skins at P45, P48 and P52. Black arrowhead indicates Msx2 +ve matrix cells (scale bar 50 μm).

Article Snippet: The following antibodies were used: Rat monoclonal anti-Ki67 (catalogue no. 14-5698-82, eBioscience), Rabbit polyclonal anti-mouse Scd1(M38) (catalogue no. #2438, Cell signalling), and Goat anti-Rabbit conjugated to Alexa Fluor A568 (catalogue no. A11036, Life Technologies).

Techniques: Expressing, Staining, Mutagenesis, In Situ, Hybridization

Journal: Scientific Reports

Article Title: Lrig1 -expressing epidermal progenitors require SCD1 to maintain the dermal papilla niche

doi: 10.1038/s41598-023-30411-7

Figure Lengend Snippet: Scd1 -deletion reduced expression of genes in key hair follicle signalling pathways, Wnt and Hedgehog. ( a ) RNA in-situ hybridisation of Scd1 in Lrig1 Control and Mutant hair bulb at P42. Purple arrowhead indicates loss of Scd1 transcript expression. ( b ) Duplex RNA in-situ hybridisation of Scd1 and Axin2 in Lrig1 Control and Mutant hair bulb at P42. Purple arrowhead indicates loss of Axin2 transcript expression. ( c ) Duplex RNA in-situ hybridisation of Scd1 and Wls in Lrig1 Control and Mutant hair bulb at P42. Purple arrowhead indicates loss of Wls transcript expression. ( d ) Scd1 , Axin2 , Wls , PPIB and DapB transcripts by spot detection in P42 Lrig1 Control and Mutant mice (n = 3 animals, 12 hair follicles analysed per animal). *p < 0.05 and **p < 0.001. Yellow solid line indicates area of representative image analysed for transcripts. ( e ) Duplex RNA in-situ hybridisation of Scd1 and Shh in Lrig1 Control and Mutant hair bulb at P42. Purple arrowhead indicates loss of Shh transcript expression. ( f ) Shh transcripts by spot detection in adjacent matrix of P42 Lrig1 Control and Mutant mice (n = 3 animals, 12 hair follicles analysed per animal). *p < 0.05 and **p < 0.001. Yellow solid line indicates area of representative image analysed for transcripts. ( g ) Duplex RNA in-situ hybridisation of Scd1 and Ptch1 in Lrig1 Control and Mutant hair bulb at P42. ( h ) Duplex RNA in-situ hybridisation of Scd1 and Gli1 in Lrig1 Control and Mutant hair bulb at P42. Purple dotted line denotes boundery of DP cells. Purple arrowhead indicates loss of Shh/Ptch1/Gli1 expression and area analysed (scale bar 20 μm). ( i ) Ptch1 and Gli1 transcripts by spot detection in DP of P42 Lrig1 Control and Mutant mice (n = 3 animals, 12 hair follicles analysed per animal). *p < 0.05 and **p < 0.001. Yellow solid line indicates area of representative image analysed for transcripts. ( j ) Duplex RNA in-situ hybridisation of Scd1 and Ptch1 in Lrig1 Control and Mutant hair bulb at P45. Purple arrowhead indicates loss of Ptch1 transcript expression. ( k ) Duplex RNA in-situ hybridisation of Scd1 and Gli1 in Lrig1 Control and Mutant hair bulb at P45. Purple arrowhead indicates loss of Gli1 transcript expression. ( l ) Ptch1 and Gli1 transcripts by spot detection in DP of P45 Lrig1 Control and Mutant mice (n = 3 animals, 12 hair follicles analysed per animal). *p < 0.05 and **p < 0.001. Yellow solid line indicates area of representative image analysed for transcripts. ( m ) Ptch1 and Gli1 transcripts by spot detection in Matrix of P45 Lrig1 Control and Mutant mice (n = 3 animals, 12 hair follicles analysed per animal). *p < 0.05 and **p < 0.001. Yellow solid line indicates area of representative image analysed for transcripts. Purple dotted line denotes boundary of DP cells (scale bar 20 μm).

Article Snippet: The following antibodies were used: Rat monoclonal anti-Ki67 (catalogue no. 14-5698-82, eBioscience), Rabbit polyclonal anti-mouse Scd1(M38) (catalogue no. #2438, Cell signalling), and Goat anti-Rabbit conjugated to Alexa Fluor A568 (catalogue no. A11036, Life Technologies).

Techniques: Expressing, In Situ, Hybridization, Mutagenesis

Journal: Cell Host & Microbe

Article Title: Aspergillus fumigatus hijacks human p11 to redirect fungal-containing phagosomes to non-degradative pathway

doi: 10.1016/j.chom.2023.02.002

Figure Lengend Snippet:

Article Snippet: Horse polyclonal anti-mouse IgG, HRP-linked antibody , Cell Signaling Technology , Cat# 7076; RRID: AB_330924.

Techniques: Recombinant, Cell Culture, Protease Inhibitor, Transfection, SYBR Green Assay, Mass Spectrometry, CyQUANT Assay, LDH Cytotoxicity Assay, Purification, Expressing, Construct, Transformation Assay, Clone Assay, Plasmid Preparation, Software, Imaging, Blocking Assay, Lysis

Journal: Cells

Article Title: Interstitial Macrophages Lead Early Stages of Bleomycin-Induced Lung Fibrosis and Induce Fibroblasts Activation

doi: 10.3390/cells12030402

Figure Lengend Snippet: Conditioned medium from BLM-sorted IM induce fibroblasts proliferation and differentiation into myofibroblasts. (A) Experimental design. After CD45 enrichment, IM were sorted from dissociated lung samples on day 7 after saline (IM untreated) or BLM (IM treated) administration and cultured for 24 h to collect the CM. MLg, a fibroblast cell line, was cultured with control medium, conditioned medium from untreated IM, or treated IM. After 48 h, the (B) metabolic activity was measured by MTS assay, (C) cell proliferation by Ki-67 staining, and (D) respective quantification was performed. Differentiation state was analyzed by (E) immunofluorescence with vimentin (red) and α-SMA (green), nuclei are stained with DAPI (blue), and (F) respective quantification, also by ( G ) Western blot, and ( H ) quantification of relative expression. Scale bar = 200 µm. Original magnification, 100×. Data are shown as the mean ± SEM of one representative experiment of three independent experiments. ** p < 0.01, *** p < 0.001: treated IM vs. treated AM.

Article Snippet: Cells were washed twice with PBS Tween 0.5% for 10 min, and blockage of non-specific binding was performed with newborn calf serum (proliferation protocol; 1/10—186427, Biochrom) or normal goat serum (transdifferentiation protocol; 1/10—G9023-0010, Sigma-Aldrich) for 1 h. Next, cells were incubated with a rabbit anti-mouse Ki-67 (1:250 in blocking solution—AB9260, Millipore), rabbit polyclonal anti-mouse vimentin (1:150 in blocking solution—#5741, CellSignalling Technology, Danvers, MA, USA), and mouse anti-mouse α-smooth muscle actin (SMA, 1:100 in blocking solution—ab32575, Abcam, Cambridge, UK) primary antibodies overnight.

Techniques: Cell Culture, Activity Assay, MTS Assay, Staining, Immunofluorescence, Western Blot, Expressing

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Detailed staining conditions and order of the Opal™-TSA mIHC 7-marker panel for human FFPE tissue sections

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Staining, Concentration Assay, Incubation

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Detailed staining conditions and order of the Opal™-TSA mIHC 6-marker panel for formalin free zinc salt-fixed tissue sections

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Staining, Incubation

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Multiplex manual staining of a representative human lymph node FFPE tissue section using the 7-marker TLS panel Representative region of interest (ROI) showing merge (on left) and single staining (on right): CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), DC-LAMP (orange), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Multiplex Assay, Staining, Marker

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Multiplex automated staining of a representative human lymph node FFPE tissue section using the 7-marker TLS panel Representative region of interest (ROI) showing merge (on left) and single staining (on right): CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), DC-LAMP (orange), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Multiplex Assay, Staining, Marker

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Proposed grading system for the accurate classification of immune cell infiltrates, immune cell aggregates, immature TLSs and mature TLSs using the 7-marker TLS panel on human prostate cancer FFPE tissue sections Representative ROIs showing merge staining: CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), DC-LAMP (orange), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Marker, Staining

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Multiplex manual staining of a Grade 2 immature TLS (upper panel) and a Grade 3 mature TLS (lower panel) in human melanoma tissue sections using the 7-marker TLS panel Representative ROIs showing merge (on left) and single staining (on right): CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), DC-LAMP (orange), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Multiplex Assay, Staining, Marker

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Multiplex automated staining of a Grade 2 immature TLS (upper panel) and a Grade 3 mature TLS (lower panel) in human prostate cancer tissue sections using the 7-marker TLS panel Representative ROIs showing merge (on left) and single staining (on right): CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), DC-LAMP (orange), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Multiplex Assay, Staining, Marker

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Proposed grading system for the accurate classification of immune cell infiltrates, immune cell aggregates, immature TLSs and mature TLSs using the 6-marker TLS panel on murine gastric cancer tissue sections Representative ROIs showing merge staining: CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), PNAd (cyan), and DAPI (blue). Images were taken at 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Marker, Staining

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet: Multiplex manual staining of a Grade 2 immature TLS (upper panel) and a Grade 3 mature TLS (lower panel) in murine gastric cancer tissue sections using the 6-marker TLS panel Representative ROIs showing merge (on left) and single staining (on right): CD4 (red), CD8 (magenta), CD19 (green), CD21 (yellow), PNAd (cyan), and DAPI (blue). Images were taken at a 20× magnification, and scale bars indicate 100 μm.

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Multiplex Assay, Staining, Marker

Journal: STAR Protocols

Article Title: Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

doi: 10.1016/j.xpro.2022.101961

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-mouse CD19 antibody , Cell Signaling Technology , Cat#3574 (1:200 dilution).

Techniques: Plasmid Preparation, Recombinant, Software, Staining, Imaging