Journal: Communications Biology

Article Title: CRISPR-clear imaging of melanin-rich B16-derived solid tumors

doi: 10.1038/s42003-023-04614-7

Figure Lengend Snippet: a Comparison of antibody labeling of intact melanin (+/–) tumors. HER2 is detected with an anti-HER2 antibody and stained with an Alexa-488 labeled secondary antibody (shown in white), tdTomato-expressing B16 cells (shown in purple), imaged at increasing depths. Images were acquired using a 25x objective. Scale bar 150 μm (yellow). b Overview of workflow for 3D imaging, localization and visualization of nanoparticles in melanin(–) tumors. The method consists of 2 modular steps: (1) multi-dye vessel-staining (highlighted in Fig. ) and injection of nanoparticles in tissue, and (2) clearing for high imaging quality using 3D confocal microscopy. For clarity, separate channels are shown for nanoparticles (Alexa 488), tdTomato-expressing melanin(–) B16 melanoma cells and vasculature (Alexa 647). Scale bar 300 μm. A 10x/0.45 objective was used for imaging. c 3D visualization of digital reconstruction of nanoparticles at 500 µm depth of melanin(–) tumors cleared with PACT . Purple is vasculature, cyan are reconstructed tdTomato-expressing melanin(–) B16 melanoma cells, and green represents nanoparticles. Scale bar 300 μm. White box insert on the left is a blow-up of the box on the right. Scale bar 20 μm for insert. A 10x/0.45 objective was used for imaging.

Article Snippet: The primary antibodies used for passive staining were rabbit anti-HER2 IgG (Polyclonal Antibody, ThermoFisher PA5-14635).

Techniques: Antibody Labeling, Staining, Labeling, Expressing, Imaging, Injection, Confocal Microscopy

Journal: The Journal of investigative dermatology

Article Title: EGFR Signaling Is Overactive in Pachyonychia Congenita: Effective Treatment with Oral Erlotinib.

doi: 10.1016/j.jid.2022.08.045

Figure Lengend Snippet: Figure 2. EGFR signaling is overactive in suprabasal layers of PC-lesional skin. (a‒c) Representative immunostaining for (a) HER1‒EGFR, (b) HER2, (c) and phospho-RPS6 (a biomarker of mTOR activation), and phospho-ERK in sections from PC-lesional skin (plantar calluses), PC-non lesional skin, and healthy control. Bars ¼ 50 mm in panel. (d) Representative in situ RNA hybridization of VEGF-A in PC-lesional skin and PC-nonlesional skin. Cell nuclei are stained in blue, and target mRNAs are stained in red. Bars ¼ 250 mm (50 mm in insets). (e) Quantification of the average number of VEGF-A probe signals/area detected in

Article Snippet: List of Antibodies and Probes Used Antigen Supplier Reference Isotype EGFR‒HER1 Cell Signaling Technology #4267 rabbit IgG HER2 Cell Signaling Technology #4290 rabbit IgG Phosphorylated p44/42 MAPK (Thr202/Tyr204) Cell Signaling Technology #4376 rabbit IgG phospho-RPS6 (Ser240/Ser244) Cell Signaling Technology #5364 rabbit IgG FLG Invitrogen #MA5-13440 mouse IgG1 Keratin 6 Abcam #ab18586 Mouse IgG1 Keratin 10 Abcam #ab20208 mouse IgG1 Ki-67 Leica #RTU-Ki67-MM1 Mouse IgG1 TRPV3 mRNA Bio-Techne #573471 Epiregulin mRNA Bio-Techne #313081 TGFa mRNA Bio-Techne #313131 VEGFa mRNA Bio-Techne #423161 AREG mRNA Bio-Techne #313111 HB-EGF mRNA Bio-Techne #524821 Abbreviations: #, number; phospho-RPS6, phosphorylated RPS6.

Techniques: Immunostaining, Biomarker Discovery, Activation Assay, Control, In Situ, Hybridization, Staining