rabbit anti desmoglein 2  (Bethyl)

Journal: Frontiers in Immunology

Article Title: Role of Dsg1- and Dsg3-Mediated Signaling in Pemphigus Autoantibody-Induced Loss of Keratinocyte Cohesion

doi: 10.3389/fimmu.2019.01128

Figure Lengend Snippet: Genetic deletion of Dsg3 and Dsg2 using CRISPR/Cas9 in HaCaT keratinocytes. (A) Schematic of sequencing results after inducing a DSB with NHEJ repair in Exon 5 of either Dsg3 or Dsg2 using CRISPR/Cas9. (B) Cell line characterization by immunostaining of desmosomal proteins ( n = 3). (C) Cell line characterization by immunoblot with representative images on the left and densitometric quantification on the right ( n = 3, one-way ANOVA, * p ≤ 0.05 vs. WT). (D) Dispase assay of WT and Dsg-deficient cell lines under baseline treatment. ( n = 7, one-way ANOVA, * p ≤ 0.05 vs. WT) (E) Cell lines incubated with IgG fractions for 24 h and subjected to Dispase assays ( n = 5, two-way ANOVA, # p ≤ 0.05 vs. respective c-IgG, * p ≤ 0.05 vs. WT).

Article Snippet: Membranes were blocked by 5% bovine serum albumin (BSA) in Tris-buffered saline with 0.05% Tween (TBS-T) for 1 h at room temperature and following primary antibodies were used overnight at 4°C in blocking solution: phospho-EGF Receptor mAb (Tyr845) (Cell Signaling Technologies, Danvers, USA), EGF Receptor mAb (CST), phospho-p44/42 MAPK mAb (CST), p44/42 MAPK mAb (CST), GAPDH (Santa Cruz, Heidelberg, Germany), Desmoplakin I/II (H-300) (Santa Cruz), α-Tubulin (Abcam, Cambridge, UK), Dsg3 pAb (Biozol, Eching, Germany), PG (Progen, Heidelberg, Germany), E-Cad (BD Transduction), Dsg2 (OriGene, Herford, Germany), Desmocollin (Dsc) 3 (Progen).

Techniques: CRISPR, Sequencing, Immunostaining, Western Blot, Incubation

Journal: Frontiers in Immunology

Article Title: Role of Dsg1- and Dsg3-Mediated Signaling in Pemphigus Autoantibody-Induced Loss of Keratinocyte Cohesion

doi: 10.3389/fimmu.2019.01128

Figure Lengend Snippet: Signaling pathway modulation by pemphigus IgG fractions in Dsg-deficient cell lines. Analysis of ERK activation after 30 min application of IgG fractions by immunoblot in Dsg3- and Dsg2-deficient cell lines. (A) Representative immunoblot and (B) densitometric analysis ( n = 9, two-way ANOVA, * p ≤ 0.05). (C) Representativ graph of Fura-2 ratiometric Ca 2+ imaging reveals a PF-IgG mediated Ca 2+ influx in all cell lines used ( n = 3).

Article Snippet: Membranes were blocked by 5% bovine serum albumin (BSA) in Tris-buffered saline with 0.05% Tween (TBS-T) for 1 h at room temperature and following primary antibodies were used overnight at 4°C in blocking solution: phospho-EGF Receptor mAb (Tyr845) (Cell Signaling Technologies, Danvers, USA), EGF Receptor mAb (CST), phospho-p44/42 MAPK mAb (CST), p44/42 MAPK mAb (CST), GAPDH (Santa Cruz, Heidelberg, Germany), Desmoplakin I/II (H-300) (Santa Cruz), α-Tubulin (Abcam, Cambridge, UK), Dsg3 pAb (Biozol, Eching, Germany), PG (Progen, Heidelberg, Germany), E-Cad (BD Transduction), Dsg2 (OriGene, Herford, Germany), Desmocollin (Dsc) 3 (Progen).

Techniques: Activation Assay, Western Blot, Imaging