human collagen type i/iii  (Advanced Biomatrix Inc)

Journal: The Journal of pathology

Article Title: A 3D tri-culture system reveals that activin receptor-like kinase 5 and connective tissue growth factor drive human glomerulosclerosis

doi: 10.1002/path.4960

Figure Lengend Snippet: (A) MC 3D monoculture without the addition of TGFβ, and (B) in the presence of TGFβ, shown by H&E staining of paraffin-embedded sections. Human collagen type I/III (immunostained with an antibody that recognises both collagen type I and III) (C) and human collagen type IV (D) can be demonstrated in nodules by immunoperoxidase staining. (E) High-power view of MC nodule by scanning electron microscopy in cross section. (F) MC nodule count and collagen type I alpha1 (COL1a1), and collagen type IV alpha1 (COL4a1) RNA quantification (n=3). (G) Effect of ALK5 inhibition (Alk5i) on MC nodule formation (n=3). (H) Effect of SMAD2 or SMAD3 siRNA knockdown in MCs on nodule formation (n=3) with immunoblots demonstrating degree of knockdown. [DharmaFECT (DH), non-targeting siRNA control (siCP), siSMAD2 (siS2), siSMAD3 (siS3), total SMAD2 (tSMAD2), total SMAD3 (tSMAD3)]. Error bars represent 1 s.d. NS, not significant; ***, p<0.001; **, p<0.01: 2-tailed Student’s t test.

Article Snippet: Staining of human collagen in MC 3D cultures After culture, matrices were embedded in HistoGel, fixed in 4% paraformaldehyde and paraffin wax-embedded for sectioning and staining for human collagen type IV (mouse anti-human collagen IV monoclonal antibody 2150–0121, Bio-Rad, CA, USA) and human collagen type I/III (rabbit anti-human collagen I/III polyclonal antibody 2150–2210, Bio-Rad, CA, USA).

Techniques: Staining, Immunoperoxidase Staining, Electron Microscopy, Inhibition, Western Blot

Journal: Wound Repair and Regeneration

Article Title: The altered mechanical phenotype of fetal fibroblasts hinders myofibroblast differentiation

doi: 10.1111/wrr.12677

Figure Lengend Snippet: Fibronectin induces more focal adhesions than types I and III collagen. (A) Type I and (B) type III collagen were incorporated into rigid PAAs yielding uniform coatings on the substrate surfaces as identified by immunostaining with representative wide‐field immunofluorescence images shown. Quantitation of (C) the average fluorescence intensities and (D) their standard deviations of types I and III collagen at the surfaces of rigid PAAs. (E) The elastic moduli of the rigid PAAs with types I and III collagen were similar to those with fibronectin and calculated based on measurements of the shear elastic moduli (G′) using rheometry at a constant strain of 0.05% and frequency of 1 Hz assuming E = 3G′ and a Poisson's ratio of 0.5. Quantitation of the (F) number of total focal adhesions per cell which were divided into (G) immature (<2 μm 2 ) and (H) mature (>2 μm 2 ) from vinculin immunostaining of adult and fetal fibroblasts on rigid PAAs with fibronectin, type I collagen, and type III collagen. (I) Quantitation of α‐SMA average pixel intensities from α‐SMA immunostaining of adult and fetal fibroblasts on rigid PAAs with fibronectin, type I collagen, and type III collagen. All data are presented as box and whisker plots with the black lines indicating the medians, the whiskers representing the 10th and 90th percentiles, and * indicating p < 0.05 for n = 60 areas from three independent experiments for (C) and (D), n = 7–8 gels from two independent experiments for (E), n = 22–26 cells from three independent experiments for (F), (G), and (H), and n = 30 cells from three independent experiments for (I) using lots 12,925 and 10,111. Scale bar represents 40 μm.

Article Snippet: Human types I and III collagen were purchased as sterile solutions (Advanced Biomatrix, Carlsbad, CA).

Techniques: Immunostaining, Immunofluorescence, Quantitation Assay, Fluorescence, Shear, Whisker Assay