Figure 1.
Figure 1. 3D collagen lattices of varying origin and cross-link status: Assembly speed, stiffness, and pore sizes at different polymerization conditions. (A) Assembly speed and (B) organization of acid-extracted pepsinized bovine dermal compared with acid-extracted rat tail collagen matrices after in vitro reconstitution (1.7 mg/ml). (A) Real-time confocal reflection microscopy of fibril formation, represented as kymograph (top panel; B, bovine dermal collagen; R, rat tail collagen) and time-dependent signal intensity curves normalized to the end-point of complete polymerization (bottom panel, horizontal solid line; n = 3 ± SD, shaded areas). Dashed lines, half-maximal polymerization after 8 min (bovine) and 30 s (rat tail, also see inset). (B) Collagen fibril diameter and network geometry, detected by transmission and scanning electron microscopy (TEM, SEM) of fixed dehydrated lattices and confocal reflection microscopy (horizontal, xy; vertical, xz) of native hydrated samples. Marked areas in xz represent signal-free regions referred to as pores. Brackets (TEM) indicate representative fibril calibers. (C) Determination of elastic modulus of bovine and rat tail collagen lattices by AFM. Left, force curves while probing bovine dermal or rat tail collagen lattices (1.7 mg/ml). Dotted curves, medians of all values fitted by the Hertz formula. Oscillatory example curves represent each one of 5–35 repeats. Vertical lines, deformation depth at 1,500 pN applied force. Right, stiffness for different collagen types and concentrations. Unless stated otherwise, polymerization temperature was 37°C. ***, P < 0.0001; **, P < 0.01; *, P < 0.05; n.s., not significant; data points represent 5–35 measurements from different locations as average of 2–20 repetitive tappings/position. Gray horizontal lines, median. (D–F) Pore cross sections quantified by confocal reflection xz sections of bovine dermal or rat tail collagen lattices of different polymerization conditions. Horizontal lines, medians. Data show 1–2 representative experiments from n = 3–4; 21 data points each. ***, P < 0.001; **, P < 0.01; *, P < 0.05; n.s., nonsignificant. (F) Right column, SEM from rat tail collagen lattices assembled at different temperatures. Bars: (A; B, confocal reflection) 10 µm; (B, F, SEM) 1 µm; (B, TEM) 0.1 µm.

3D collagen lattices of varying origin and cross-link status: Assembly speed, stiffness, and pore sizes at different polymerization conditions. (A) Assembly speed and (B) organization of acid-extracted pepsinized bovine dermal compared with acid-extracted rat tail collagen matrices after in vitro reconstitution (1.7 mg/ml). (A) Real-time confocal reflection microscopy of fibril formation, represented as kymograph (top panel; B, bovine dermal collagen; R, rat tail collagen) and time-dependent signal intensity curves normalized to the end-point of complete polymerization (bottom panel, horizontal solid line; n = 3 ± SD, shaded areas). Dashed lines, half-maximal polymerization after 8 min (bovine) and 30 s (rat tail, also see inset). (B) Collagen fibril diameter and network geometry, detected by transmission and scanning electron microscopy (TEM, SEM) of fixed dehydrated lattices and confocal reflection microscopy (horizontal, xy; vertical, xz) of native hydrated samples. Marked areas in xz represent signal-free regions referred to as pores. Brackets (TEM) indicate representative fibril calibers. (C) Determination of elastic modulus of bovine and rat tail collagen lattices by AFM. Left, force curves while probing bovine dermal or rat tail collagen lattices (1.7 mg/ml). Dotted curves, medians of all values fitted by the Hertz formula. Oscillatory example curves represent each one of 5–35 repeats. Vertical lines, deformation depth at 1,500 pN applied force. Right, stiffness for different collagen types and concentrations. Unless stated otherwise, polymerization temperature was 37°C. ***, P < 0.0001; **, P < 0.01; *, P < 0.05; n.s., not significant; data points represent 5–35 measurements from different locations as average of 2–20 repetitive tappings/position. Gray horizontal lines, median. (D–F) Pore cross sections quantified by confocal reflection xz sections of bovine dermal or rat tail collagen lattices of different polymerization conditions. Horizontal lines, medians. Data show 1–2 representative experiments from n = 3–4; 21 data points each. ***, P < 0.001; **, P < 0.01; *, P < 0.05; n.s., nonsignificant. (F) Right column, SEM from rat tail collagen lattices assembled at different temperatures. Bars: (A; B, confocal reflection) 10 µm; (B, F, SEM) 1 µm; (B, TEM) 0.1 µm.

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