Reagents

Extracted and purified type I collagen is available from biochemical suppliers.

20 40 60

Incubation time (min)

20 40 60

Incubation time (min)

Figure 6.45 Reconstitution of type I collagen fibrils at 34 °C, I 0.2, pH 7.4. (a) Different initiation routes are possible in changing from cold acid to warm neutral conditions and this can affect both the assembly pathway and kinetics of fibril formation. (b) Typical turbidimetric curve (obtained at 313 nm) after 'warm start' initiation with I 0.2, pH 7.4 phosphate buffer at 34 °C. The turbidity is proportional to the extent of fibril assembly. (c) Typical 'early fibril' with tapered tips, found throughout the lag and early growth phases of reconstitution (scale bar = 300 nm). (d) Typical final gel of D-periodic fibrils (D = 65 nm in the dried and negatively stained preparation) with no fibril tips apparent (scale bar = 300 nm)

Figure 6.45 Reconstitution of type I collagen fibrils at 34 °C, I 0.2, pH 7.4. (a) Different initiation routes are possible in changing from cold acid to warm neutral conditions and this can affect both the assembly pathway and kinetics of fibril formation. (b) Typical turbidimetric curve (obtained at 313 nm) after 'warm start' initiation with I 0.2, pH 7.4 phosphate buffer at 34 °C. The turbidity is proportional to the extent of fibril assembly. (c) Typical 'early fibril' with tapered tips, found throughout the lag and early growth phases of reconstitution (scale bar = 300 nm). (d) Typical final gel of D-periodic fibrils (D = 65 nm in the dried and negatively stained preparation) with no fibril tips apparent (scale bar = 300 nm)

Suitable acetic-acid-soluble type I collagen can alternatively be extracted and purified by well-established procedures [3] with modifications to ensure minimal loss of the telopeptides [4]. Laboratory reagents should be of high purity.

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