Fourier analysis of electron micrographs of positively stained collagen fibrils: application to type I and II collagen typing.
Type I and II collagen (native-type) fibrils, positively stained with uranyl acetate, present typical periodic (D = 67 nm) cross-striation patterns. Although the two patterns are similar, the distributions of charged amino acids along the type I and II collagen molecules are different. After optical diffraction analysis or computer image processing of electron micrographs, different Fourier transforms were obtained from type I and II collagen fibrils, either as native fibrils or after in vitro reconstitution from purified molecules. With tissues such as tendon and cartilage, better results were obtained after mild trypsin treatment, which allowed better isolation and staining of the collagen fibrils. The main difference observed in the Fourier transforms was the presence in type II collagen fibrils of a strong tenth-order peak (corresponding to the tenth harmonic of the fundamental frequency). In order to discriminate between the two collagens, we measured the ratio (R) of the areas under the ninth- and tenth-order peaks. In trypsin treated tissues, the distributions of these ratios were clearly separated: below 1.0 for type II collagen fibrils and above 1.5 for type I collagen fibrils. This method appears to be suitable for rapid typing of type I and II collagen fibrils and might be useful for determining the exact composition of fibrils in tissues, such as intervertebral discs, that contain these both types of collagen.
Institut de Biologie et Chimie des Protéines, UPR CNRS 412, Lyon, France.
M C
MC
Ronzière
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B
Herbage
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Herbage
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JC
Bernengo
D000818:Animals; D002358:Cartilage, Articular / Q000737:chemistry; D002417:Cattle; D003094:Collagen / Q000032:analysis* / Q000737:chemistry / Q000648:ultrastructure*; D005583:Fourier Analysis; D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron; D055095:Optics and Photonics; D009942:Organometallic Compounds / Q000737:chemistry; D051381:Rats; D012867:Skin / Q000737:chemistry; D013194:Staining and Labeling / Q000379:methods*; D013710:Tendons / Q000737:chemistry
D009942:Organometallic Compounds; C005460:uranyl acetate; D003094:Collagen