Sepúlveda P, Murgui A, López-Ribot J L, Casanova M, Timoneda J, Martínez J P
Departamento de Microbiología, Facultad de Farmacia, Universitat de València, Spain.
Infect Immun. 1995 Jun;63(6):2173-9. doi: 10.1128/iai.63.6.2173-2179.1995.
Rabbit polyclonal antibodies (PAbs) directed towards the amino-terminal cysteine-rich 7S domain (PAb anti-7S), the major internal collagenous domain (PAb anti-type IV), and the C-terminal noncollagenous region (PAb anti-NC1) of the type IV collagen molecule were probed by indirect immunofluorescence against Candida albicans blastoconidia and germinated blastoconidia. Most nongerminating cells and mother blastoconidia from which germ tubes originated showed strong fluorescence when PAb anti-7S was used, whereas with PAb anti-type IV, fluorescence was found almost exclusively on the surface of filamentous forms. A patched fluorescent pattern rather than a homogenous confluent fluorescence was observed in all cases. No fluorescent cells were observed with PAb anti-NC1. By Western immunoblotting, PAb anti-type IV cross-reacted primarily with a polypeptide of 37 kDa present in wall extracts obtained from intact cells of both growth forms by treatment with beta-mercaptoethanol, whereas PAb anti-7S recognized a major 58-kDa antigen also present in both extracts, along with some other high-molecular-mass (> 106-kDa) polydisperse species present only in the material released from blastoconidia with beta-mercaptoethanol. No reactive bands were observed when PAb anti-NC1 was used as a probe in Western immunoblotting experiments. The sensitivities or resistances to collagenase digestion of the different polypeptides that cross-reacted with PAbs anti-type IV and anti-7S suggest the existence of cell wall components in C. albicans that contain epitopes that mimic the collagenous domains of the type IV collagen molecule.
针对IV型胶原分子的氨基末端富含半胱氨酸的7S结构域(抗7S多克隆抗体)、主要的内部胶原结构域(抗IV型多克隆抗体)和C末端非胶原区域(抗NC1多克隆抗体)的兔多克隆抗体,通过间接免疫荧光法检测白色念珠菌芽生孢子和发芽芽生孢子。当使用抗7S多克隆抗体时,大多数未发芽细胞和长出芽管的母芽生孢子显示出强荧光,而使用抗IV型多克隆抗体时,荧光几乎仅在丝状形式的表面发现。在所有情况下均观察到斑点状荧光模式而非均匀融合荧光。使用抗NC1多克隆抗体未观察到荧光细胞。通过蛋白质免疫印迹法,抗IV型多克隆抗体主要与一种37 kDa的多肽发生交叉反应,该多肽存在于通过β-巯基乙醇处理从两种生长形式的完整细胞获得的细胞壁提取物中,而抗7S多克隆抗体识别出两种提取物中均存在的一种主要的58 kDa抗原,以及仅存在于用β-巯基乙醇从芽生孢子释放的物质中的一些其他高分子质量(>106 kDa)的多分散物种。当在蛋白质免疫印迹实验中使用抗NC1多克隆抗体作为探针时未观察到反应条带。与抗IV型和抗7S多克隆抗体发生交叉反应的不同多肽对胶原酶消化的敏感性或抗性表明,白色念珠菌细胞壁成分中存在模拟IV型胶原分子胶原结构域表位的表位。